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PHYLOGENETICS OF TRIGYNASPIDA (ACARI: MESOSTIGMATA)
DISSERTATION
Presented in Partial Fulfillment of the Requirements for
the Degree of Doctor of Philosophy in the Graduate
School of The Ohio State University
By
Cheol-Min Kim, B.S., M.S.
The Ohio State University 2001
Dissertation Committee: Approved by John W. Wenzel, Co-Adviser [xj.
Hans Klompen, Co-Adviser
David J. Horn / / Â d v i s ^
Norman F. Johnson Department of Entomology UMI Number 3031216
UMI’
UMI Microform 3031216 Copyright 2002 by Bell & Howell Information and Leaming Company. All rights reserved. This microform edition is protected against unauthorized copying under Title 17, United States Code.
Bell & Howell Information and Leaming Company 300 North Zeeb Road P.O. Box 1346 Ann Arbor. Ml 48106-1346 Copyright by Cheol-Min Kim 2001 ABSTRACT
A generally accepted viewpoint on the superfamilial and familial level evolutionary relationships and the classification among the members of the Trigynaspida (Arachnida: Acari: Mesostigmata) has been reassessed and a new paradigm has been proposed based on the results from the study of both morphology and DNA sequences.
Morphological study includes 51 taxa and 55 characters, and molecular study includes NS5-NS8 region of 18S (18 taxa, 714 characters) and V-X region of 285 (21 taxa, 738 characters) nuclear ribosomal DNA (rONA) genes. Combined analyses of these data sets (51 taxa, 1507 characters), along with separate analyses of each data set, were also performed. The principle of maximum parsimony was applied to all the analyses as the optimality criterion.
The results from morphological, molecular, and combined study supported bifurcating lineages of Cercomegistina and Antennophorina within the monophyletic Trigynaspida. However, currently accepted superfamily of Fedrizzioidea, which contains four families that are associated with passalids (Fedrizziidae and Klinckowstroemiidae), carabids (Promegistidae), and diplopods and squamates (Paramegistidae), is separated into three groups. While Fedrizziidae and Klinckowstroemiidae remain in the current superfamily of Fedrizzioidea, Promegistidae move to Parantennuloidea that displays the association with carabids and ii tenebrionids, and the remaining Parame^stidae stand alone as 3l new superfamily Paramegistoidea.
The phylogenetic position of Parantennuloidea was basal within the Antennophorina in morphological analyses but was sister to Paramegistoidea in both molecular and combined analyses. Two strict ant associates, Antennophoroidea and Aenictequoidea, maintained the sister group relationships in all the analyses.
This new paradigm is also strongly supported by the information of hosts that are associated with these mites, in which it implies that the wide spread passalid-association was initiated in the early evolution of these mites. The information of the plate tectonics and the global biogeographic patterns among the Trigynaspida and its hosts imply the fact that the origin of Trigynaspida is linked, at the latest, to the early Mesozoic (Upper Triassic).
Ill Nature created species; Man created genera.
IV ACKNOWLEDGMENTS
There would be nobody who raises the question to argue with the fact that this study was conducted in the dark age of my life. Despite this, this humble piece of work is now being shown up to the world thanks to the invaluable support and assistance from my family, colleagues, friends, and open-minded nameless helpers from all around the world. Without them, this work would have hardly been performed in its present form.
In losing my tenure, I thank my adviser and committee members, John Wenzel, Dave Horn, Norm Johnson, and Hans Klompen.
Bev 'Wally' Gerdeman not only offered critical materials for the study but very well digested my humor to the world that are often cynical, skeptical, and nihilistic. Thank you, Bev.
Dave Walter often gave me the momentum to move forward with this project. I am especially deeply gratitude for his open-heartedness, generosity, and incisive criticism on this research. I am sure that his letter of support allowed me to get my travel grant, which allowed me to perform field work in Australia.
It was my own pleasure to be acquainted with Owen Seeman, who not only generously offered some of his collected materials but offered guided field trips along with the information from his specialty. It will always be nice to go into the rainforest with Owen again sometime in the future, I hope.
Barry OConnor offered 2 individuals of Antennophorus, a very critical taxon for this study, along with the arrangement of loans of Pyrosejidae and many Costa Rican materials. I just did not have the guts to use only one of them for the DNA study, leaving the voucher specimen of Antennophorus at the UMMZ! Evert Lindquist also kindly offered fresh Costa Rican materials. The next step after this work should be conducting the remaining taxonomic works and to return the borrowed materials to them as quickly as possible. John Moser sent me lots of articles and materials on bark beetle associates. Eddie Ueckermann offered Davacaridae from the Antarctic Marion Islands. Sabina Swift surveyed the reptile collection in Bernice P. Bishop Museum to find Ophiomegistus soaked in formalin for me.
Marisa Castagnoli kindly supplied the priceless photographs of Antennomegistus caputcarabi in the Berlese Collection, which allowed me to write a key to the genera of Paramegistidae. Jerry Krantz offered me the permission to use the illustrations in his
1 have to express my sincere thanks to the late Professor Donald Earl Johnston (1934-1994), a mentor, a scholar, and a truly gifted teacher. Although it was somewhat a short period of time, I was just lucky enough to cross his path.
This study was supported in part by the Korean Honor Scholarship, Graduate Student International Research Travel Grant, Graduate Student Alumni Research Award, and International Student Grant.
The efforts of Katherine Harris and the Suprêmes allowed me to have sporadic intermissions while having somewhat boring time of writing the final draft. I can say to my fellows of America that, however, I did not have any relationship with that woman, and never will. An opportunity to honor the distillers and brewers of those cheap Scotches and Budweisers, which are thought to be useful for the destruction of those obnoxious prions in the brain, should not be ignored. Uncounted hundreds of those beautiful trigynaspids that have been killed for this ruthless work also well-deserve the honor and gratitude. Thank you all. I am a proud Gaean. C-M.
Vll VITA
January 11,1965 ...... Bom - Seoul, Korea
1983-1987 ...... B.S., Seoul National University
1987-1989 ...... M.S., Seoul National University
PUBLICATION
Kim, C-M. 1990. A new species of eriophyid mite, Aceria zelkoviana sp. n. (Acari: Eriophyidae) associated with Zelkova serrata Makino (Ulmaceae) from Korea. Internat. J. Acarol. 16(2):85-87.
V lll HONORS AND AWARDS
Korean Honor Scholarship The Korean Honor Scholarship Committee, Washington, D C., 1996
Graduate Student International Research Travel Grant The Ohio State University, Columbus, OH, 1998
Graduate Student Alumni Research Award The Ohio State University, Columbus, OH, 1998
International Student Grant The Ohio State University, Columbus, OH, 2000
FIELD OF STUDY
Entomology
IX TABLE OF CONTENTS
Abstract ...... ii
Acknowledgments ...... v
V ita...... viii
List of Tables ...... xii
List of Figures ...... xiv
Chapters:
1. General Introduction ...... 1
2. A new genus and species of Paramegistidae ...... 15 Introduction ...... 15 Materials and methods ...... 18 Description ...... 19 Discussion ...... 24 A key to the genera of the family Paramegistidae (based on female)... 26
3. Morphological Analyses ...... 35 Introduction ...... 35 Materials and methods ...... 39 Bauplan of Trigynaspida ...... 44 Character selection, description, and coding ...... 58 Results and discussion ...... 90 A note on Paramegistus australis Banks, 1916 ...... 94 A key to the families of Trigynaspida (based on female) ...... 95 Diagnoses of the superfamihes of Trigynaspida ...... 106 4. Molecular A nalyses ...... 110 Introduction ...... 110 Materials and methods ...... 112 Results and discussion ...... 120
5. Combined Analyses ...... 127 Introduction ...... 127 Materials and methods ...... 130 Results and discussion ...... 135
Appendix A. Aligned NS5-NS8 region of the 18S rRNA sequences ...... 148 Appendix B. Aligned V-X region of the 28S rRNA sequences ...... 156
References ...... 164
XI LIST OF TABLES
Table Bags
2.1 Biogeography, host association, and numbers of described species of Paramegistidae. {* = new records) ...... 16
2.2 Leg chaetotaxy of Meristomegistus vazquezus n. sp ...... 22
3.1 List of taxa used for the morphological study. S.F.: Superfamily ...... 42
3.2 Taxon-Character matrix used for morphological study. ?: not available ...... 89
3.3 Proposed classification for Fedrizzioidea ...... 92
4.1 List of taxa used for molecular study. S.F. = Superfamily ...... 113
4.2 Sequences of primers used for the amplification of 185 (NS5 and NS8; from Black et ai, 1997) and 28S (V and X; from Hillis and Dixon, 1991) nuclear rONA ...... 116
4.3 Tree statistics ...... 121
5.1 List of taxa used for both of morphological and molecular study. S.F. = Superfamily; M = Morphological study; 185, 285 = Molecular study ...... 132 xii 5.2 Test for compatibility among the partitioned data sets, ns: not significant ...... 136
5.3 Proposed classification for currently known Fedrizzioidea. This proposal is corroborated by the morphological, molecular, and combined analyses ...... 144
X lll LIST OF FIGURES
Figure EagS
1.1 Schematic diagram of the chaetotaxy of femur IV of (a) Trigynaspida and Diarthrophallina, and (b) Sejina. Open circles represent for the dorsal setae and the filled circles represent for the ventral setae ...... 3
1.2 Phylogenetic tree re-drawn from Kethley (1977b). Note that two ant associates of Antennophoroidea and Aenictequoidea are not sisters to each other. Paramegistidae, Celaenopsidae, and Schizogyniidae are not monophyletic ...... 10
2.1 Meristomegistus vazquezus n. sp., female, idiosoma, dorsum ...... 27
2.2 Meristomegistus vazquezus n. sp., female, idiosoma, venter ...... 28
2.3 Meristomegistus vazquezus n. sp., female, gnathosoma. a: Gnathotectum; b: Ventral view (palp tarsal setae are eliminated); c: Palp tibia and tarsus, dorsal view; d: Chelicera ...... 29
2.4 Meristomegistus vazquezus n. sp., female, leg I, dorsal view, a: Overview; b: Tarsus I; c: Tarsus I, distal blunt-end sensilla ...... 30
2.5 Meristomegistus vazquezus n. sp., female, legs U-IV, dorsal view, a: Leg H; b: Leg HI; c: Leg I V ...... 31
2.6 Meristomegistus vazquezus n. sp., male, idiosoma, dorsum ...... 32 xiv 2.7 Meristomegistus vazquezus n. sp., male, idiosoma, venter ...... 33
2.8 Meristomegistus vazquezus n. sp., male, gnathosoma. a: Gnathotectum; b: Ventral view ...... 34
3.1 Fedrizzia sp. (Fedrizziidae). Note entire prestemal, bearing stl and stpl, and stemogynial shield, bearing stp3. (modified from Johnston, 1968) ...... 46
3.2 (a) Rhodacarus roseus Oudemans (Rhodacaridae) and (b) Gamasellus vibrissatus Emberson (Gamasellidae), showing prestemal platelets and postanal seta (pon). Note the presence of 3 pairs of stp and 4 pairs of St on sternal shield. Prestemal and metastemal shields are absent, (modified from Johnston, 1968) ...... 47
3.3 Celaenopsis badius (C. L. Koch) (Celaenopsidae), showing paired metastemal (MT) and ventromarginal (VTM) shields. Postanal seta is absent in all the postlarval Trigynaspida. (modified from Johnston, 1968) ...... 48
3.4 Pergamasus crassipes (Linnaeus) (Parasitidae), showing prestemal platelets, metastemal shields (MT), and postanal seta (pon). For the difference between latigynial and metastemal shields, see the text, (modified from Johnston, 1968) ...... 49
3.5 Promegistus armstrongi Womersley (Promegistidae), showing pseudostemogynium (PSG; = pseudostemum). Each metastemal shield (MT) is fused to the endopodal element ...... 50
3.6 Antennoseius delicatus Berlese (Ascidae). st4 are on the soft cuticle. (modified from Johnston, 1968) ...... 52
3.7 Dinychus sp. (Uropodidae), showing 'hologynaspid' genital system. (after Krantz, 1978) ...... 54
XV 3.8 Cercoleipus sp. (Cercomegistidae). Gnathotectum ...... 62
3.9 Seiodes ursinus Berlese (Seiodidae), (a) female and (b) male gnathotectum ...... 62
3.10 (a) Long and slender chelicera of Uropodoidea. Cheliceral excrescences are absent, (b) Short and robust chelicera of Celaenopsoidea. Excrescences are dendritic, (after Krantz, 1978) ...... 64
3.11 Klinckowstroemia sp. (Klinckowstroemiidae). (after Krantz, 1978) ...... 65
3.12 Comiculi. (a) Fedrizzia sp. (Fedrizziidae); (b) Paramegistus confrater Trâgârdh (Paramegistidae); (c) Echinomegistus wheeleri (Wasmann) (Paramegistidae); (d) Antennophorus sp. (Antennophoridae); (e) Euzercon latus (Banks) (Euzerconidae) ...... 67
3.13 (a) weakly fused (Paramegistus) and (b) completely fused (Ophiomegistus) palp tibia and tarsus ...... 69
3.14 Megacelaenopsis sp. (Megacelaenopsidae). Palp tarsus with 3-tined apotele ...... 70
3.15 Cercoleipus sp. (Cercomegistidae). Fused tritostemal laciniae ...... 71
3.16 Ptochacarus silvestrii Womersley (Ptochacaridae), showing paired stemovaginal sclerites ...... 73
3.17 Choriarchus reginus Kinn (Schizogyniidae), showing fused metastemal shield (MT). Heads of vaginal sclerites are well-developed ...... 76
3.18 Seiodes ursinus Berlese (Seiodidae), female sternal-genital region 78
xvi 3.19 Paramegistus confrater Trâgârdh (Paramegistidae). Weakly-developed (or vestigial) vaginal sclerites are shown in dotted line. Stemogynial shields are paired ...... 79
3.20 Ptochacarus sp. (Ptochacaridae), male stemogenital region showing paired eugenital setae ...... 87
3.21 Strict consensus tree from 3 equally most parsimonious trees for comparative morphological data under no topological constraints. Numbers shown are the values of branch support. Names of the superfamilies are from this study ...... 91
4.1 Strict consensus of 3 equally parsimonious trees deduced from NS5- NS8 region of 185 rRNA sequences under no topological constraints. Gaps were treated as missing. Numbers on the branches indicate branch support sensu Bremer (1988) ...... 122
4.2 Strict consensus of 9 equally parsimonious trees inferred from the V-X region of 285 rRNA sequences under no topological constraints. Gaps were treated as missing. Numbers on the branches indicate branch support sensu Bremer (1988) ...... 123
4.3 5trict consensus of 3 equally parsimonious trees deduced from 185+285 sequence data under no topological constraints. Gaps were treated as missing. Numbers on the branches indicate branch support sensu Bremer (1988) ...... 124
4.4 Sequence alignment of 77 and matching 77' regions of 28S rRNA, showing compensatory base pair changes. The first position (base) of 5'- of 77 stem matches to the last position (base) of 3 - in 77' stem. Note that the group Trigynaspida is uniquely distinguished from the outgroups. The region between 77 and 77' is omitted. Brackets are used to define the stem region ...... 126
xvii 5.1 Strict consensus tree ûront ISd equally most parsimonious trees with no topological constraints. Numbers shown are the values of branch support ...... 137
5.2 Strict consensus tree from combined morphology and molecules with host information ...... 143
xviu CHAPTER 1
GENERAL INTRODUCTION
^Æites (Subclass Acari) represent the most diverse group of arachnids
{i.e., spiders, scorpions, daddy-long-legs, pseudoscorpions, mites, ticks, etc.) and are second only to the insects in terms of the number of species among animals (Johnston, 1982; Lindquist, 2000; Wheeler, 1990). They are critical ecological elements in the process of succession in the ecosystem as well as important pests of agriculture, and human and animal hygiene. Among the groups of mites, the Order Mesostigmata, which occupies diverse habitats in the ecosystem, is often divided into two subgroups, known as Trigynaspida and Monogynaspida. Among these, the Trigynaspida, first grouped and named by Camin and Gorirossi (1955), is composed of 273 nominal species in 97 genera in 27 families. This grouping and the name of the group are based primarily on morphological features of genital area of the females, in which they usually have three-pieced genital ('tri-gynaspid') shields, whereas the members of the other group of Mesostigmata {i.e., Monogynaspida) have usually a single or sometimes no genital shield in the venter of the females. The typical 'trigynaspid' genital system is composed of a mesogynial shield flanked by two latigynial shields, and the genital orifice, made by these shields, becomes an ovipore of the female. In monogynaspid mites, trigynaspids' two latigynial shields are thought to be fused with endopodal elements or with mesogynial (epi^mialX shield^ and the remained mesogynial shield alone becomes often sac- or tongue-shaped 'epigynial shield'.
This 'trigynaspid' genital system, however, is not unique to the members of Trigynaspida as some early derivative Uropodina (e.g., Trachytes) also show 'trigynaspid-like' genital structure (see Chapter 3). On the contrary, in some trigynaspids, such as Celaenopsoidea, characteristic latigynial or mesogynial shields (or both) are often fused to the ventral shield elements, showing a more or less monogynaspid (or agynaspid) condition. Accordingly, more detailed and generalized characters that are not restricted to the female genital shields are required to define the group Trigynaspida.
Diagnosis of Trigynaspida
In general, although the Trigynaspida can be identified prima facie by its female genital structures, with the unique exception of the paedomorphic millipede-associate Neotenogynium malkini Kethley, the group Trigynaspida is diagnosed by retaining the combination of the following characters in the adults:
1. Presence of 8 setae on femur IV (shared with Diarthrophallina) 2. Presence of 'trigynaspid' and its derivative genital structures in female (shared with some Uropodina) 3. Presence of the setae av4 and pv4 on tarsus IV (shared with Sejina) 4. Absence of postanal seta (shared with Diarthrophallina and some Uropodina). 5. Presence of 4 anterolateral setae (al) on tarsi 11-lV Tri^naspida, along with Diarthrophallina sensu Trâgârdh (1946b), retains 8 setae on femur IV by having a posterolateral seta (pi), which lead to the chaetotactic formula of (1,2/1, 2/1,1). This simple character, which can be applied to both male and female, is compared with that of Sejina (including Sejus, Epicroseius, Uropodella) and Uropodina, in which they retain only 7 setae on femur IV by lacking pi (1,2/1, 2/1,0) (Figure 1.1).
si • O " avi pvl
(a) (b)
Figure 1.1: Schematic diagram of the chaetotaxy of femur IV of (a) Trigynaspida and Diarthrophallina, and (b) Sejina. Open circles represent for the dorsal setae and the filled circles represent for the ventral setae.
Trigynaspids also share the ventral setae av4 and pv4 on tarsus IV (Figure 2.5, c) with Sejina. These setae, often very minute, are added in the deutonymph, and usually occur on a free ventral intercalary sclerite in the adult. In the protonymph, the same paired av and pv setae appear on a free 3 intercalary sderite. Although, these seta& are called az?3 and a&in the Figure 47-6 of Krantz (1978), they are thought to be homologous to av4 and pv4 appearing on such sclerite in the deutonymph and adult. The intercalary sclerite is fused to the telotarsus in Celaenopsoidea.
Unlike the majority of mesostigmatid mites, trigynaspids do not retain an unpaired postanal seta (pon) in the adult. This seta appears in the larva, but is absent in subsequent stases. This character of the absence of pon in the adult is also shared by the members of Diarthrophallina and some Uropodina (such as Trachytes, Polyaspinus, and Uroseius). In contrast, Sejina (including Sejus, Epicroseius, Asternolaelaps, and Uropodella), the remaining Uropodina (such as Oplitis and Polyaspis), and other mesostigmatid mites (including Epicriina, Zerconina, Dermanyssina, Parasitina, etc.) retain this seta throughout all the stases.
Furthermore, not all, but many trigynaspids retain paired eugenital setae on the male genital opening, a character shared with some Uropodina, such as Phaulodinychus repletus (Berlese), Clausiadinychus, Cilliba cassidea (Hermann), Trachytes, Polyaspis, and Polyaspinus. This is compared to Sejina and Diarthrophallina, which lack male eugenital setae.
While trigynaspid mites share the characters above with other mesostigmatid mites, Trigynaspida has an apparently unique set of setal characters of retaining 4 anterolateral setae {al) on tarsi II-IV (Figures 2.5, a, b, c). This is compared with the remaining mesostigmatid mites that have a maximum of 3 anterolateral setae. With this character, trigynaspids typically retain 19 setae on tarsi n UI (4,4/3,4/2,2), and 21 on tarsus IV (4,4/4,4/3,2), which are compared with the other non-trigynaspid mesostigmatid mites, having the maximum of 18 on tarsi Il-in, and 20 on tarsus IV. Evans (1965) recognized a 'medioventrai hair (seta)' on tarsi II-IV as a character unique to Trigynaspida. Using such a term of 'medioventrai' hair (seta), however, is not appropriate as this term implies a 7-setae whorl (verticil) system, which is aberrant to the standard 6-setae whorl in Mesostigmata. These setae are actually av2 in tibia IV and av3 in tarsi II-IV in the adult. Moreover, these 'medioventrai' setae are also found in other Mesostigmata, including Sejina, and, therefore, are not unique to Trigynaspida.
Biology and Distribution
Trigynaspid mites are usually large' in their body size, ranging about 0.5 to 5 mm in the adults. Despite their large body size, however, they are rarely seen (Walter, 1997). In fact, many species have never been recovered since their original description. This fact of rareness linked with small population size in nature might have led to relatively insufficient studies of their taxonomy. Although about 20 species of trigynaspid mites have been reported from temperate Europe and North America, the vast majority of species have been reported from tropical and subtropical realms. Taxonomic literature on this group, such as Funk (1980), Hunter (1993a), Hunter and Butler (1966), Hunter and Rosario (1988), Hyatt (1964), Trâgârdh (1950a), and Turk (1948), report more than 200 species from the Neotropical region. Furthermore, it is believed that many of these are known as the fauna of former Gondwanaland, which is the great southern supercontinent appeared in geological history of the Earth, formerly comprising South America, New Zealand, Australia, India, Africa, Madagascar, and Antarctic Islands, including South Georgia, Crozet, and Marion Is., because they are predominantly collected from the territories of that ancient landmass. Trigynaspida is composed of two bifurcating lineages, which represent the suborders Cercomegistina and Antennophorina (Camin and Gorirossi, 1955). At present, Cercomegistina contains 17 described species in 6 families (Astemoseiidae, Cercomegistidae Davacaridae, Pyrosejidae, Saltiseiidae, and Seiodidae) within the single superfamily Cercomegistoidea (Trâgârdh, 1937). With a few exception of the species of Cercomegistidae that show association with bark beetles (as in Cercomegistus and Cercoleipus; Kinn, 1971), social spiders (Holocercomegistus; Evans, 1958), or pagurid crabs (Vitzthumegistus; André, 1937), both immature and adult stages of cercomegistines are free- living (saprophagous to predacious) in moist and decaying vegetation mostly found in the Southern Hemisphere.
The members of Antennophorina are more diverse than those of Cercomegistina, containing 21 families within 7 superfamilies (Kethley (1977b) proposed 6 superfamilies of Antennophorina.). One peculiar feature of the biology and behavior of the antennophorines is that the adult stage of most of these mites exhibits well-known associations with passalids, scolytids, formicids, or diplopods, which are mostly gregarious or social groups of insects or arthropods. Three genera {i.e., Ophiomegistus (Paramegistidae), Ophiocelaeno (Diplogyniidae), and Indogynium (Schizogyniidae)) are associated with squamates. Among these hosts, passalids serve as the major host for trigynaspids in terms of the diversity of species (Hunter, 1993a; Hunter and Rosario, 1988). A few species of Antennophorina, such as the members of Triplogyniidae and Megacelaenopsidae along with Mixogynium proteae Ryke (Schizogyniidae), are known to be free-living as adults.
While adult antennophorines are often associated with their hosts, nearly all the known immatures, excluding Micromegistus (Parantennulidae), are free-living. These immatures are commonly found in 6 surrounding habitats of the hosts, feedingon nematodes, coUembolan eggs, fungal hyphae, and other organic debris (Butler and Hunter, 1968; Hunter and Davis, 1965; Kinn, 1971). As to the Micromegistus, all the postembryonic stases are found on a carabid beetle. Unlike other arthropods that serve as the hosts for Antennophorina, carabids are not gregarious and often move fast through the wide range of habitat. Although such behavior of the host can offer higher chance of dispersal to Micromegistus, it is not difficult to imagine that it would likely to cause the state of orphanhood to immatures if they are not on the beetle. Instead of feeding on organic debris around the host habitat, imm ature Micromegistus feeds on fungal hyphae or other organic debris on the beetle, presumably acquired by the beetle's behavior (movement), preferring moist and decaying habitats (Nickel and Elzinga, 1970b; pers. observation).
It should be noted that these types of mite-host relationships appearing in most Antennophorina are not parasitic but can be phoretic, and often are paraphagic, taking the body secretions from the hosts, to commensal (Butler and Hunter, 1968; Hunter, 1993a; Hunter and Davis, 1965; Kinn, 1971; Nickel and Elzinga, 1970b; Trâgârdh, 1907). For the Antennophoridae of Antennophorina, well-known associates of ants, Franks et al. (1991) argued Antennophorus grandis Berlese as an obligate ectoparasite on Lasius. Although imitating ants' trophallactic behavior by mites to take liquid foods as observed by Franks et al. (1991) was well-reported in previous literature (Banks, 1915; Donisthorpe, 1927; fanet, 1897), there is no clear evidence indicating that Antennophorus is parasitic to (or exploit) the host or hosts' colony. While the mite may not mimic ants' colony odor, any roaming mite is picked up by the ant from any colony, and furthermore, when the mite's antenniform leg I are artificially amputated, any ant picks up the mite and delivers it to the ants' brood pile and place among the larvae or among the soil debris (Franks et al., 1991: p. 66). More detailed research is therefore 7 necessary to understand this subtle and enignatic relationships between the mites and their host ants. Perhaps, like many antennophorine mites, Antennophorus might also be mutualistic with their hosts by attacking the hosts' natural enemies. For another antennophorid species on ant, Antennophorus donisthorpei Wheeler, Hunter and Rosario (1998) also stated "However, direct harm (from the mite) to the host (ant) does not occur". Trâgârdh (1907) speculated paraphagic behavior of Neomegistus julidicola Trâgârdh (Paramegistidae) on its host millipede. Despite the known and empirical facts that the adult trigynaspid mites are associated with typical groups of hosts stated above, their biology is generally poorly known (Walter, 2000).
Systematics and Classification
The major systematic and biological studies on trigynaspids were carried out by Camin and Gorirossi (1955) for Trigynaspida; Funk (1968,1974, 1977,1980) for Celaenopsoidea; Hunter (1970) for Davacaridae; Hunter and Davis (1965) for Euzerconidae; Rosario and Hunter (1987, 1988) for Klinckowstroemiidae; Kim and Klompen (2001) for Paramegistidae; Kinn (1966,1967,1970,1971) for Schizogyniidae, Cercomegistidae, and Celaenopsidae; Lindquist and Moraza (1993) for Pyrosejidae; Nickel and Elzinga (1970b) for Parantennulidae; Seeman and Walter (1997) for Triplogyniidae; Trâgârdh (1906,1907,1946a, 1948,1950a, 1950b) for Celaenopsidae, Diplogyniidae, Schizogyniidae, and Paramegistidae; Walter (2000) for Saltiseiidae; and Womersley (1958a, 1958b, 1959) for Diplogyniidae, Parantennuloidea, and Fedrizziidae. Despite these efforts, however, the trigynaspids are still one of the poorly-known groups among the Acari in terms of the number of described species, ecology, and phylogenetic relationships within and among the trigynaspids and other mesostigmatid m ites. 8 The evolutionary relationships of trigynaspid mites at higher-level was first hypothesized by Trâgârdh (1907,1937,1946a) as the parts of his studies on systematics of Mesostigmata. His works, based on the viewpoint of evolutionary taxonomy that groups the taxa with diagnostic characters which are often autapomorphic to the taxon, are outdated as many new taxa have been reported since then. Furthermore, many of his major groups contain a mixture of many members from apparently different superfamilies (also see Chapter 3). Funk (1968) hypothesized the relationships among the families of Celaenopsoidea, one of the superfamilies that constitute Trigynaspida, based on numerical phenetic UPGMA (Sokal and Sneath, 1963) by using all the discrete and continuous characters, such as measurements of setae, appendages, and the body. Although UPGMA (and other distance methods) may have incomparable superiority of algorithmic unambiguity to other phylogenetic methodologies, it has no relationship with the organisms' ancestry (see Felsenstein, 2001). It also has another innate defect of always assuming equal rate of evolution along the sister branches from their co ancestor. In nature, the rate of evolution is not necessarily equal in two sister taxa.
Problem status quo
The current scheme of higher-level relationships and classification of trigynaspid mites was proposed by Kethley (1977b), where he suggested 24 families, including 7 new families, within 7 superfamilies based on his study on the patterns of the distribution of setae {i.e., chaetotaxy) on the legs. In his work, Kethley assumed that Trigynaspida is a natural group, indicating all of its members have a single ancestor (Figure 1.2). His results also confirmed bifurcating internal lineages of Cercomegistina and Antennophorina within Trigynaspida, proposed by Camin and Gorirossi (1955). However, despite the 9 8 V 8 8 2 o o 8 I s !o m % o
O) 0) O) § 0) 0) 0) (D Cl) CD CB o O) O 0) O) III o > ; Z I o z o o IIIa uj CO OCD < U u YYYJ
Figure 1.2: Phylogenetic tree re-drawn from Kethley (1977b). Note that two ant associates of Antennophoroidea and Aenictequoidea are not sisters to each other. Paramegistidae, Celaenopsidae, and Schizogyniidae are not monophyletic.
10 general fact that his classification has been widely accepted by many textbooks, such as Krantz (1978), Evans (1992), or Alberti and Coons (1999), it should be noted that Kethley's results from the study that relied only on leg chaetotaxy contain several problems.
First, although chaetotaxy is often useful in acarine systematics as these patterns are often conserved within a group, almost invariable between sexes, and often variable among the groups (Evans, 1963a, 1963b, 1964,1965,1969, 1972), interpretation of setal homology can often be controversial when the base of setae are moved in position (see Lindquist and Moraza, 1993). Evans' (1965) failure to recognize anteroventral (av) setae on tibiae and tarsi as 'medioventrai {mv)' setae is a good example. In other words, although chaetotactic data may offer phylogenetic information to some extent, interpretation of those patterns might likely be difficult enough to cause the confusion.
Second, relying only on one (or limited) set of morphological characters in systematics, such as the leg chaetotaxy for Trigynaspida as in Kethley (1977b) or chelicerae or mouthparts for the "Gangsystematik" Uropodina as in Hirschmann (1975, 1993) or Wisniewski and Hirschmann (1993), is equivalent to using a small set of a single gene to deduce phylogenetic relationships that may yield a gene tree, which is dangerous and should not be encouraged. In short, no single character of morphological attributes should be used in inferring evolutionary relationships at the higher level (Evans, 1972). Instead, morphological analyses should rely on multiple sets of morphological attributes, such as those from the mouthparts and reproductive organs, which are important for pursuing specific functions and behavior of the organisms as these features are also often conserved in the context of evolution. As the members of Trigynaspida have, for example, very peculiar genital structures along with sternal and often anal shield 11 structures in the female, these characteristics should not be disregarded in phylogenetic analyses.
Third, there is no clear coding scheme in Kethley's analyses. Although character coding can be achieved by relying either on outgroup comparisons or on information on ontogeny (Watrous and Wheeler, 1981), his analyses do not include any outgroups other than Trigynaspida per se, and there is no information regarding this question. Without including closely related outgroup taxa that allow the test of monophyly of Trigynaspida, he implicitly assumed that the group Trigynaspida is monophyletic, leaving his results of systematics unreliable. The test of monophyly of the ingroup under study by including the outgroups is fundamental in phylogenetic systematics as the outgroups serve as the control group' in phylogenetic systematics. The philosophy of phylogenetic systematics is simply based on the idea that all members of the ingroup taxa are the descendants derived from a single ancestor in a history of evolution {i.e., monophyly). Furthermore, even in the phylogeny of Trigynaspida proposed by Kethley (1977b), the appearance of non-monophyly even within a family is also observed. For example, in his cladogram, genera of Paramegistidae (including Paramegistus, Neomegistus, Ophiomegistus, and two species of Echinomegistus), Celaenopsidae and Schizogyniidae are not monophyletic (see Figure 1.2).
Finally, his classification based on his analyses shows wide spread of host association, in which, especially, the associations with carabids, ants, and millipedes are not conserved within his classification. For example, his two superfamilies that exhibit the associations with the ants {i.e., Aenictequoidea and Antennophoroidea) show disparate separation, showing non-sister group relationships within his cladogram. Species of Micromegistus (Parantennulidae) and Promegistus (Promegistidae) that are associated with carabids are also distantly related in his cladogram. Although, the co- 12 evolution among the hosts and parasites does not have to follow Fahrenholtz's rule (Brooks and McLennan, 1993; Mitter and Brooks, 1983) showing somewhat mirror-like topologies of evolutionary trees of these entities, it is worthwhile to suspect and examine if there are conserved relationships among the hosts and these mites at the higher level.
Goal of the Study
Although Kethley's paradigm retains these questions, nobody has studied higher-level relationships among these mites since then. Accordingly, as Lindquist and Moraza (1993) noted, the higher-level relationships among trigynaspid mites and their allies need to be re-examined based on a phylogenetic viewpoint (Wiley, 1981), which is the most common and reasonable approach towards the understanding of evolutionary relationships among the biological organisms.
To address the answer to the questions of monophyly of Trigynaspida and the higher-level relationships among its members, studies of comparative morphology (Chapter 3) and molecules (nuclear ribosomal DNA sequences; Chapter 4) have been conducted. That is, by using both morphological and molecular data, the present study is intended to test if the group Trigynaspida is indeed monophyletic. Although good diagnostic characters to define the group Trigynaspida were empirically given above, global test of monophyly is required. Using two different sets of data in inferring evolutionary relationships is ideal as the 'hypothetical' results (i.e., estimated relationships among the taxa) from one data set can be 'confirmed' by another. These analyses, by themselves, also allow the examination of the higher-level phylogenetic relationships among the superfamilies and families of trigynaspid mites simultaneously. Selection of outgroup taxa was based on the proposed phylogeny of Johnston in Norton et aL (1993, Figure 13 12). Outgroups were used only for rooting, the trees» and therefore» inferring the relationships among the outgroups is beyond the scope of this study.
The scope of this study also expands to the relationships among the trigynaspid mites and their hosts (Chapter 5). That is, by adding the information of the hosts to the estimated phylogeny, the evolutionary patterns among these mites and their hosts have been introduced. Parallel to this approach, biogeographic patterns of global distribution of these mites on different continents were also briefly discussed (Chapter 5). With an aid from plate tectonics, this approach allows the estimation of the age of Trigynaspida in a geological time.
The building of evolutionary relationships among the major groups of trigynaspid mites based on morphological and molecular data is of considerable importance to acarology, because this is the first interpretation on this group based on the viewpoints of phylogenetic systematics. This study has challenged the currently accepted relationships of Trigynaspida proposed by Kethley (1977b), and the resultant relationships and the diagnoses of the superfamilies along with the new key to the families from the present study have newly been proposed. The data and results from this study can also be used as a stepping stone to a broader research agenda, such as the study of phylogenetic relationships among the entire groups of Acari {i.e., mites and ticks), which can provide valuable framework for examining a variety of ecological and physiological problems in the future.
14 CHAPTER 2"
A NEW GENUS AND SPECIES OF PARAMEGISTIDAE (MESOSTIGMATA: TRIGYNASPIDA) ASSOCIATED WITH MILLIPEDES FROM MEXICO
INTRODUCTION
T he Mesostigmatan trigynaspid mite family Paramegistidae, proposed by Trâgârdh (1946a), currently includes 25 described species in five known genera, Antennomegistus Berlese, 1904; Echinomegistus Berlese, 1904; Neomegistus Trâgârdh, 1906; Ophiomegistus Banks, 1914; and Paramegistus Trâgârdh, 1906. Among these, Antennomegistus and Echinomegistus species are associated with carabid beetles (Banks, 1904; Berlese, 1888,1892,1904; Hyatt, 1964; Krantz, 1978; Nickel and Elzinga, 1970a), Ophiomegistus species are well- known associates of lizards, skinks, and snakes (Banks, 1914; Domrow, 1978, 1984,1987; Goff, 1979,1980a; 1980b; Grant, 1947; Gunther, 1942,1951; Voss, 1966; Womersley, 1958b), while Neomegistus and Paramegistus species have been reported from julid millipedes (Trâgârdh, 1906, 1907, 1948; Womersley, 1958b). Unlike the majority of Mesostigmata, the body in Paramegistidae is somewhat wider than long. Like most adult members of the infraorder
^ This Chapter has been accepted by Acarologia, and is in print.
15 Antennophorina^ the body of Paramegistidae i& well-sclerotized. Little is known about their biology or about the morphology of the immature stases, except for a few comments on Echinomegistus wheeleri by Nickel and Elzinga (1970a) and on Neomegistus julidicola by Trâgârdh (1907). A brief summary of biogeography, host associations, and number of described species is given in Table 2.1.
Genus Distribution Host No. of described species Antennomegistus Paraguay carabid beetle 1 Echinomegistus USA, Nicaragua*, carabid beetle 2 Venezuela Meristomegistus Mexico m illipede 1 Neomegistus S. Africa, m illipede 1 Madagascar* Ophiomegistus Indo-Australia squamate 20 Paramegistus S. Africa millipede 1
Table 2.1: Biogeography, host association, and numbers of described species of Paramegistidae. (* = new records)
Recently, we could examine some mite-bearing millipedes collected in southern Mexico. Among the mites collected were specimens that looked very similar to the known members of the family Paramegistidae in terms of morphology of the genital structures and the chaetotaxy of the appendages. However, these specimens do not key to Paramegistidae in the family key for 16 Trigynaspida as proposed by Kethley (1977b). In this key, Kethley used fusion of the palp tibiae and tarsi of the adults as one of the diagnostic characters of Paramegistidae. In the new material, however, the palp tibiae and tarsi are distinctly separated, disallowing inclusion in the Paramegistidae. Neither can these specimens be included in Kethley's Promegistidae, the proposed sister family of Paramegistidae (Kethley, 1977b). That family shares the separate palp tibiae and tarsi, but has no stemogynial or prestemal (jugular) shields, shields that are present in our material and in all currently known members of the Paramegistidae. In addition, Promegistidae retain only 6 (not 7 as in most Antennophorina) setae on palp genua, a peculiar character shared with Micromegistus gourlayi (Parantennulidae) and Philodana johnstoni (Philodanidae).
Accordingly, instead of raising a new family, 1 describe the above specimens as a new genus and species, and provide a revised diagnosis of the family Paramegistidae, allowing inclusion of the newly proposed genus. A key to the genera in the family is provided. This is the first record of Paramegistidae associated with millipedes in the New World.
17 MATERIALS AND METHODS
Notations for leg chaetotaxy follow Evans (1963a, 1965,1969). Mites were mounted in Hoyer's medium (Krantz, 1978). All measurements are in micrometers (|im).
In this description, we adopt a simplified terminology for the structure(s) on the ventral side of the idiosoma located between tritostemum and the sternal shield(s), previously referred to as prestemal shield(s)' (Kethley, 1977b; Krantz, 1978), jugular shield(s)' (Krantz, 1978), 'jugularium' (Evans, 1992), 'tetartostem um ' (Kethley, 1977b), or tetratostem um ' (Hunter, 1993b). We propose to use prestemal shield(s) for all sclerotized plates anterior to the sternal shield(s), that are characterized by the presence of sternal setae stl and/or lyrifissures stpl. Any small sclerotized platelets appearing in the prestemal area, but lacking both setae and lyrifissures, are designated as prestemal platelets'.
18 DESCRIPTION
Meristomegistus n. gen.
Type Species: Meristomegistus vazquezus n. sp.
Diagnosis: Palp tibiae and tarsi distinctly separated (Figure 2.3, c). Pilus dentilis on fixed cheliceral digit absent. Prestemal shields weakly fused or paired. Idiosomal venter with distinctive, stylus-like setae. Female: sternal shields paired; stemogynial shields subtriangular, weakly-separated; mesogynial shield squared, fused posteriorly with ventrianal shield (Figure 2.2). Male: eugenital setae present.
ETYMOLOGY: Reflecting the characteristic feature of clear articulation of the palp tibiae and tarsi. 'Meristo' in Greek means separation, and a suffix, 'megistus', derived from a Latin word of 'megistanes’, means magnate. Genus name is masculine in gender.
Meristomegistus vazquezus n. sp. (Figures 2.1-2.8, b)
Diagnosis: Same as for genus. F e m a le (Figures 2.1-2.5, c). Body oval; idiosoma 829 long, 608 w ide at middle (at the line between coxae IV). Body margin with 16 pairs of strong, smooth spinous setae of 19-61 long. 19 Idiosoma. Dorsum (Figure 2.1). Covered by an unomamented holodorsal shield bearing numerous small setae of similar size {ca. 25 long). Venter (Figure 2.2). Tritostemal base ellipsoid, arising from anteromedian area of prestemal shields; two barbed laciniae (108), reaching almost to the base of comiculi. Prestemal shields unomamented, weakly coalesced medially, each bearing 1 smooth, stylus-like seta stl (42) plus 1 sternal lyrifissure (stpl). Paired sternal shields unomamented, small, more or less triangular, each with 3 smooth, stylus-like setae, st2, st3, and st4 (40-74), plus 1 lyrifissure (stp2). Paired stemogynial shields weakly sclerotized, each bearing 1 stemal lyrifissure (stp3). Paired latigynial shields right-angled triangular, slightly covering mesogynial shield; each shield with 1 glandular pore plus 2-4 minute glandular poroids along with 2 smooth, stylus-like setae of similar size (57). Mesogynial shield square-shaped, fused posteriorly with ventrianal shield. Ventral and anal shield fused to form ventrianal shield. Ventrianal shield pyriform, with 15 setae. Anal opening located in distal end of ventrianal shield, bearing 2 anal valves, each with a small lyrifissure; adanal, postanal, and euanal setae absent. Postanal cribrum absent, but paired lyrifissure present. Metapodal shields broad, fused with exopodal and peritrematal shields; notch present near stigmatal opening; 1 smooth, stylus-like seta, 1 glandular pore, along with 8 minute glandular poroids located in distal end of each metapodal shield. Stigmatal opening between coxae UJ and IV. Peritremes long, reaching almost to the anterior end of idiosoma.
Gnathosoma (Figures 2.3, a, b). Gnathotectum more or less pentagonal, with anteromedian spade-shaped projection bearing obscure dorsomedian keel; anterolateral margin smooth. Ventrally all 4 pairs of gnathosomal setae smooth. Hypostomal setae hsl longer than any other hypostomal setae. Comiculi not strong, more or less membranous, distinctly bifurcate. Deutostemal groove with 3-4 rows of minute denticles. 20 Palpi (Figures 2.3, b, c). Trochanters 57 long, without a distinct apophysis, each with 2 setae; femora 57 long, each with 5 setae; genua 38 long, each with 7 setae, dorso-proximal lyrifissure present; tibiae 29 long, each with 15 setae; tarsi 23 long, each with 15 setae. Palp tibiae and tarsi clearly articulated (Figure 2.3, c). Palptarsal claw (apotele) 2-tined.
Chelicerae (Figure 2.3, d). Chelate. Movable digit edentate, fixed digit with row of 5 small distal teeth. Excrescences hyaline; medial filament thickened and finely plumose. Cheliceral seta on middle of fixed digit antiaxial. Two dorsal lyrifissures present at base of fixed digit. Pilus dentilis on fixed digit absent.
Legs (Figures 2.4, a-2.5, c). Leg chaetotaxy of coxae, trochanters, femora, genua, tibiae, and tarsi of legs I-IV as in Table 2.2. Setal notations as figured. Coxae I- IV each with lyriform pore in anterior proximal position (Figure 2.2). Coxal base n-rv with rows of minute comb-like denticles. Base of femora I-IV with circumsegmental fissure. Tarsi I without claw or ambulacrum, with 7 lobe like blunt-end sensilla at distal end (Figure 2.4, c). Tarsi II-IV each with paired claws and a fan-like ambulacrum. Tarsi II-III each with ventral intercalary sclerite in circumsegmental fissure with no setae. Tarsi IV with setae av4 and pv4 on ventral intercalary sclerite in circumsegmental fissure.
Male (Figures 2.6-2.S, b). Body shape and size same as female.
Idiosoma. Dorsum (Figure 2.6). Similar to female but with additional lateral opisthosomal setae on the shield (possibly UR series). Venter (Figure 2.7). Prestemal shield separated from fused holoventral shield. Genital opening between coxae HI and IV, more or less ellipsoid, covered by 2 valves; anterior 21 Segment I n m IV Coxa 2 2 2 2 Trochanter 6 5 5 5 (1,1/2,0/1,1) (1,0/2,0/1,1) (1,1/2,0/1,0) (1,1/2,0/1,0) Fem ur 12 10 7 8 (1,2/2,2/3,2) (2,2/1,2/2,1) (1,2/1,2/1,0) (1,2/1,2/1,1) Genu 11 12 10 10 (1,3/1,3/1,2) (2,3/1,3/1,2) (2,3/1,2/1,1) (2,3/1,2/1,1) Tibia 13 10 9 10 (2,3/2,2/2,2) (2,2/1,2/1,2) (2,2/1,2/1,1) (2,2/2,2/1,1) Tarsus 48 19 19 21 (4,4/3,3/2,3) (4,4/3,3/2,3) (4,4/4,4/3,2)
Table 2.2: Leg chaetotaxy of Meristomegistus vazquezus n. sp.
valve with paired smooth eugenital setae (11) at laterobasal margin. Stemal and ventrianal region separated by notch postero-laterally to genital opening. Ventrianal region with 19 smooth, stylus-like setae plus paired glandular pores. Metapodal shield broad, each with 1-2 setae and 1 glandular pore. 5-8 minute glandular poroids on hee small circular platelets posterolateral to metapodal shields, instead of on the distal end of metapodal shields (as in female).
Gnathosoma (Figures 2.8, a, b). Gnathotectum triangular, without anteromedian spade-shaped projection; dorsomedian keel obscure. Setae hs2 longer (93) than any other hypostomal setae, barbed up to the middle but
2 2 smooth at distal end; much longer than in the female. Comiculi not bifurcate, not strong, more or less membranous.
Palpi and Chelicerae: As in female.
Legs: Chaetotaxy as in female.
IMMATURES: U nknow n.
ETYMOLOGY: This species is named in honor of the collector. Dr. MA. M agdalena VAzquez, Universidad de Quintana Roo, Mexico.
TYPES: A holotype female (OSAL 000387) and an allotype male (OSAL 000388) deposited in Universidad Nacional Autonoma de Mexico (UNAM), Mexico City, Mexico. Two para type females in alcohol (OSAL 000389,000390) deposited in the Acarology Laboratory (OSAL), The Ohio State University, Columbus, Ohio, U.S.A.
TYPE DATA: MEXICO: Quintana Roo, Noh-Bec, 19°7'24"N 88°20'20"W, high tropical forest, coll. M. Vazquez, 4 Sep. 1997, ex Aceratophallus n.sp. (Diplopoda: Polydesmida: Rhachodesmidae), coll. no. AL5565. Host millipede in collection of Dr. Richard L. Hoffman, Virginia Museum of Natural History, Martinsville, Virginia, USA, labelled "CMK 97-I005-I, mites removed"
23 DISCUSSION
According to Kethley (1977b), the mite family Paramegistidae has two major diagnostic characters: 'paired stemogynial shields' and fused palp tibiae and tarsi'. The latter character is applicable only to the genera Antennomegistus, Echinomegistus, Neomegistus, and Ophiomegistus, but not to Paramegistus, because Paramegistus retains a weak separation of the palp tibiae and tarsi. We thus propose a revised diagnosis of the family Paramegistidae as follows:
Oval, flattened mites with minute dorsal setae and distinctly larger stylus-like, spinous, or foliate ventrianal setae. Prestemal shield always present. Stemal shield paired or two shields connected by narrow bridge. Stemogynial shields, bearing stp3, paired. Mesogynial shield free from or fused to large ventrianal shield; latigynial shields generally well-developed. Palp tibia and tarsus fused or rarely separated; chelicerae edentate, movable digit with filamentous or more or less dendritic excrescences. Comiculi membranous, often with sclerotized tip. Male eugenital setae often present.
The new mite genus shares with Paramegistus the partial or complete separation of the palp tibiae and tarsi. It differs from Paramegistus by the structure of the mesogynial shield. In female Paramegistus (i.e., P. confrater) the mesogynial shield is completely free from the ventrianal shield, a
24 character shared with Antennomegistus and Echinomegistus. In contrast, in the new genus, the mesogynial shield is completely fused posteriorly with the ventrianal shield. This character state is shared with the paramegistid genera Neomegistus and Ophiomegistus.
25 A KEY TO THE GENERA OF THE FAMILY PARAMEGISTIDAE (BASED ON FEMALE)
1. Palp tibiae and tarsi fused ...... 2 1'. Palp tibiae and tarsi not fused; associated with m illipedes ...... 5
2. Mesogynial shield free, not fused with ventrianal shield; associated with carabid beetles ...... 3 2'. Mesogynial shield fused with ventrianal shield; associated with millipedes or squamate reptiles ...... 4
3. Latigynial shields paired, free, not fused with ventrianal shield ...... Antennomegistus 3'. Latigynial shield fused with ventrianal shield posteriorly ...... Echinomegistus
4. Latigynial shields with numerous setae, each shield at least with 3 setae (usually with 5 to 30 setae); posterior end of metapodal element with short stout subulate setae; associated with squam ates ...... Ophiomegistus 4% Latigynial shields without numerous setae,, each shield bearing 2 setae; posterior end of metapodal element without short stout subulate setae; associated with millipedes ...... Neomegistus
5. Mesogynial shield free from ventrianal shield ...... Paramegistus 5'. Mesogynial shield fused posteriorly with ventrianal shield; idiosoma longer than w id e ...... Meristomegistus
26 Figure 2.1: Meristomegistus vazquezus n. sp., female, idiosoma, dorsum. 27 e s
K i K
Figure 2.2: Meristomegistus vazquezus n. sp., female, idiosoma, venter. 28 100 100
Figure 2.3; Meristomegistus vazquezus n. sp., female, gnathosoma. a: Gnathotectum; b: Ventral view (palp tarsal setae are eliminated); c: Palp tibia and tarsus, dorsal view; d: Chelicera. 1 z
Figure 2.4: Meristomegistus vazquezus n. sp., female, leg I, dorsal view, a: Overview; b: Tarsus I; c: Tarsus I, distal blunt-end sensilla. F
a df al2 9d1 agg /a iO a» _a„
pd3 % w
Figure 2.5: Meristomegistus vazquezus n. sp., female, legs IIIV, dorsal view, a: Leg II; b: Leg III; c: Leg IV. Figure 2.6: Meristomegistus vazquezus n. sp., maie, idiosoma, dorsum.
32 i
a
V
X
t e
Figure 2.7: Meristomegistus vazquezus n. sp., male, idiosoma, venter.
33 b
100
100
Figure 2.8: Meristomegistus vazquezus n. sp., maie, gnathosoma. a: Gnathotectum; b: Ventral view.
34 CHAPTER 3
MORPHOLOGICAL PHYLOGENETICS OF TRIGYNASPIDA (MORPHOLOGICAL ANALYSES)
INTRODUCTION
Ever since Aristotle, external and internal morphological characteristics have been used by naturalists to describe and to examine the 'creatures' in nature. With the contribution of Carl von Linne's Systema Naturae, 1758, this taxonomic tradition is continued in systematics, which deduces the relationships among these 'species ' in an orderly manner. Though it has relatively a short history, acarology, the science of mites and ticks, was not an exception to this tradition. Indeed, acarologists even developed the system of recognizing homologous hairs (setae) on the body and appendages! (Grandjean, 1939; Lindquist and Evans, 1965; Sellnick, 1944)
Mites are usually small in their body size, and with this bodily constraint, they occupy a variety of habitats by their highly adapted features of the body. These features often result in high degree of morphological convergences (as in body shape, in mouthparts, etc.) or retrogressive evolution by the reduction of characteristics (as in many parasitic mites). Although this often led to poor description (as in many early taxonomic
35 literature on Eriophyoidea and ÂstigmataXor incorrect identification of mites at low levels, it also influenced higher level systematics, leaving confusion and controversies on higher level relationships of Acari. Questions on the phylogenetic position of Eriophyoidea (Lindquist, 1996), Endeostigmata (Norton et al, 1993; OConnor, 1984), or those contradicting hypotheses on the monophyly versus non-monophyly of Acari as a whole would be a few good examples (Van der Hammen, 1977,1989; Lindquist, 1984; Shultz, 1990; Weygoldt and Paulus, 1979; Zachvatkin, 1952). A word from Krantz (1978), saying, "Acarology is, in fact, in a state of systematic turmoil similar to that experienced by entomologists a century ago.", very well summarizes status quo of acarology.
The hypotheses of evolutionary relationships of trigynaspid mites at the higher-level were proposed by Trâgârdh (1907,1937,1946a) as the parts of his works on systematics of Mesostigmata. His studies were based on the point of view of 'evolutionary taxonomy', which groups the taxa with diagnostic' characters which are often unique to the taxon. Regardless of the accuracy of this methodology or his results, his works are outdated as many new taxa have been reported since then. Many of his major groups contain a mixture of members from apparently different superfamilies. Later, in his unpublished Ph.D. dissertation. Funk (1968) hypothesized relationships among the families of Celaenopsoidea, one of the superfamilies comprising Trigynaspida, based on numerical phenetic ÜPGMA by using all the discrete and continuous characters (such as, measurements of setae, appendages, and body). Although UPGMA (and other distance methods) may have incomparable superiority of algorithmic unambiguity to other phylogenetic methodologies, it has no direct relationship with the organisms' ancestry. It also has another innate defect of always assuming equal rate of evolution along the sister branches from their co-ancestor. In nature, the rate of evolution is not necessarily equal in two sister taxa (Ayala, 1997; Li, 1997). 36 The current scheme on higher-level relationships among the trigynaspid mites as a whole was proposed by Kethley (1977b). His work, based on the study on the patterns of distribution of the setae (i.e., chaetotaxy) of the legs (see Figure 1.2), was adopted by Krantz (1978) and has become the standard in subsequent major textbooks, including, but not limited to, Evans (1992) and Alberti and Coons (1999). Chaetotaxy is often useful in acarine systematics as these patterns are often conserved within a group, are almost invariable between male and female, and are often variable among the groups. (Evans, 1963a, 1963b, 1964,1965,1969). The interpretation of setal homology, however, can often be controversial when the base of setae are moved in position (see Lindquist and Moraza, 1993). For example, Evans (1965) recognized medioventral hairs' (setae), appearing on tarsi II-IV of mesostigmatid mites as a unique character for Trigynaspida. This interpretation, which implies a 7-setae whorl (verticil) system rather than a 6- setae one, is incorrect as these are actually anteroventral (av) setae, one of the sets of setae that constitute standard 6-setae whorl system in Mesostigmata (Evans, 1969). In other words, although chaetotactic data can offer phylogenetic information, interpretation of chaetotactic patterns, other than the number of setae on a segment, might be difficult enough to cause the confusion. Relying on one or a limited set of morphological characters in systematics, such as chaetotaxy for Trigynaspida (Kethley, 1977b) or chelicerae or mouthparts for Uropodina (Hirschmann, 1993), is somewhat equivalent to using a small set of a single gene to deduce phylogenetic relationships, which, if not experimental, should be cautioned. As the members of Trigynaspida retain very peculiar genital structures along with sternal and often anal shield structures in female, these characteristics should be included in phylogenetic analyses.
3 7 This study was performed to re-examine the higher level phylogenetic relationships and classification of Trigynaspida proposed by Kethley (1977b), with 'possible total morphological characters'.
38 MATERIALS AND METHODS
Taxon Sampling
Representatives from all the currently-known superfamilies were included to the study. This includes a total of 51 species, including 40 species in 23 families of Trigynaspida along with 11 species in 9 families of their 'monogynaspid' outgroup taxa (Table 3.1). Selection of outgroup taxa was based on the proposed phylogeny of Johnston in Norton et al. (1993, Figure 1.2). Outgroups are used for rooting the trees, and therefore, inferences of outgroup relationships are beyond the scope of this study. Except for the uropodine outgroup species of Thinozercon, this study was performed based on the slide-mounted specimens housed at the Acarology Laboratory (OSAL), Museum of Biological Diversity, Department of Entomology, The Ohio State University.
To test Kethley's hypothesis of the relationships between the two superfamilies of ant associates, species of Antennophorus (Antennophoroidea) along with Aenicteqiies and Ptochacarus (Aenictequoidea) were included. For the investigation of the monophyly of Paramegistidae, species from all the known genera (i.e., Antennomegistus, Echinomegistiis, Neomegistus, Ophiomegistiis, and Paramegistus) along with Meristomegistus, a new genus recently described (Kim and Klompen, 2001), were included. Finally, to examine the relationships among the associates of
39 caiabids^ Micromegistus (PaiantennulidaeX and Promegisttis (Promegistidae) were included.
In addition to the above sampling scheme focused directly on the testing of Kethley's cladogram, the following strategies were also considered. That is, to examine the relationships among the species within a same family from the same continent, Fedrizzia and Neofedrizzia (Fedrizziidae), passalid associates from Australia were included. As a possible back up of investigating this relationship, two species of Klinckowstroemiidae (Klinckowstroemia), another passalid-associated sister family to Fedrizziidae, from Central America were also included. Although these two families are very similar to each other in terms of morphology, their biogeographic distribution is quite peculiar, showing no overlap in the global distribution (Hunter, 1993a).
In an expansion of this sampling scheme, Micromegistus bakeri (Parantennulidae) from North America and M. gourlayi from Australia were included to investigate the relationships within a same genus (family) but from different continents. In addition, to examine the relationships between two closely related species (genera) that are associated with two different host groups, passalid-associated Euzercon latus (Euzerconidae) and millipede- associated Neoeuzercon sp. (Euzerconidae) were included to the study.
Analyse?
All the characters were coded with equal weight. Multistate characters were coded as multistate. Heuristic searches with TBR branch swapping along with random addition of taxa with 100 replicates were performed to avoid local optima. Maximum parsimony, which seeks the minimum number of steps of evolutionary change within the data, implemented in PAUP* 4.0b8 40 (Swoffotd, 2001) was used as the optimality cntenoiu Branches were collapsed if the maximum branch length is zero. To assess the confidence level on the deduced phylogeny, tranch support' (Bremer, 1988) (= 'decay index' sensu Donoghue et al., 1992; 'Bremer support' sensu Kâllersjô et ai, 1992) estimated by AutoDecay 4.0.2 (Eriksson, 1999) using global reverse constraints of topologies was performed.
For the characters of #15, and #26 through #29, lOOX differential interference contrast (DIC) objective lens (Zeiss Plan-Neofluar) with oil immersion was applied for the all taxa for the consistency in the character coding. Terminologies used here follow Evans (1992), otherwise specified.
41 Superfamily and Family Species SUBORDER CERCOMEGISTINA
S.F. Cercomegistoidea
Astemoseiidae Astemoseius sp. Cercomegistidae Cercoleipus coelonotus Cercomegistus evonicus Davacaridae Davacarus gressitti Pyrosejidae Pyrosejus sp. Pyrosejidae n. sp. Saltiseiidae Saltiseius sp. Seiodidae Seiodes sp. SUBORDER ANTENNOPHORINA
S.F. Celaenopsoidea
Celaenopsidae Pleuronectocelaeno drymoecetes Diplogyniidae Cryptometastemum sp. Ophiocelaeno sellnicki Passalacarus sylvestris Trichodiplogynium sp. Euzerconidae Euzercon latus Neoeuzercon sp. Megacelaenopsidae Megacelaenopsis oudemansi Megacelaenopsis sp. Schizogyniidae Paraschizogynium odontokeri Triplogyniidae Funkotriplogynium vallei Triplagynium sp. S.F. Parantennuloidea
Parantennulidae Micromegistus bakeri Micromegistus gourlayi Philodanidae Philodana johnstoni SJ^. Aenictequoidea
Aenictequidae Aenicteques chapmani Ptochacaridae Ptochacarus sylvestrii S.F. Antennophoroidea
Antennophoridae Antennophorus wasmanni
(Continued)
Table 3.1: List of taxa used for the morphological study. S.F.: Superfamily
4 2 Table 3.1 (Continued)
S.F. Megisthanoidea
Hoplomegistidae Stenostemum truittae Stenostemum sp. Megisthanidae Megisthanus floridanus S.F. Fedrizzioidea
Fedrizziidae Fedrizzia sp. Neofedrizzia leonilae Klinckowstroemiidae Klinckowstroemia starri Klinckowstroemiella blumae Paramegistidae Antennomegistus caputcarabi Echinomegistus wheeleri Meristomegistus vazquezus Neomegistus julidicola Ophiomegistus sp. Paramegistus confrater Promegistidae Promegistus armstrongi SUBORDER UROPODINA
Thinozerconidae Thinozercon michaeli Trachyuropodidae Oplitis sp. Poiyaspididae Polyaspis lamellipes Trachytes sp. SUBORDER SEJINA
Sejidae Squs sp. Epicroseius sp. Ichthyostomatogasteridae Astemolaetaps sp. Uropodellidae Uropodella sp. SUBORDER MICROGYNIINA
Microgyniidae Microgynium rectangulatum SUBORDER HETEROZERCONINA
Discozerconidae Discozercon sp. Heterozerconidae Heterozerconidae n. sp.
43 BAUPLAN OF TRIGYNASPIDA
As the word Trigynaspida' indicates, the structures of female genital region of trigynaspid mites are crucial in recognizing/identifying this group of mites. However, as the female genital structures, which are often unique to each trigynaspid superfamily, are contiguous with sternal structures along the venter of the body, standardized criteria to distinguish genital structures from those of sternal are required as the first step of homology assessment among these structures.
Definition of Sternal Region
In Mesostigmata, including Trigynaspida, the venter of the female is composed of 4 major regions. From anterior to posterior, they are referred to as sternal, genital, ventral, and anal region. In males, shields in these regions are quite often fully fused to form a single hologastric shield. Although sexual dimorphism of the venter of mesostigmatid mites is often remarkable, the sternal region in both sexes is easily identified as the area posterior to (or around) the base of tritostemum.
The sternal region in the female is defined as the area, posterior to (or around) the tritostemum, retaining 4 pairs of setae (stl~st4) and/or 3 pairs of lyrifissures (stpl-stpS). In males, which usually have more than 4 pairs of setae on the fused stemogenital or hologastric shield, the area bearing s tl‘St4 and/or 3 pairs of lyrifissures is regarded as sternal in origin. In fact, the setae 44 sf5 (and often st6\ in. the male is thought to be homologous to the setae appearing in the genital shield of the females (i.e., latigynial in Trigynaspida or epigynial shield in most of Monogynaspida, including Oermanyssina), which bear these 5th or 6th setae. Subsequently, the area posterior to this sternal region becomes the beginning of the genital region in female.
The sternal region is often occupied by a single large shield (i.e., sternal shield), but in many cases, especially in Trigynaspida, this region is fragmented to three different kinds of shields based on the distribution of the setae and lyrifissures. From anterior to posterior, they are referred to as prestemal, sternal, and metastemal shields.
Prestemal shields, often paired, are defined as the shield(s) retaining stl and/or stpl (Figures 2.2 and 3.1). Often, stl are located off the shield(s) but stpl are positioned onto the shield(s), allowing recognition of the prestemal shield(s). This interpretation includes previous terms of 'prestemal shield(s)' (Kethley, 1977b; Krantz, 1978), 'jugular shield(s)' (Krantz, 1978), jugularium' (Evans, 1992), 'tetartostem um' (Kethley, 1977b) or tetratostem um ' (Hunter, 1993b). Any small sclerotized platelets appearing in the prestemal area, but lacking both setae and lyrifissures, are designated as prestemal platelets' (Kim and Klompen, 2001) (Figure 3.2, a).
Similarly, metastemal shields, usually paired, are defined as the shield(s) retaining both st4 and stp3. Metastemal shields are commonly found in Celaenopsoidea of Trigynaspida (Figure 3.3), Dermanyssina and Parasitina (Figure 3.4). Although these metastemals often resemble the latigynial shields prima facie, it should be noted that the latigynials, which are not stemal but genital in origin, never retain st4 and stp3 (but often bear st5 and st6). They should, therefore, not be confused or homologized with metastemal shields.
45 Figure 3.1: Fedrizzia sp. (Fedrizziidae). Note entire prestemal, bearing stl and stpl, and stemogynial shield, bearing sfp3. (modified from Johnston, 1968)
46 stl
sî2 ,st3 St4
stp3
pon
(a) (b)
Figure 3.2: (a) Rhodacarus roseus Oudemans (Rhodacaridae) and (b) Gamasellus vibrissatus Emberson (Gamasellidae), showing prestemal platelets and postanal seta (pon). Note the presence of 3 pairs of stp and 4 pairs of St on stemal shield. Prestemal and metastemal shields are absent. (modified from Johnston, 1968)
47 3 ^ n
\ / vu
/
Figure 3.3: Celaenopsis badius (C. L. Koch) (Celaenopsidae), showing paired metastemal (MT) and ventromarginal (VTM) shields. Postanal seta is absent in ail the postlarval Trigynaspida. (modified from Johnston, 1968)
48 pnstemal ptatBht
\ /
pon
Figure 3.4: Pergamasus crassipes (Linnaeus) (Parasitidae), showing prestemal platelets, metastemal shields (MT), and postanal seta {pon). For the diAerence between latigynial and metastemal shields, see the text, (modified from Johnston, 1968) 49 latigynial shields
Figure 3.5: Promegisttis armstrongi Womersley (Promegistidae), showing pseudostemogynium (PSG; = pseudostemum). Each metastemal shield (MT) is fused to the endopodal element.
Often, in the females of Astemoseiidae, Seiodidae, Fedrizziidae, Klinckowstroemiidae, and Paramegistidae, there appears a shield, bearing only a paired stp3 (but with no seta), in the midstemal or seemingly pregenital region (Figure 3.1). This type of shield, often rhombic in shape, is referred to as the stemogynial shield (= stemogynium), and appears as a pair in Paramegistidae (Figure 2.2). A similar structure, but lacking any seta or lyrifissure, is found in Aenictequoidea, Promegistidae, and Parantennulidae, and is called pseudostemogynium (= pseudostemum) (Figure 3.5). I assume that pseudostemogynium may often be fused to stemogynial shield bringing an elongated stemogynial-pseudostemogynium complex. However, when the pseudostemogynium (= pseudostemum) is present alone, its origin is
50 thought to be genital rather than stemal (because of the lack of setae or lyrifissure). This pseudostemogynium is thought to be homologous to the pregenital shield in Holothyrida. Both of these structures (i.e., stemogynial shield and pseudostemogynium) lack any form of the setae and, therefore, are distinguished from metastemal shield(s), bearing both st4 and stp3.
From this new interpretation on the stemal region of the Mesostigmata, with few exceptions, such as the hypertrichy in stemal region (or venter) or highly modified stemal region of the female (as in Heterozerconina), the following rules are proposed. Exceptions to these rules in Mesostigmata are rare, but are applied to Heterozerconidae and Discozerconidae, where stemal elements are fractured and fused to endopodal elements.
1. Stemal 'region' is composed of the combination of three different sclerotized shields. From anterior to posterior they are, prestemal, stemal, and metastemal shields.
2. Stemal region' retains 4 pairs of setae (sf) a n d /o r 3 pairs of lyrihssures (stp).
3. If stemal 'shield' has 4 pairs of setae and/or 3 pairs of lyrihssures, then neither prestemal nor metastemal shield(s) is present (Figure 3.2). Prestemal or metastemal platelets, devoid of any seta or lyrifissure, however, may be found, {e.g., Arctacaridae, Zerconidae, Rhodacaridae, Ologamasidae, etc.)
4. If stemal shield has 3 pairs of setae and/or 2 pairs of lyrifissures, then either prestemal or metastemal shield(s), bearing additional pair of setae and/or lyrifissures, is present (Figures 3.3 and 3.4). However, in this case, if neither prestemal nor metastemal shield(s) is present, then stl or st4 may be on soft cuticle (Figure 3.6). 51 Figure 3.6; Antennoseius delicatus Berlese (Ascidae). st4 are on the soft cuticle, (modified from Johnston, 1968)
5. If stemal shield has 3 pairs of setae along with 1 pzdr of lyrifissures, then both prestemal, bearing stl, and a stemogynial shield, bearing stp3 are present (Figure 3.1).
6. If stemal shield has 2 pairs of setae and/or only 1 pair of lyrifissures, then both prestemal and metastemal shields are expected to be present. Rare exceptions may be found when stl, stpl, st4, and stp3 are located on the soft cuticle. Such possible exception, however, has not been found yet.
52 Definition of Genital Region
The genital region of the mites that belong to Mesostigmata shows remarkable sexual dimorphism. As stated above, the area posterior to the stemal structures constitutes the beginning of the genital region of the female. Often, this area is identified by the presence of st5 (and st6). Like the stemal region, the genital region is also composed of several shields that collectively cover the genital opening, which serves as the ovipore of the female. In male, circular genital opening, covered by a single or two valves, is either midstemal {Le., located between coxae in or IV) (Figure 2.7) or prestem al {i.e., located beneath the base of tritostemum) in position.
In female Mesostigmata, three major types of genital shield system are present based on the structure and function of the genital shields.
The first system, which 1 tentatively call 'hologynaspid' system, appears in Diarthrophallina and Uropodina. In this system, females have one complete horseshoe- or often inverted U-shaped genital shield (currently known as epigynial shield) that covers the female genital orifice (Figure 3.7). The whole shield itself is hinged at the posterior margin and is opened when she delivers her eggs. Although this type of hologynaspid shield is easily identified by its unique shape, it is also recognized by the lack of any stemal setae (stl-st4) and lyrifissures {stpl-stp3) on this shield, which are confined to the stemal region. A paired st5, defining the genital area, are located mostly on endopodal elements or rarely on the epigynial shield (as in Protodinychus). In Trachytes (Polyaspidoidea, Trachytidae) of Uropodina, each lateral margin of epigynial shield can be folded and opened during the oviposition, a condition similar to that in Trigynaspida. In the hologynaspid system, metastemal shields are not present. 53 5
Figure 3.7: Dinychus sp. (Uropodidae), showing 'hologynaspid' genital system, (aher Krantz, 1978)
The second type of genital shield system is known as the trigynaspid' system, which is found in Trigynaspida and Trachytes (Uropodina: Polyaspidoidea: Trachytidae). In this case, females typically have three genital shields, i.e. a mesogynial and paired latigynial shields. In a typical trigynaspid condition, mesogynial shield, hinged to its posterior margin, is often large and triangular and the latigynials, hinged to their outside lateral margin, are
54 elongate triangular or often bar-shaped (Figure 2.2). Like in the hologynaspid genital system, none of these meso- and latigynial shields retain stl-sté or stpl-stpS. Except for Micromegistus (Paranteimulidae), genital setae st5 (and st6) are always located on the latigynial shields. Although these three shields are opened to contribute oviposition, the trigynaspid system is thought to be more efficient in egg delivery compared to hologynaspid system in the context of evolution. Variations from this ancestral trigynaspid condition, however, commonly occur throughout Trigynaspida.
The third type of genital shield system — females have rather a small membranous genital orifice formed by the anterior extension of a single epigynial shield — is tentatively called 'agynaspid' system, and is conunon in the remaining groups of Mesostigmata, such as Zerconina, Epicriina, Arctacarina, and many Dermanyssina (Figures 3.2, a and 3.6). In Parasitina and in some Dermanyssina, such as Macrochelidae, the epigynial shield is opened during oviposition, rendering retrogressive hologynaspid genital system (Figure 3.4). In these cases, however, this system is easily distinguished by the presence of paired metastemal shields, which are absent in hologynaspid system. Setae st5 are usually located on the epigynial shield. Along with the hologynaspid system described above, the agynaspid system constitutes the current concept of 'Monogynaspida'.
In agynaspid system, the latigynial shields of the trigynaspid system are thought to be fused with the mesogynial shield or rarely with the endopodal elements, and the mesogynial shield of trigynaspid system per se remains as 'epigynial' shield. This is evidenced by the presence of st5 on this epigynial shield. In the context of evolution, agynaspid system is thought to be the most derived condition compared to hologynaspid system, as it allows the least loss of water (or body fluid) and it might be the least vulnerable from the potential attacks of parasites during the oviposition. In all of these genital systems 55 described above, oviposition. occurs at the anterior end of mesogynial (Le., epigynial) shield.
With regard to these interpretations on the genital systems of Mesostigmata, it should be mentioned that it was the Swedish acarologist Ivar Trâgârdh (1946a) who first applied the terms of 'Agynaspida' and 'Eugynaspida' into the classification of Mesostigmata. Indeed, he speculated that the Mesostigmata is composed of dichotomous lineages, each represented by the members that belong to one of these two groups. However, his interpretation is different from what I present herein, as his Agynaspida does not include any of Dermanyssina, the very typical agynaspid mite group. In addition, he assigned Megisthanidae of Trigynaspida into Agynaspida, while assigning current Celaenopsoidea (Celaenopsina sensu Trâgârdh, 1946a), Cercomegistidae, Paramegistidae, Antennophoridae, and Fedrizziidae into Eugynaspida. If his interpretation on genital system had been correct, all of these 'Trigynaspids' should have belonged into the one same grouping of, possibly, his Eugynaspida.
Definition of Ventral or Ventrianal Region
The area posterior to the genital region constitutes the ventral or ventrianal region. In Trigynaspida, the posterior margins of the latigynial shields or mesogynial shield are often fused to the ventral shield. In Heterozerconina, paired disc- or pad-like structures appear in this region. Although these discs are thought to be used as the measures for the attachment to the host's integument, detailed adhering mechanism is not known yet. Interestingly, however, each side of these discs in Heterozerconidae retains a small shield, bearing at least st4 along with st5 and st6, implying that the sternal and genital regions of this group of mites may
56 have been extended to posterior region compared to other mesostigmatid mites.
Parallel to the ventral shield, strap-like ventromarginal shields are often present along the length of the venter in Celaenopsoidea and some Uropodina (Figure 3.3). Free metapodal shields, located posterior to the coxae IV, if present, also appear in this region.
While cercomegistines always retain a ventrianal shield in the adult stage, some antennophorines, such as Megisthanoidea, Fedrizziidae, Klinckowstroemiidae, Parantennulidae, Euzerconidae, and Triplogyniidae, have a free anal shield, separated from the ventral shield. In Fedrizziidae and Klinckowstroemiidae, the separation is weak and only a thin line of separation is often present (Figure 3.1).
The anal opening, composed of two anal valves, retains a pair of euanal setae in the larvae in most Parasitiformes sensu lato, including Trigynaspida. These setae are lost in subsequent stases in Trigynaspida, but often the same area is occupied by a pair of lyrifissures. While most mesostigmatid mites carry a postanal seta (pan) posterior to the anal opening throughout all the stases (Figure 3.2), Trigynaspida, along with Diarthrophallina and some Uropodina, retain this seta only in the larval stase.
57 CHARACTER SELECTION, DESCRIPTION, AND CODING
While few characters from the immatures were selected as these immatures are very poorly known for most of Trigynaspida, those from the females constitute the majority of the data set. A total of 55 morphological features drawn from all the different body parts, which allow the assessment of structural homologies, were selected as the characters. These include the characters from dorsum, mouthparts, legs, and those from the venter, such as sternal, genital, ventral, and anal regions. In using body and leg chaetotaxy, each seta was regarded as a single character.
Morphological features that were used as characters for this study, their brief descriptions throughout taxa, and their coding schemes are as follows. Definitions of the terminologies for the sternal and genital regions are given in the section, Bauplan of Trigynaspida', described above. A Taxon-Character matrix for the analyses is shown in the Table 3.2.
I. Number of dorsal shields in adult
[0] 3 or more (podonotal + mesonotal + pygidial) [1] 2 (podonotal + opisthonotal) [2] 1 (holodorsal)
Among the members of Trigynaspida, antennophorines usually have a holodorsal shield, while most cercomegistines, excluding Astemoseiidae and Saltiseiidae, retain two or more shields in the adult. More than two dorsal
58 shields in the adult are found in Davacaridae, which retains four (single podonotal, weakly paired mesonotal, and a pygidial shield) and in Neotenogynium malkini (Neotenogyniidae), a paedomorphic trigynaspid that shows 3 dorsal shields in the female (Kethley, 1974). It should be noted that an undescribed new genus of Neotenogyniidae from Brazil carries a holodorsal shield in the female. Outside of the Trigynaspida, presence of more than two dorsal shields is found in Sejina, Microgyniina, and some Uropodina (e.g., Trachytes retains four dorsal shields).
Kethley (1977a) showed two dorsal shields in antennophorine Philodanidae. However, re examination of the holotype and paratype specimens revealed that the female philodanid retains a single shield with a vestigial line of fusion. The male shows an immaculate holodorsal shield.
2. Dorsal setae
[0] non-holotrichy [1] holotrichy
While holotrichous setation in dorsum allows setal notations in each setae on dorsum (Evans and Till, 1965; Lindquist and Moraza, 1998), non- holotrichous setation in dorsum indicates that such identifications are not possible.
With an exception of Pyrosejidae, all the cercomegistines show hypertrichy (i.e., a form of non-holotrichy). This hirsute situation is also found in antennophorine Philodanidae, Antennophoridae, Aenictequoidea, and most Megisthanoidea. Paramegistidae and Celaenopsoidea, however, show holotrichous (non-hypertrichous) state.
59 3. Idiosomal setae fUnorderedl
[0] dorsal and ventral setae nearly same in shape (and length) [1 j dorsal setae are longer or stout [2] ventral setae are longer or stout
This is a variable character within Trigynaspida. Although nearly similar length and shape of setae on the body are found in general, often, especially in early stases, such as larvae or protonymphs, or in the beetle- associated mesostigmatid mites, longer dorsal setae are found. In Ophiomegistiis, often foliate or stylus-like, or stout subulate setae are found in venter (Goff, 1979).
4. Setae on venter
[0] simple, setiform, smooth [1] barbed, or foliate, lanceolate, stylus-like
While simple setae are often short and straight, barbed ones are usually longer and often curved.
5. Spinous marginal setae
[0] absent [1] present
Trigynaspid members of Parantennuloidea, Promegistidae, Paramegistidae, Aenictequoidea, and Antennophoridae along with Discozerconidae, an outgroup, carry spinous or subulate setae along the membranous margin of the idiosoma. These setae are located in the lateral end of the body, and are not homologized with the setae of dorsum or venter.
60 6. Gnathotectum
[0] with anterior projections or serrations [1] without anterior projections or serrations
Gnathotectum is the structure covering dorsal area of the gnathosoma, and is also called epistome sensu Berlese (1900) or tectum sensu Snodgrass (1948). While the presence of, often multiple, anterior projections in the gnathotectum is observed in many groups of Mesostigmata, this character was once regarded as a good diagnostic character for the entire Cercomegistina (Figure 3.8) within Trigynaspida (Kethley, 1977b). Within Cercomegistina, however, Seiodidae lack such projections (Figure 3.9). The members of Antennophorina, excluding Philodanidae, do not show multiple anterior projections.
7. Gnathotectum
[0] not triangular, somewhat roundish [1] triangular
8. M edian keel of gnathotectum
[0] absent [1] obscure [2] present
Gnathotectum often has a roof-like structure, showing a median keel like line, which can be seen dorsally and ventrally. Ventral keels are often more prominent than those observed dorsally.
The presence of median keel in gnathotectum was once regarded as a synapomorphy for the entire Antennophorina. However, detailed observation reveals that this character is variable, and conspicuous in Celaenopsoidea, Megisthanoidea, and Promegistidae within
61 Figure 3.8: Cercoleipiis sp. (Cercomegistidae). Gnathotectum.
(a) (b)
Figure 3.9: Seiodes ursinus Berlese (Seiodidae), (a) female and (b) maie gnathotectum.
62 Antennophorina. This structure is absent in Parantennuloidea and some Paramegistidae along with the Cercomegistina. When it is obscure, it only has a thin longitudinal line as in Klinckowstroemiidae, Fedrizziidae, Aenictequoidea, Antennophoridae, and the remaining Paramegistidae.
9. Chelicerae
[0] elongate [1] not elongate
Elongate cheliceral segments are typical to Uropodina, in which all of its members retain long and slender cheliceral segments with much shorter and smaller digits (Figure 3.10). In many uropodines, the length of the entire chelicerae is more than a half or close to 3/4 of the idiosomal length. In other groups, including Trigynaspida, Sejina, Heterozerconina, and Microgyniina, their chelicerae are shorter, reaching at best less than the half of the length of the body. In these groups, the cheliceral digits are nearly equal to the length of the cheliceral segments.
10. Cheliceral digits
[0] tapered, often edentate (or with numerous minute teeth) [1] robust, with large proximal tooth on digits
This character is thought to be related to the kinds of food that the mites uptake. In other words, while the tapered and often edentate cheliceral digits are used for the ingestion of liquid or soft food, those of robust and often dentate ones are thought to be used to grab more solid food. The robust and dentate chelicerae are often used for the phoresy by holding the host's body part, such as seta (Evans and Hyatt, 1963). Moreover, it is assumed that the mites with tapered and edentate ones are not predatory or more or less paraphagic, while those with robust ones are more or less predatory (Kinn,
63 1971). In. Tri^m aspida, Parantennuloidea,. Promegistidae^ Paramegistidae,. Aenictequoidea, and Antennophoridae have tapered and edentate digits (Figure 2.3, d). All the Cercomegistina along with the antennophorine Megisthanoidea, Fedrizziidae, Klinckowstroemiidae, and Celaenopsoidea have robust and dentate digits.
(a) (b)
Figure 3.10: (a) Long and slender cheHcera of Uropodoidea. Cheliceral excrescences are absent, (b) Short and robust chelicera of Celaenopsoidea. Excrescences are dendritic, (after Krantz, 1978)
64 11. Cheliceral excrescences
[0] reduced or absent [1] dendritic (or brush-like) [2] filamentous (hypertrophied)
Figure 3.11: Klinckowstroemia sp. (Klinckowstroemiidae). (after Krantz, 1978)
Cheliceral excrescences are thought to be homologous to the arthrodial brushes that are often observed at the proximal region of the movable digit of chelicerae in many Mesostigmata, including, but not limited to, Microgyniina, Heterozerconina, Dermanyssina, and Parasitina. In Trigynaspida, Cercomegistoidea, Megisthanoidea, and Celaenopsoidea show dendritic or brush-like forms (Figure 3.10, b), while the remaining groups, such as Parantennuloidea, Promegistidae, Paramegistidae, Aenictequoidea, Antennophoridae, Fedrizziidae, and Klinckowstroemiidae show more or less filamentous forms (Figure 3.11). Uropodina and Sejina lack this character on their chelicerae.
65 12. Location of cheliceral excrescences
[0] proximal [1] middle or mediodistal
In most Cercomegistoidea, Parantennuloidea, Promegistidae, and Celaenopsoidea, the cheliceral excrescences originate from the proximal region of the movable digit of the chelicera. This contrasts with the condition in Fedrizziidae, Klinckowstroemiidae (Fedrizzioidea), Megisthanoidea, in which they arise from the mediodistal region of the movable digit (Figure 3.11).
13. Com iculi
[0] strong, stout at base, heavily sclerotized [1] not heavily sclerotized, sometimes membranous, or almost seta-like
The comiculi are also known as the external malae, which are the parts of the endites of the coxa of the pedipalps in Mesostigmata (Evans, 1992). The analogous structures are the rutella appearing in acariform Sarcoptiformes. In Trigynaspida, all the members of Cercomegistina along with antennophorine Fedrizziidae, Klinckowstroemiidae, Megisthanoidea, and Celaenopsoidea retain strong and heavily sclerotized comiculi (Figure 3.12, a, e). In contrast, the members of Parantennuloidea, Promegistidae, Paramegistidae, Antennophoridae, and Aenictequoidea show somewhat membranous or almost setiform comiculi (Figure 3.12, b, c, d). As typical to Mesostigmata, all the outgroups have heavily sclerotized comiculi in the adult.
14. Com iculi
[0] smooth, not branched, hom-like [1] 2-tined [2] 3- or more tined, somewhat serrate 66 While retaining smooth and hom-like comiculi is typical to the Mesostigmata, often 2-tined (bifed; as in Triplogyniidae, Meristomegistus (Paramegistidae; see Figure 2.3, b), etc.) or multi-tined (serrate; as in Antennophoridae, Euzerconidae, etc.) comiculi are present. A
(a) (b) (c) (d) (e)
Figure 3.12: Comiculi. (a) Fedrizzia sp. (Fedrizziidae); (b) Paramegistus confrater Trâgârdh (Paramegistidae); (c) Echinomegistus wheeleri (Wasmaim) (Paramegistidae); (d) Antennophoriis sp. (Antennophoridae); (e) Euzercon latus (Bauaks) (Euzerconidae)
15. Subcapitular setae
[0] barbed (coarsely (heavily) or lightly) [1] smooth
Subcapitular setae sensu Evans and Till (1965,1979) and Krantz (1978) are also known as palpcoxal setae sensu Evans (1992); gnathosomal setae sensu Funk (1968) and Ghilyarov and Bregetova (1977); deutostomal setae
67 sensu Nickel and Elzinga (1970b). While smooth setae are simple and with no serrations, barbed ones are bipectinate.
16. Hypostomal setae
[0] form a straight line [1] not form a straight line, angled
Linear arrangement of hypostomal setae along the axis of the body has extensively been used to diagnose Uropodina and Diarthrophallina. Although this is true for these groups, in Trigynaspida, Saltiseiidae, Philodanidae, Klinckowstroemiidae, Fedrizziidae, Hoplomegistidae, and Megacelaenopsidae also show linear arrangement of these setae.
17. Palp genu
[0] with 5 setae [1] with 6 setae [2] with 7 setae
With few exceptions, presence of 7 setae on palp genua in Antennophorina is unique in Mesostigmata. These 7 setae are composed of 3 dorsal (d), 2 anterolateral (al), 1 posterolateral (pi), and 1 ventral (v) seta. Exceptions to this in Antennophorina are found in Micromegistus bakeri (Parantennulidae), which has only 5 (by lacking a ventral and an anterolateral seta), and in M. gourlayi (Parantennulidae), Philodanidae, Promegistidae, Echinomegistus (Paramegistidae), and Neotenogyniidae, all of which have 6 setae as in Cercomegistina. In outgroups, while most of Uropodina, excluding Thinozerconidae, retain 5 setae on this palp segment, most mesostigmatids have 6 setae by lacking a ventral seta.
68 18. Palp tibia and tarsus
[0] completely articulated [1] slightly (or partially) fused or fused but with line [2] (insensibly or completely) fused
(a) (b)
Figure 3.13: (a) weakly fused {Paramegistus) and (b) completely fused (Ophiomegistus) palp tibia and tarsus.
When the palp tibia and tarsus are completely articulated, the diameter of the base of tarsus usually becomes smaller than the diameter of the distal end of tibia, and the intersegmental membrane is observed at the ventral side of the palp (Figure 2.3, c). As to the fused tibia and tarsus, there is no such separation, and the membrane or the line of separation is not present (Figure 3.13, b). In this case, the presumed area of palp tarsus can be estimated by the location of apotele, which is located in the base of palp tarsus in Mesostigmata. Often, this separation or fusion between these two palpal
69 segments is obscure, but bearing a thin line of separation between the two segments (Figure 3.13, a). When they show this feature, the entire palp tibia and tarsus become tapered, and no intersegmental membrane is present.
19. Palptarsal claw (= apotele)
[0] 2-tined [1] 3-tined
Within the group Trigynaspida, all the members of Cercomegistina (Cercomegistoidea) carry a 3-tined palptarsal claw, while those of Antennophorina usually have a 2-tined one. Exceptions to this in Antennophorina are observed from Parantennulus (not included in this study) and Micromegistus (Parantennulidae), Megisthanidae, and Megacelaenopsidae, all of which have 3-tined palptarsal claw (Figure 3.14).
Figure 3.14: Megacelaenopsis sp. (Megacelaenopsidae). Palp tarsus with 3-tined apotele.
70 20. Tritostem al laciniae
[0] free, separated (occasionally fused in the proximal portion; base of tritostemum often wider than long) [1] fused (occasionally separated terminally; base of tritostemum often longer than wide)
In his key to the families, Kethley (1977b) recognized the fused tritostemal laciniae as one of the main characters for Cercomegistina (Figure 3.15). However, exceptions are found in three cercomegistine families, such as Astemoseiidae, Davacaridae, and Seiodidae, which retain free tritostemal laciniae (Figure 3.18). Along with this character, Seiodidae also show smooth edge of gnathotectum, which is atypical to Cercomegistina. All the members of Antennophorina and outgroups, except for Polyaspis and Oplitis, retain separated (free) tritostemal laciniae.
Figure 3.15: Cercoleipus sp. (Cercomegistidae). Fused tritostemal laciniae.
71 21. Presternal shield (in female)
[0] paired [1] entire (not paired) [2] absent
See the Introduction to this Chapter (also see Figures 3.1, 3.5, 3.16 and 3.17).
22. Sternal shield (in female)
[0] entire (not fragmented; fused) [1] fragmented, divided, paired, not entire
As for this character, the presence of presternal or metastemal shield(s) is not regarded as the fragmented, paired, or divided sternal shield. Instead, this character refers symmetric separation (i.e., pairing) of the sternal shields along the median line of the body (Figure 2.2) (For the definition of the sternal shield, see the Introduction to this Chapter.). Fragmented or paired sternal shields appear in many Cercomegistina, Paramegistidae, Antennophoridae, and Sejina, Microgyniina, and Heterozerconina.
In Microgyniina, two sternal shields, each bearing two setae of stl-stl and st3-st4, are separated longitudinally. As each shield retains two pairs of sternal setae, it is assumed that these shields are neither presternal nor metastemal but simply separated stemals. In Heterozerconina, the stemal structures, bearing stl-st4 and stpl-stpS, are separated. While the bar-shaped stemal shield is located in its original location, the remaining structures, identified by retaining stemal lyrifissures (stp), are fused to the endopodal elements.
72 23. Stemovaginal sclerites
[0] absent [1] present
The Aenictequoidea has a pair of posterior internal stemal shield extensions, called the stemovaginal sclerites (Figure 3.16). As these intemal structures are originated from the stemal elements, they cannot be homologized with the vaginal sclerites.
Figure 3.16: Ptochacarus silvestrii Womersley (Ptochacaridae), showing paired stemovaginal sclerites.
73 24. Stem al shield
[0] bearing metastemal setae [1] not bearing metastemal setae
In Mesostigmata, metastemal setae refer to the 4th pair of the stemal setae (st4). While the setae st4 on stemal shield is common (Figures 3.1, 3.2 and 3.16), often these setae are located on soft integument or on separate metastemal shields outside the stemal shield (Figures 3.3, 3.4, 3.5, 3.6 and 3.17). When the setae st4 are on stemal shield, metastemal shield is expected to be absent (see the 'rules' described in the Introduction). However, presence of the st4 outside the sternal shield does not necessarily mean that the metastemal shield is present as the st4 are often located in soft cuticle.
25. Stem al setae 1 (stl)
[0] on integument [1] on stemal shield [2] on prestemal shield
See the definition of stemal area described in the Introduction to this Chapter, (also see Figures 3.1-3.6)
26. First stem al setae (stl)
[0] smooth [1]barbed
27. Second stem al setae (st2)
[0] smooth [1] barbed
7 4 28. Third stemal setae (st3)
[0] smooth [1] barbed
29. Fourth stemal setae (sM)
[0] smooth [1] barbed
30. Metastemal shield
[0] absent [1] entire, fused together [2] paired, divided
In this study, metastemal shield is defined as a shield bearing st4 and stp3 in adult. This is compared with stemogynial shield (stemogynium) which bears stp3 only, and with latigynial shields, each of which often bears Sts or st6. In many mesostigmatid mites, metastemal shield, if present, usually appears as a pair (Figure 3.3). In some Celaenopsoidea, this shield is thin and transversely narrow, and is often weakly connected together by a bridge (Figure 3.17).
31. Posterior end of stemal element
[0] not fused with endopodal elements [11 fused with endopodal elements
In most mesostigmatid mites, stemal elements, including prestemal, stemal, or metastemal shields, if present, are free and distinct Often, stemal or metastemal shields are elongated and fused to endopodal elements as in Micromegistus (Parantennulidae), Promegistidae, and Aenictequoidea. In Promegistidae, the metastemal shields, bearing st4, are extended posteriorly (Figure 3.5), and in Aenictequoidea, the posterior sides of stemal shield are
75 MT
Figure 3.17: Choriarchus reginits Kinn (Schizogyniidae), showing fused metastemal shield (MT). Heads of vaginal sclerites are well-developed.
extended. This structure, however, is not intemal, and therefore, is distinguished from stemovaginal elements.
32. Stemogynial shield
[0] absent [1] entire (not divided; fused) [2] divided, paired
76 Stemogynial shield (stemogynium^ i& defined as a shield bearing stpS only (Figures 2.2 and 3.1). When a shield retains additional st4, it is defined as metastemal shield (Figures 3.3 and 3.4). A stemogynial shield appears in Astemoseiidae and Seiodidae (Figure 3.18) in Cercomegistina, and Klinckowstroemiidae, Fedrizziidae, Paramegistidae, and Hoplomegistidae in Antennophorina. Despite the fact that Kethley (1977b) used this character to identify his Fedrizzioidea that includes Klinckowstroemiidae, Fedrizziidae, Paramegistidae, and Promegistidae, Promegistus (Promegistidae) lacks this character. Paramegistidae and Hoplomegistidae are diagnosed by the presence of paired stemogynial shields (Figures 2.2 and 3.19).
33. Pseudostemogynium (= pseudostemum sensu Kethley, 1977b)
[0] absent [1] present
A pseudostemogynium sensu Krantz (1978) is defined by a shield or a sclerite located between stemal and genital region that lacks any seta or lyrifissure. This structure is, therefore, distinguished from stemogynial shield (stemogynium) that retains stp3. The pseudostemogynium is rhombic in Promegistidae or rectangular in MjcromegisfMs(Parantennulidae). As this structure lacks any seta or lyrifissure, which delimits the stemal region, it is thought to be the genital in origin, and is thought to be homologous to the pregenital shield in Holothyrida. It is found in Saltiseiidae, Parantennulidae, Promegistidae, and Aenictequoidea in Trigynaspida (Figure 3.5).
34. Vaginal sclerites
[0] absent [1] present, but head reduced [2] head well-developed
77 N: £
Figure 3.18: Seiodes ursinus Berlese (Seiodidae), female sternal-genital region.
78 Vaginal sderites, whidt axe thought ta be the internal genital structures that support egg delivery, are well-developed in Celaenopsoidea and Megisthanoidea (Figure 3.17). Often, the heads are vestigial and arms look like thin straps in other groups of Trigynaspida, including Parantennuloidea, Promegistidae, Paramegistidae, Aenictequoidea, Antennophoridae, Klinckowstroemiidae, and Fedrizziidae (Figure 3.19). Similar structures are also found in uropodine mite, Polyaspinus migginsi Camin. Cercomegistina and other outgroup members used for this study lack this character.
Figure 3.19: Paramegistus confrater Trâgârdh (Paramegistidae). Weakly- developed (or vestigial) vaginal sclerites are shown in dotted line. Stemogynial shields are paired.
79 35. M esogynial shield
[0] absent [1] reduced, obscure, or fused with ventral (or ventrianal) shield [2] present (distinct, free, well-developed)
In Cercomegistina, the mesogynial shield is free, usually larger than that of Antennophorina, and mostly triangular. This structure becomes variable in Antennophorina, in which the shield is triangular (as in Aenictequidae or Ptochacaridae), rectangular (as in Micromegistus (Parantennulidae)), reduced (as in Philodanidae, Diplogyniidae, or Triplogyniidae), or completely fused to ventral or ventrianal elements (as in Fedrizziidae, Megisthanoidea, Euzerconidae, Celaenopsidae, or Megacelaenopsidae).
36. M esogynial shield
[0] triangular or subtriangular [1] rectangular, subrectangular, or bar-shaped
37. Latigynial shield(s)
[0] reduced or incorporated in other shield elements [1] two shields fused together [2] present, distinct, free, well-developed
Retaining the functional latigynial shields that are opened during oviposition is often characteristic to female Trigynaspida. These shields are absent in other mesostigmatid mites, excluding Trachytes of Uropodina, but are thought to be fused to the endopodal elements in Uropodina (Protodinychus is an exception) and Diarthrophallina. In higher mesostigmatids, such as Dermanyssina, latigynial shields are thought to be fused to the epigynial shield. As to this character, however, the presence of
80 endopodal elements^ whicltare not related to ov^osition, is not regarded as the same as the presence of latigynial shields, and are coded as 0 state.
When the latigynials are present as free and paired, they are usually triangular (as in most Cercomegistina, Promegistidae, Paramegistidae, Diplogyniidae, etc.) in shape, but (sub)rectangular (as in some Aenictequoidea) or rare roundish form (as in Philodanidae) is also present. Often two shields are fused together to form a single shield as in Hoplomegistidae. In addition, in some Celaenopsoidea, such as Euzerconidae, Celaenopsidae, or Megacelaenopsidae, these shields are completely fused to ventrianal shield (Figure 3.3).
As for the outgroups, uropodine Trachytes shows paired latigynial shields which seems to be the functional at oviposition. No such latigynial shields is known in other Mesostigmata.
38. Ventromarginal shields
[0] absent (fused with peritrematal plate) [1] present (free from peritrematal plate)
See the Introduction to this Chapter for the definition of ventromarginal shield (see Figure 3.3).
39. Anal or ventrianal shield
[0] fused with dorsal shield [1] not fused with dorsal shield
40. Anal shield (in female)
[0] (insensibly) fused with ventral shield (ventrianal shield) [1] contiguous with ventral shield [2] present, free, not fused with ventral shield 81 Anal shield is defined as a free shield bearing anal opening. When this shield is not free, it is fused to ventral shield to form a ventrianal shield. In Trigynaspida, all the Cercomegistina lack free anal shield. In Saltiseiidae, weakly developed sclerotization around anal opening is present (Walter, 2000), this sclerotization, however, is embedded into the ventral element covering the entire ventrianal area. Fedrizziidae and Klinckowstroemiidae have a thin line between ventral and anal area.
41. Anal discs ('suckers')
[0] absent [1] present
This enigmatic character defines Heterozerconina, including Heterozerconidae and Discozerconidae. In these two families, paired circular pad-like structures appear in ventral-anal elements. Although it is assumed that these pads offer tighter attachment to the host's integument, detailed mechanism is not known yet.
42. Postanal seta ipon) (in adult)
[0] present [1] absent
43. Pedofossae (= foveae pedales sensu Krantz, 1978)
[0] present [1] absent
Pedofossae are the depression of the cuticle around the base of coxae IV, which are thought to be used for the resting of the legs IV. This character is observed in Klinckowstroemiidae and Fedrizziidae and in Uropodoidea of Uropodina (outgroup). 82 44. Peritreme
[0] well-developed [1] (greatly) reduced or peritreme-like groove (pseudoperitreme) present
While normally developed peritremes reach almost to the base of coxae I or to the base of gnathosoma, reduced peritremes reach only to the area around / between coxae II-UI. In Trigynaspida, Parantennulidae and ant associates of Aenictequoidea and Antennophoridae show short peritremes.
45. Tarsus I
[0] with claws [1] without claws
In Trigynaspida, all the members of Antennophorina lack claws on tarsus I (Figure 2.4, b). This character is shared with cercomegistine members of Cercoleipiis and Cercomegistus (Cercomegistidae), undescribed new genus of Pyrosejidae, and Saltiseiidae, along with uropodine outgroup, Polyaspis (Polyaspididae).
46. al3 on Tarsus II
[0] present [IJ absent
Unlike Monogynaspid mites, trigynaspids retain an additional anterolateral seta (al; alx in Evans, 1965,1969) on tarsi II-IV in adult, yielding a total of 4 al setae on these leg segments. Although this seta is identified as al3 in the adult, in the context of the sequence of ontogeny this seta appears as the fourth one.
83 47. a/3 on Tarsus III
[0] present [1]absent
Although this character is similar to the character #46, character independence is assumed as the setae (characters) are not located in the same leg. Hence, these are two different characters on two different locations. This character becomes synapomorphic to the Trigynaspida.
48. Femur IV
[0] with 8 setae [1] with 7 setae [2] with 6 setae
When the femur IV in the adult bears 8 setae, it has a chaetotaxy of 1- 2/1-2/1-1 (Figures 1.1 and 2.5, c). The posterolateral (pi) seta becomes absent when this leg segment has a total of 7 setae; and the seta pi and posteroventral (pv) seta are both absent if the femur IV has 6 setae.
49. Tarsus IV
[0] with 4 al setae [11 with 3 or fewer al setae
50. Ventral intercalary sclerite on tarsus IV (adult)
[0] free, well-developed [1] weakly developed [2] absent
The presence of this sclerite, bearing av4 and pv4 on tarsus IV in adult, is shared by the Sejina and most of the Trigynaspida, excluding Celaenopsoidea. In Celaenopsoidea, the sclerite is partially fused to telotarsus IV. In Epicriidae (not used for this study), a similar sclerite is present at the 84 circumsegmental region of tarsus IV. It should be noted that this sclerite is not ventral but dorsal in position, and bears ad3 and pd3. In all adult Trigynaspida and Sejina, circumsegmental region of tarsus ll-UI also bears this sclerite (Figures 2.5, a, b, c).
51. Proximoventral region of tarsus IV
[0] av4 and pv4 present [1] av4 and pv4 absent
All the Trigynaspida, including Celaenopsoidea, and Sejina retain paired setae of av4 and pv4 on ventral intercalary sclerite appearing in the circumsegmental fissure between basi- and telotarsus IV (Figure 2.5, c) in deutonymph and adult. In the protonymph, the same paired av and pv setae appear on a free intercalary sclerite. Although these setae are called av3 and pv3 as in Krantz (1978, Figure 47-6), they are thought to be homologous to av4 and pv4 appearing on such a sclerite in the deutonymph and adult. Presence of these setae outside the Trigynaspida and Sejina has not been reported in Mesostigmata.
52. Male genital opening
[0] midstemal (= located between H-FV) [1] prestemal (= located beneath the base of tritostemum)
In primitive mesostigmatid mites, its location is more or less posterior, located in an area between coxae 111 or IV (Figure 2.7). Most of the Trigynaspida along with the outgroups of Uropodina and Sejina retain this ancestral state. In Celaenopsoidea, the opening is located beneath the base of tritostemum, which is observed in many derived groups, such as Dermanyssina and Parasitina. Heterozerconina also bears prestemal position in male genital opening.
85 53. Male eugenital setae
[0] absent [1] present
Some trigynaspids carry paired eugenital setae on the male genital opening, a character shared with some members of Uropodina, such as Trachytes and Polyaspis. In Cercomegistina, this character is found in Astemoseiidae, Cercoleipiis (Cercomegistidae), and Davacaridae. antennophorine groups of Paramegistidae (excluding Echinomegistus), Aenictequoidea, Antennophoridae, Klinckowstroemiidae, and Megisthanoidea also retain this character (Figures 2.7 and 3.20). This is compared to Sejina, which lacks male eugenital setae.
54. Salivary styli
[0] present [1] absent
Trigynaspida does not retain this character while most other groups of Mesostigmata show this character (Camin and Gorirossi, 1955; Evans, 1992). When they are present, they often appear parallel to the labrum and are often associated with the comiculi.
55. Phoretic deutonymph
[0] present [1] absent
In Mesostigmata, phoretic deutonymphs are found in certain Uropodina (Uropodinae), some Sejina, Microgyniina, certain Parasitina (Parasitinae) and some Dermanyssina. The phoretic deutonymph often
86 Figure 3.20: Ptochacarus sp. (Ptochacaridae), maie stemogenital region showing paired eugenital setae.
entails a specialized body form, such as anal pedicel and anal gland in Uropodinae or modified anal region in Microsejus (Evans and Till, 1979), both of which offer the function of an attachment to the host. When these phoretic deutonymphs are associated with their host for dispersal, they are sessile and probably non-feeding. In Micromegistus (Parantennulidae) of Trigynaspida, all the postembryonic stages are found at a single beetle host. Although its deutonymphal stage is associated with its host, it is not regarded 87 as the phoretic deutonymph, because* along with other stases, those deutonymphs are always active and feed on fungi. Unlike phoretic monogynaspid mites that have a deutonymphal dispersal stage, trigynaspids, excluding Micromegistus, disperse as adults (Hunter, 1993a).
88 Taxon Character
Asternoseius 20100??011110011101020001000000100202010011000000000111 Cercoleipus 1000001011100001101101002111100000202010011010000000111 Cercomegistus 1010001011100001111121002111100000202010011010000000011 D avacarus 0000001011110001101000002000000000202010011000000000111 S e io d e s 1010011011100011101020001000000100202010011000000000011 P yro seju s 1110001011100011111101002000000000212000011000000000011 Pyrosejidae, n. sp. 1110001011100011111101002000000000212000011010000000011 S a ltis e iu s 2001000011110000101120011100020010202010011010000000011 Micromegistus bakeri 2100110010201211001020010100001011212012011110000000011 Micromegistus gourlayi 2100110010201011101020010100001011212012011110000000011 Philodana 200010101020101012002000? 00000? 0011? 2111011010000000011 Promegistus 21211112102010111000200110000210111?2010011010000000011 Antennomegistus 2121111010211011120001002000000201202010011010000? 00?11 Echinomegistus 2121111010211011120001002000000201200010011010000000011 Meristomegistus 2121111110211111200001002000000201112010011010000000111 Neomegistus 21201111102112012200010020000002011? 0010011010000000?11 Ophiomegistus 2121111010211011220001002000000201102010011010000000111 Paramegistus 2121111110211211210001002000000201202010011010000000111 Aenicteques 200111111021121122002010100000101120201001111000000??11 Ptochacarus 2000111110211211220000102000001011202010011110000000111 Antennophorus 2000111110211201200011002000000001172010011110000000111 Klinckowstroemia 2120011111210110200010002000000101202111010010000000111 K. v i c t o r i a e 2100011111210110200010002000000101202111010010000000111 F e d rizz ia 21000111112100? 02000100020000001010? 0111010010000000011 Neofedrizzia 2100011111210010200010002000000101070111010010000000011 Megisthanus 20000112111101112010100020000007 020?0012011010000000111 Stenosternum truitae 2000011211110100200000002000000202071012011010000000111 Stenosternum sp. 2100011211110110200000002000000202071002011010000000111 Pleuronectocelaeno 210001121110000120002001110002000207 0110011010000101011 Trichodiplogynium 2100011211110001200020011000010002102100011010000101011 Cryptometasternum 2100011211110001200020011000010002102100011010000101011 Ophiocelaeno 2100011211110001200020011000010002102100011010000101011 Passalacarus 2100011211110001200020011000010002102100011010000101011 Euzercon 212001121111020120002001111102000207 0112011010000101011 Neoeuzercon 212001121111020120002001111102000207 0112011010000101011 Megacelaenopsis sp. 210001121110000020102001100000000207 0100011010000107011 M. oudemansi 210001121110000020102000100000000207 010001101000010? Oil Paraschizogynium 2120011211100001200020011000010002110110011010000101011 Funkotriplogynium 2110011211100101210020011000000002102112011010000101011 Triplogynium 2100011211100101210020011000000002102112011010000101011 Thinozercon 00000010000?00101000100000000070000700100?1001121210?01 P o ly a sp is 111000100007 000000012000100000100007 0010001011121210100 T rach ytes 0110002 0000?0000000020001000001000072010011001111210101 O p l i t i s 210000? 0000? 00000001200010000010000? 0010000001111210? 00 Asternolaelaps 20000110110? 0101110027 00100000? 00007 0000001001111000001 Epicroseius 0? 07 0000110? 00011100210017? 000? 00007 0000001001111000000 Se ju s 07 000000110?000111002100100000? 00007 000000100111100000? U ropodella 01010100110? 0000120021001110002000071000001101111000001 Microgynium 01100010101? 001110002001000000? 00007 01000011011112107 00 Discozercon 210011001010001110000101210000100007 0110101001111211001 Heterozerconidae, n.sp. 2100010010100011100001012000001000070010101001111211001
Table 3.2: Taxon-Character matrix used for morphological study. ?: not available
89 RESULTS AND DISCUSSION
From the data set, three equally most parsimonious trees (L = 267 steps; Cl = 0.27; RI = 0.75) were deduced under no topological constraints. A strict consensus tree recovered monophyletic Trigynaspida, carrying the bifurcating lineages of Cercomegistina and Antennophorina (Figure 3.21). As to the higher relationships within Trigynaspida, Kethley's (1977b) superfamily Fedrizzioidea, containing Fedrizziidae, Klinckowstroemiidae, Promegistidae, and Paramegistidae, was not supported as this superfamily is not monophyletic. In addition, the phylogeny showed that the two ant associates, Antennophoroidea and Aenictequoidea, are not distantly related but monophyletic.
Regarding the status of Fedrizzioidea sensu Kethley (1977b), while two sister families of Fedrizziidae and Klinckowstroemiidae, passalid associates, form a monophyletic group and remain in this superfamily, Promegistidae and Paramegistidae require the separate new groupings. Promegistidae, represented by a single known species, Promegistus armstrongi Womersley that is associated with Pamborus (Coleoptera: Carabidae) in Australia, become the sister to the two species of Micromegistus (Parantennuloidea: Parantennulidae), carabid associates from North America and Australia. This new group, along with another known parantennuloid, Philodana johnstoni (Philodanidae), forms a clade of the revised superfamily Parantennuloidea. Unlike Fedrizziidae and Klinckowstroemiidae that retain entire (= impaired)
90 Aatemoseius Seiodes Cetcdeipus Cercomegistus Pyrosejus Cercomegistoidea Pyrosejidae n.sp. Davecerus Saltiseius Micromegistus iMkeri M. goudayt Promegistus Parantennuloidea Philodana Antennomegistus • 1 - ^ Echinomegistus Ophiomegistus Neomegistus PaiMiatistaidea Paramegistus Meristomegistus Aenicteques Ptochacarus Aenictequoidea Antennophorus Antennophoroidea Klinckovstroemia K. victoriae Fedrizzia Fedrizzioidea Neofedrizzia Megisthanus Stenosternum Megisthanoidea S.truittae Trichodiplogynium Crypt ometastemum Passalacarus Ophiocelaeno Pleuronectocelaeno Euzercon Neoeuzercon Celaenopsoidea Megacelaenopsis 1 Megacelaenopsis 2 Paraschizogynium Triplogynium Funkotriplogynium Thinozercon Polyaspis Oplitis Uropodina Trachytes Microgynium MkrogynHna Discozercon Heterozerconidae Heterozerconina Asternolaelaps Sejus Epicroseius Sejkw Uropodella
Figure 3.21: Strict consensus tree from 3 equally most parsimonious trees for comparative morphological data under no topological constraints. Numbers shown are the values of branch support sensu Bremer (1988). Names of the superfamilies are hrom this study.
91 stemogynial shield, heavily sderotized comiculi, and robust dentate cheliceral digits, Promegistidae retain no stemogynial shield, more or less membranous base of the comiculi, and tapered edentate chelicerae, which are characteristic of the members of Parantennuloidea.
The remaining family Paramegistidae, displaying the associations with diplopods, squama tes, or carabids, become monophyletic and share sister group relationships with the ant-associates, Antennophoroidea and Aenictequoidea. This new relationship suggests the removal of Paramegistidae from current Fedrizzioidea to form a new superfamily, Paramegistoidea. The breakdown of Fedrizzioidea and the new status for its members are summarized in the Table 3.3.
Current Fedrizzioidea Host Proposed Superfamily Fedrizziidae Passalids Fedrizzioidea Klinckowstroemiidae Passalids Fedrizzioidea Paramegistidae Millipedes, Reptiles, Paramegistoidea, NEW Carabids Promegistidae Carabids Parantennuloidea
Table 3.3: Proposed classification for Fedrizzioidea.
When the monophyletic Fedrizzioidea sensu Kethley (1977b) was enforced from the data set, 10 additional steps (i.e., 277 steps; 47 trees) were required. A strict consensus tree from these enforced trees, however, failed 92 not only to discover the hierarchy within Trigynaspida but to recognize monophyletic Cercomegistina and Antennophorina.
This study shows that the two current superfamilies of ant-associates, Antennophoroidea and Aenictequoidea, are not as distantly related as suggested by Kethley (1977b). Instead, they are sister groups, suggesting either uniting them in a single superfamily (of Antennophoroidea), or retaining two superfamilies as is by keeping their current names. These ant-associates are not that different in terms of morphology, as they share several characters, such as short peritremes, eugenital setae on male genital opening, and non- sclerotized comiculi.
Within the Antennophorina, reshuffled Parantennuloidea, including Promegistidae, branched first. Although the phylogeny from this study is different from that of Kethley's, this initial branching of Parantennuloidea in Antennophorina is consistent with the Kethley's. This group is supported by the characters such as the presence of 5 or 6 setae (not 7) on palp genua, the presence of a pseudostemogynium, and the absence of prestemal shield and male eugenital setae.
93 A n o te o n Paramegistus australis Banks. 1916
Banks (1916:230) reported this species from a single male associated with an ant, Polyrhachis hexacantha (Erichson, 1842), in eastern Australia, and assigned it into Paramegistidae. Kethley (1977b:142) recognized that this species does not belong to Paramegistus or even to Paramegistidae, and proposed a new genus, Pseudomegistus, to hold this species within Antennophorina of Trigynaspida. This species, however, bears only 7 setae on femur IV, by lacking pi seta (1-2/1-2/1-G), a character not present in Trigynaspida (see Figure 1.1, b). Accordingly, this species is removed from the Trigynaspida. As this species was described from a single male, and the specimen is in poor condition, the designation of the family of Mesostigmata for this species has not yet been made {incertae sedis).
94 A key to the families of Trigynaspida (based mainly on females) is provided below. The figure numbers written in bold correspond to those of Krantz (1978) otherwise specified as in Saltiseiidae, Pyrosejidae, Philodanidae, Triplogyniidae, and Costacaridae. New figures of the female venter and the gnathotectum of both sexes of Seiodidae have been supplied in Figure 3.9.
A Key to the Families of Trigynaspida (based on Female)
1. Often with two or more dorsal shields (if one, then often a line of fusion exists between anterior podonotal and posterior opisthonotal shields) (Figs. 45-2, 45-7). Gnathotectum usually with anterior projections or serrations, often not coming to point, and without median keel (Fig. 45-5) (if edge of gnathotectum smooth with pointed anterior end, then tritostemum with 2 distinct laciniae; as in Seiodidae). Palp genu usually with 6 setae. Palptarsal claw (= apotele) 3-tined. Chelicerae often robust, dentate or with large proximal teeth. Cheliceral excrescences dendritic, originating from proximal or medioproximal region of movable digit. Tritostemal laciniae usually fused for half or more of length, often rod-like, occasionally separated terminally (Fig. 45-4) Πcompletely separated at base (as in Seiodidae (Figure 3.2) and Davacaridae). Claviform vaginal sclerite absent. Mesogynial shield often large and triangular. Anal opening almost always on ventrianal shield. Tarsus I with or without claws ...... Cercomegistina, Superfamily Cercomegistoidea ----- 2
1*. With a single dorsal shield (sometimes line of fusion exist as in Philodanidae; if more than one shield, then claviform vaginal sclerites well-developed as in Neotenogyniidae) (Figs. 50-3, 50-4). Gnathotectum more or less triangular or rounded, smooth, often with a median keel (Figs. 47-5, 51-6). Palp genu usually with 7 setae (rarely with 5-6 setae). Palptarsal claw (= apotele) usually 2-tined (rarely 3-tined). Chelicerae often
95 tapeied^ edentate or with minute teeth OR robust, dentate. Cheliceral excrescences filamentous or rarely dendritic, originating horn various region of movable digit. Tritostemal laciniae free. Claviform vaginal sclerites (Figs. 50-3,52-1) or stemovaginal processes (Figs. 49-2,49-4) often present, sometimes head of vaginal sclerites reduced. Anal opening on ventrianal or free anal shield. Tarsus I without claws ...... Antennophorina ----- 7
2. Stemogynial shield, bearing 3rd stemal lyrifissure {stp3), present (Figure 3.18). Tarsus I with claws. Chaetotactic formula of dorsal setae on genua I- rV, 5-S-6-6...... 3
2'. Stemogynial shield, bearing 3rd stemal lyrifissure (sfp3), absent. Tarsus I with or without claws. Chaetotactic formula of dorsal setae on genua I-IV not as above ...... 4
3 Holodorsal shield. Prestemal shields paired. Stemal shields paired. Stemal setae 1 (stl) inserted in integument posterior to paired prestemal shields (Fig. 45-6). Male genital opening with paired eugenital setae ...... ASTERNOSEODAE
3'. 2 dorsal shields. Prestemal shield absent. Stemal setae 1 (stl) on an entire stemal shield. Gnathotectum more or less triangular, with smooth edge (Figure 3.9). Tritostemal laciniae free, not fused to each other. Latigynial and mesogynial shields not distinctly separated (Figure 3.18). Peritreme reduced. Male genital opening with no eugenital setae SEIODIDAE
4. Holodorsal shield. Gnathosoma strongly prognathous. Stemal setae stl-4 of female on a common stemal shield. Anal opening in small shield bearing
96 only paranal setae (Waltei^2000; Fig. 2). Tarsus L without claws. Legs IV hypertrophied, modified for jumping. Peritrematal shield with longitudinal groove behind coxae IV ...... SALTISEIIDAE
4'. Dorsum with 2 or more shields ...... 5
5. With four dorsal shields (Fig. 45-7). Metapodal shields free. Mesogynial shield well-developed, elongate-subrectangular (Fig. 45-8). Tarsus I with claws. Male genital opening with paired eugenital setae. Chaetotactic formula of dorsal setae on genua I-IV, 6-5-6-6 ...... DAVACARIDAE
5'. With two subequal dorsal shields (Fig. 45-2). Free metapodal shields absent. Mesogynial shield reduced or lost, or triangular in shape (Fig. 45-1). 6
6. Dorsal shields with relatively few setae. Lateral idiosomal setae on marginal shields (Lindquist and Moraza, 1993; Fig. 2), not on platelets in soft cuticle. Ventrianal shield fused posteriorly to opisthonotal and marginal shields. Tarsus I with or without claws. Paralaciniae present. Chaetotactic formula of dorsal setae on genua I-IV, 5-S-6-6. Free-living...... PYROS^IDAE
6'. Dorsal shields hypertrichous. Lateral idiosomal setae on platelets in soft cuticle (Fig. 45-1). Ventrianal shield free posteriorly; tarsus I without claws. Ventrianal shield free posteromarginally, not fused with dorsal shield. Paralaciniae absent. Chaetotactic formula of dorsal setae on genua I-IV, 6-6- 6-6...... CERCOMEGISTIDAE
7. Palp genu with 5-6 setae. Stemogynial shield absent. Pseudostemogynium often present. Claviform vaginal sclerite absent or head greatly reduced.
97 Median keel of gnathotectum^ reduced or absent. Chelicerae tapered, edentate, digits reduced with filamentous excrescences. Comiculi setiform or multi-tined. Setae av4 and pv4 on tarsus IV often short. Male eugenital setae absent ...... Superfamily Parantennuloidea 8
7’. Palp genu with 7 setae (if 6, then paired stemogynial shield present as in Echinomegistus (Paramegistidae), or dorsum with 3 dorsal shields as in Neotenogynium malkini (Neotenogyniidae)). Claviform vaginal sclerite present, or head often reduced. Median keel of gnathotectum often present. Chelicerae dentate or edentate. av4 and pv4 on tarsus IV variable. Male eugenital setae present or absent ...... 10
8. Palp tibia and tarsus fused. Palptarsal claw 2-tined. Mesogynial shield vestigial or absent. Anal shield wide, strap-like, with more than 5 pairs of setae. Dorsal shield hypertrichous, with more than 50 pairs of setae. Associated with tenebrionid beetles. (Kethley, 1977a; Fig. 2 ) ......
...... PHILODANIDAE
8*. Palp tibiae and tarsi normally articulated. Palptarsal claw 2 or 3-tined. Mesogynial shield well-developed or fused to ventrianal shield. Pseudostemogynium often present. Anal area with 2 pairs of setae. Dorsal shield not as above ...... 9
9. Stemal shield paired or fragmented. Mesogynial shield free, not fused to ventral shield (Fig. 50-1). Anal shield separated from ventral shield with no more than 2 pairs of setae present. Peritreme more or less reduced, not reaching to the base of coxae I. Chaetotactic formula of dorsal setae on genua I-IV, 5-S-6-6. Associated with carabid beetles or myriapods ......
...... PARANTENNULIDAE
98 y Stemal shield entire^ not hagmented (Figure Mesogynial ^ e ld fused to ventrianal shield. Metastemal shields fused to endopodal elements. Pseudostemogynium ellipsoid. Peritreme not reduced, reaching to the base of coxae I. Chaetotactic formula of dorsal setae on genua I-IV, 4-4-4-4. av4 and pv4 on tarsus IV short. Comiculi membranous with homed tip. Associated with carabid or passalid beetles ...... PROMEGISTIDAE
10. Prestemal shield(s) usually present (if absent, then pseudostemogynium or triangular mesogynial shield well-developed as in Physalozerconidae and Aenictequidae). Stemal shield entire, or paired, or rarely fragmented. Metastemal shield, bearing st4 and stp3, absent. Mesogynial shield well- developed or fused to ventral element. Head of vaginal sclerite often vestigial. Stemogynial or stemovaginal elements often present. Gnathotectum smooth, triangular or more or less roundish, median keel often obscure or absent. Chelicerae robust or tapered; with dendritic or filamentous cheliceral excrescences (Figs. 46-4,47-4, 49-6). Palp genu with 6- 7 setae. Ventral intercalary sclerite on tarsus IV free, not fused to tarsus. Male genital opening located in intercoxal region II-IV, not above base of tritostemum ...... 11
10'. Prestemal shield absent (if present, then dorsum with 3 dorsal shields as in Neotenogynium malkini (Neotenogyniidae)). Stemal shield entire, not paired. Metastemal shield(s), bearing st4 and sfp3, often present. Mesogynial shield, if present, small or vestigial, often fused to ventral element. Stemogynial or stemovaginal elements absent. Claviform vaginal sclerite well-developed, head well-developed (Figs. 50-3, 52-1, 52-6). Gnathotectum smooth, triangular, median keel present. Chelicerae robust, with large proximal tooth on movable digit; with dendritic or brush-like cheliceral excrescences (Figs. 51-3,51-4). Palp genu with 7 setae (if 6, then dorsum with 3 dorsal shields as in Neotenogynium malkini 99 (Neoteno^miidae)). Ventral intercalary sclerite on tarsus IV absent or fused to tarsus. Male genital opening located above base of tritostemum ......
...... Superfamily Celaenopsoidea ------20
11. Chelicerae tapered, edentate. Movable digit with filamentous excrescences (Fig. 49-6). Comiculi membranous or setiform, often with small protuberance(s). Anal shield fused with ventral shield. Associated with ants, carabids, millipedes, or reptiles ...... 12
11'. Chelicerae robust, dentate. Movable digit with filamentous or dendritic excrescences (Figs. 46-4, 47-4). Comiculi sclerotized, hom-like. Anal shield free from or contiguous to ventral shield. Associated with passalid beetles...... 17
12. Stemogynial shield bearing stemal lyrifissures 3 (stp3) present (F ig . 47-2). Stemovaginal elements absent. Peritreme normal. Pseudoperitreme absent. Comiculi membranous. Associated with arthropods or reptiles. ..
...... Superfamily Paramegistoidea ------PARAMEGISTIDAE
12*. Stemogynial shield absent. Latigynial shield each usually with many setae. Stemovaginal element often present. Peritreme short, often with pseudoperitreme. Comiculi often setiform, often with small protuberance(s). Strictly associated with ants...... 13
13. Palp tibia and tarsus normally articulated. Mesogynial and ventral elements fused narrowly (F ig . 49-5)......
...... Superfamily Antennophoroidea ------ANTENNOPHORIDAE
13*. Palp tibia and tarsus insensibly fused. Stemal shield with paired posterior extensions devoid of setae and pores (= stemovaginal elements; Figs. 49-2,
100 49-4)^ and usually (or oAen% covered by lati^m ial shields. Meso^mial shield usually well-developed (Figs. 48-4,49-1) (if obscure or absent, then prestemal shields present as in Messoracaridae) (Fig. 48-2) ...... Superfamily Aenictequoidea 14
14. Prestemal shields paired. Latigynial shields elongate, subrectangular, with parallel mesal margins. Mesogynial shield obscure or absent. (Fig. 48-2) ...... MESSORACARIDAE
14'. Prestemal shield absent. Latigynial shields subtriangular, mesal margins not parallel. Mesogynial shield triangular (Figs. 48-4, 49-1,49-4) ...... 15
15. Latigynial shields each with two setae. Peritreme-like groove (= pseudoperitreme) present. (Fig. 48-4) ...... AENICTEQUIDAE
15'. Latigynial shields each with more than 8 setae (Figs. 49-1,49-4). Pseudoperitreme absent ...... 16
16. Stemal setae 1 (sfl) on a paired prestemal shields. Latigynial shields each with less than 12 setae. (Fig. 49-1) ...... PTOCHACARIDAE
16'. Stemal setae 1 (stl) inserted in entire stemal shield. Prestemal shield absent. Latigynial shields each with more than 30 setae. (Fig. 49-4) ...... PHYSALOZERCONIDAE
17. Chelicerae with dendritic excrescences (Fig. 47-4). Anal shield completely separated from ventral shield. Stemogynial shield entire or divided (F ig s. 47-3,48-1). Pedo fossae (= fovae pedales) absent. Palp tibia and tarsus normally articulated. Palptarsal claw (= apotele) 3-tined. Large mite, ranging over 3 mm long ...... Superfamily M egisthanoidea ------18 101 IT. Cheliceral digits short with filamentous excrescences (Fig. 46-4). Anal shield fused with or contiguous to ventral shield (Figs. 46-2, 46-6). Stemogynial shield entire, inversely triangular (Fig. 46-3). Often with pedo fossae (= fovae pedales) to accommodate folded legs. Turtle-like mite, ranging ca. 1 mm long ...... Superfamily Fedrizzioidea 19
18. Prestemal shields paired, each with stl. Latigynial shield fused together, free from ventral shield. (Fig. 48-1). Vaginal sclerites well-developed ...... HOPLOMEGISTIDAE
18'. Prestemal shield, bearing stl, usually entire, rarely paired, weakly sclerotized. Latigynial and mesogynial shields lost, represented by a ridge bordering the stemogynial elements posteriorly. Stemogynial shield divided or entire, completely covering the genital orifice (Fig. 47-3). Vaginal sclerites reduced, without heads but often with thickened arms...... MEGISTHANIDAE
19. Latigynial and mesogynial shields well-developed (Figs. 46-3,46-6). Vaginal sclerites present but head reduced. Male genital aperture ellipsoid, located between coxae III. Male eugenital setae often present ...... KLINCKOWSTRGEMnO AE
19'. Latigynial and mesogynial shields reduced or lost, covered by enlarged stemogynial shield (Fig. 46-2). Vaginal sclerites absent or obscure. Male genital aperture circular or only slightly wider than long, located between coxae n m. Eugenital setae absent ...... FEDRIZZIIDAE
20. Palp genu with 6 setae. With 3 dorsal shields plus with prestemal shield (Figs. 50-3, 50-4) (if holodorsal shield present, then prestemal shield
102 absent). Peritremes greatly reduced., Associated with Neotropical millipedes ...... NEOTENOGYNHDAE
20'. Palp genu with 7 setae. With holodorsal shield. Stemal setae 1 (stl) on an entire stemal shield (Figs. 50-5, 51-1, 51-5, 52-1, 52-6) ...... 21
21. Latigynial shields well-developed, free from ventral elements. Mesogynial shield usually small flanked by latigynial shields or reduced. (Figs. 51-1,51-2)...... 22
21'. Latigynial shields fused to ventral elements posteriorly. Mesogynial shield usually vestigial or fused to ventral elements. (Figs. 50-5, 51-5, 52-1, 52-6)...... 24
22. Anal shield fused to ventral shield (Fig. 51-1) {Neodiplogynium is exception). Mesogynial shield, if present, usually less than 1/3 the length of the latigynial shields. Associated with arthropods or reptiles ...... DIPLOGYNDDAE
22'. Free anal shield present. Mesogynial shield usually longer than 1/3 the length of the latigynial shields. Free-living or associated with insects or millipedes ...... 23
23. Metastemal shield absent. st4 and stp3 on stemal shield. Mesogynial shield at least 1/2 the length of the latigynial shields. Comiculi weak, bifurcated. Free-living in soil-litter, on bark, in bark beetle galleries, or in the nests of stingless bees. (Funk, 1977; Fig. 1) ...... TRIPLOGYNIIDAE
23'. Metastemal shield, bearing st4 and stp3 present. Mesogynial shield very small or vestigial. Comiculi hom-like. Associated with millipedes. (Hunter, 1993b; Fig. 4) ...... COSTACARIDAE 103 24. Anal shield free (Fig. 51-5) or fused with ventral shield. Often multi-tined paralaciniae present. Latigynial and mesogynial elements free hrom each other mesally, fused with ventral element posteriorly. Metastemal shield(s), bearing st4, narrow, strap-like (absent in Karkinoeuzercon). Associated with a variety of arthropods or with snakes ...... 25
24'. Anal shield fused with ventral shield. Free, narrow postanal shield, not bearing anal opening, often present. Latigynial-mesogynial complex insensibly fused (Fig. 52-1), with at most a small median notch present on anterior margin of ventral-genital element (Fig. 52-6). st4 on sternal shield or on well-developed metastemal shields. Free-living or associated with bark beetles ...... 26
25. Anal opening on free anal shield, bearing 2 pairs of setae. Median separations between mesogynial and latigynial elements not extending past level of coxae III (Fig. 51-5). Comiculi always serrate. Hypostomal setae hsl on anteriorly extended projection or lobe. Associated with passalid beetles or millipedes ...... EUZERCONIDAE 25' Free anal shield often present. Latigynial shields elongate, often extending posteriorly beyond coxae IV. Comiculi hom-like. Hypostomal setae hsl not on anteriorly extended projection. Free-living or associated with passalid beetles, bark beetles, or snakes. (Fig. 50-5) ...... SCmZOGYNIIDAE
26. Metastemal shield, bearing st4 and stp3, well-developed, paired (Fig. 52-1). Free, narrow postanal shield, not bearing anal opening, often present. Fused endopodal-metapodal-peritrematal elements fused with ventrianal 104 shield. Associated with bark, and beetles burrowing under bark. ______...... CELAENOPSIDAE
26'. Metastemal shield absent, fused to sternal shield. Setae st4 on sternal shield. Postanal shield absent (Fig. 52-6). Endopodal-peritrematal shields fused, but free from ventrianal shield. Large mites with body >1 mm in length. Free-living ...... MEGACELAENOPSIDAE
105 Diagnoses of the S u p e rfam ilies, o f Trigynaspida.
Based on the results from this study, the following 8 superfamilies within Trigynaspida are proposed. This proposal is different from that of Kethley (1977b), in which he proposed 7 superfamilies. As stated above, the difference is derived from the erection of the new superfamily, Paramegistoidea.
Superfamily Cercomegistoidea
DIAGNOSIS: Body often with moderate hypertrichy. Adult female usually with separate podonotal and opisthonotal shields or holodorsal shield, rarely with opisthonotal region with separate mesonotal and pygidial elements. Peritremes and peritrematal shields usually well developed, the latter fused anteriorly to podonotal shield. Venter with large ventrianal or genitoventral shield. Free anal shield absent. Female genital region with 3 shields or their remnants: mesogynial shield usually well-developed, latigynial shields usually well-developed; stemogynial shield (= stemogynium), if present, entire; stemal shield well-developed (entire) or fragmented. Claviform vaginal sclerites usually absent. Male genital opening on intercoxal shield, usually between coxae Ill-IV, and covered by 1-2 transverse valves. Tarsus 1 with or without claws. Tarsus IV with setae av4 and pv4 on a well-defined ventral intercalary sclerite. Tritostemal laciniae usually fused for more than half of their length, or separated at base. Chelicerae robust, chelate-dentate; movable digit with brush-like excrescences; gnathotectum often denticulate or rarely smooth, without median ventral keel; palp genu with 6 setae; palptarsal claw (= apotele) usually 3-tined.
106 Superfamily Paranteiinuloidea
DIAGNOSIS: Oval, flattened mites often with minute dorsal setae and a marginal fringe of enlarged, flattened setae. Prestemal shield absent. Stemal shield variously fragmented. Stemogynial shield bearing stp3 absent. Pseudostemogynium often present. Mesogynial shield free from or fused to large ventral or ventrianal shield; latigynial shields generally well-developed. Palp tibia and tarsus fused or separate; palp genu with 5 or 6 setae; chelicerae not robust, tapered, edentate, digits reduced, movable digit with filamentous excrescences located in proximal region of digit; fixed digit with membranous distal processes or fimbriate dorsal excrescence. Comiculi often setiform or rarely multi-tined. Male eugenital setae absent.
Superfamily Paramegistoidea, NEW
DIAGNOSIS: Oval, flattened mites with minute dorsal setae and distinctly larger stylus-like, spinous, or foliate ventrianal setae. Prestemal shield always present. Stemal shield paired or two shields connected by narrow bridge. Stemogynial shields, bearing stp3, paired. Mesogynial shield free from or fused to large ventrianal shield; latigynial shields generally well-developed. Palp tibia and tarsus fused or rarely separated; palp genu with 7 setae (Echinomegistiis with 6 setae); chelicerae edentate, movable digit with filamentous or more or less dendritic excrescences. Comiculi membranous, often with sclerotized tip. Male eugenital setae often present.
107 Superfamily Anteimophoroidea
DIAGNOSIS: Stemal setae 1 (stl) on entire prestemal shield. Mesogynial shield reduced, with anterior pedicellate ventrianal element dividing narrow latigynial shields; latigynials each with more than two pairs of setae. Peritremes short. Palp tibia and tarsus distinct, unfused; chelicerae with minute teeth, movable digit with filamentous excrescences. Male with eugenital setae. Comiculi with small protuberance(s) or more or less serrate.
Superfamily Aenictequoidea
DIAGNOSIS: Stemal setae 1 (stl) on separate prestemal shields or on entire stemal shield. Stemal shield with a pair of posterior projections lying under latigynial shields (stemovaginal processes). Pseudostemogynium present. Mesogynial shield subtriangular, separate from ventrianal shield, and overlapped or covered by latigynial shields each bearing 2 to numerous setae. Peritremes short, sometimes associated with an additional peritreme-like groove (= pseudoperitreme). Palp tibia and tarsus insensibly fused; chelicerae tapered, edentate. Comiculi with small protuberance(s).
Superfamily Megisthanoidea
DIAGNOSIS: Stemal setae 1 (stl) on paired or entire prestemal shield(s) often weakly sclerotized; stemal shield entire, with 3 pairs of setae (sf2-st4). Mesogynial shield obscure; latigynials insensibly fused to each other or weakly separated, free from ventral shield; Stemogynial shield entire or divided. Anal shield free, not fused with ventral shield. Palp tibia and tarsus 108 separate; palptarsal daw 3-tined^ chelicerae robust^ strongly toothed^ movable digit with dendritic excrescences.
Superfamily Fedrizzioidea
DIAGNOSIS: Stemal setae 1 (stl) on entire prestemal shield, stemal shield with 3 pairs of setae {st2-st4). Stemogynium entire, hinged along posterior margin of stemal shield. Mesogynial and latigynial shields well developed, or reduced and withdrawn under ventral plate. Pedofossae often present. Anal shield subtriangular, fused with or contiguous to ventral shield. Palp tibia and tarsus distinct, unfused; chelicerae robust, with short digits and dense, mop-like mass of filamentous excrescences; comiculi hom-like.
Superfamily Celaenopsoidea
DIAGNOSIS: Prestemal shield always absent (except for Neotemgynium malkini). Stemal shield entire, bearing 3-4 pairs of setae; free metastemal shield bearing sti and stp3 often present. Mesogynial shield small, triangular and free or fused with latigynial and/or ventral elements; latigynial shields free or fused to ventral plate; claviform vaginal sclerites well developed. Venter covered by a single extensive plate or plate flanked by free lateral and/or anal shields. Palp tibia and tarsus distinct, unfused; palp genu with 7 setae (except for Neotenogynium malkini carrying 6 setae); cheliceral digits robust, dentate, movable digit with large proximal tooth and dendritic or brush-like excrescences. Male genital opening on anterior margin of stemal shield. Ventral intercalary sclerite fused to tarsus IV.
109 CHAPTER 4
MOLECULAR PHYLOGENETICS OF TRIGYNASPIDA (MOLECULAR ANALYSES)
INTRODUCTION
Since the mid 1980s, with the aid of PCR technology, molecular phylogenetics has emerged as one of the integral parts of phylogenetic systematics. Despite the advances in other fields of biology, and despite the fact that acarology has many unanswered primary questions, phylogenetics of acarines based on molecules have only recently received attention. While there are a few studies on acariform mites (e.g., Fenton et al., (1997) for Eriophyidae; and Navajas et ai, (1997,1999) for Tetranychidae) and parasitiform mites (Klompen, 2000), most of the molecular phylogenetic studies have been focused on parasitiform. ticks (Black and Piesman, 1994; Black et ai, 1997; Dobson and Barker, 1999; Klompen et ai, 2000; Norris et al, 1999).
As to the phylogenetics of trigynaspid mites at the higher level, no molecular analysis has been performed. Accordingly, as a continuum of the phylogenetic systematics of Trigynaspida, molecular phylogenetics based on DNA sequences have been performed. Although this is the first attempt to
110 investigate the h i^er level relationships among the members of Trigynaspida based on molecules, its purpose is extended to test the tree inferred from morphological study.
I l l MATERIALS AND METHODS
Taxon-Sampliiig
This study was intended to survey the higher level systematics of trigynaspid mites, and therefore, inclusion of multiple representatives from currently-known superfamilies of Trigynaspida was attempted. The facts, however, that molecular studies often require fresh materials and given that these mites are more or less rare in nature did not allow the inclusion of all the known taxa for the study. Consequently, representatives of all the trigynaspid superfamilies, except for Aenictequoidea, along with the outgroup taxa in the phylogeny of Mesostigmata proposed by Johnston in Norton et al. (1993, Figure 1.2) were included. This sampling scheme includes a total of 12 ingroup and 6 outgroup taxa for 18S; 14 ingroup and 7 outgroup taxa for 28S; and 15 ingroup and 7 outgroup taxa for the combined data (Table 4.1). Although these outgroups are primarily used for rooting the trees for the test of ingroup monophyly (Barriel and Tassy, 1998; Giribet and Ribera, 1998), these multiple outgroups often allow better alignments especially in the variable (loop) regions.
While a few of these mites were collected by me, most of them were given by the professional and amateur entomologists from all around the world. For the field collection of target mites, search of known host groups of
112 Superfamily and Family Species Gene Voucher No. SUBORDER CERCOMEGISTINA
S.F. Cercomegistoidea
Astemoseiidae Astemoseius sp. 18S, 28S AL5467 Cercomegistidae Cercoleipus coelonotus 18S, 28S CMK 971201-1
SUBORDER ANTENNOPHORINA
S.F. Celaenopsoidea
Diplogyniidae Cryptometastemum sp. 18S CMK 971006-4 Passalacarus sylvestris 18S, 28S AL5190 Trichodiplogyrtium sp. 285 CMK 970330-1 Euzerconidae Euzercon latiis 18S, 28S AL5190 Neoeuzercon sp. 18S, 28S BMOC 96-1105-205
S.F. Parantennuloidea
Parantennulidae Micromegistus gourlayi 18S, 285 CMK 980716-1-9
S.F. Aenictequoidea No Taxon Used
S.F. Antennophoroidea
Anteimophoridae Antennophonis wasmanni185, 285 BMOC 99-0804-003
S.F. Fedrizzioidea
Fedrizziidae Fedrizzia sp. 285 CMK 971210-2 Neofedrizzia sp. 185, 285 CMK 971006-3 Klinckowstroemiidae Klinckowstroemia sp. 185, 285 CMK 970904-3 Paramegistidae Paramegistus confrater 185, 285 CMK 990605-1 Promegistidae Promegistus amislrongi 285 CMK 980906-4
S.F. Megisthanoidea
Megisthanidae Megisthanus floridantts 185, 285 AL5190
(Continued)
Table 4.1: List of taxa used for molecular study. S.F. = Superfamily
113 Table 4.1 (Continued)
SUBORDER UROPODINA
Trachyuropodidae Oplitis sp. 185, 28S AL5252 Polyaspididae Polyaspis lamellipes 185, 285 AL5564
SUBORDER SEJINA
Sejidae Sejus sp. 185, 285 AL5072 Epicroseius sp. 185, 285 AL5480 Ichthyostomatogasteridae Astemolaelaps sp. 185, 285 AL5564
SUBORDER HETEROZERCONINA
Discozerconidae Discozercon sp. 285 CMK 980716-1-16 Heterozerconidae Heterozerconidae n. sp. 185, 285 AL5914
animals, such as passalid beetles, centipedes, polydesmid and spirostreptid millipedes, were carried out at their specific habitats (e.g., rotten logs, under the bark, nests, etc.). In the field, each collected host specimen carrying target mites was directly preserved in a vial containing a data label and 95% ethyl alcohol. For the free-living trigynaspids, leaf litter and humus of forest were also collected. Target mites from these samples were isolated by the Berlese- TuUgren funnels and by high-powered stereo microscopes, and then preserved in 95-100% ethyl alcohol in -80°C freezer until they are materialized.
Voucher specimens, mounted on glass slides, were deposited at the Acarology Laboratory (OSAL), Museum of Biological Diversity, Department of Entomology, The Ohio State University. Mites removed from a single host or from Berlese-TuUgren funnel were first identified to genus level, and 1 to 6 114 slides were prepared for the correct identification of the species. For some, but not all, materials, a squeeze method to force the contents of the body, including muscles and body fluids containing DNA, was applied during the DNA extraction to recover the body parts, though broken, for the voucher specimens. These specimens were mounted and preserved in slide boxes for molecular vouchers in the OSAL.
Selection of Genes
The NS5-NS8 region of 18S (= from the part of 31 stem to the part of 50 stem region of Black et al. (1997) and Van de Peer et al. (1994)) and the V-X region of 28S (= from part of 70 to part of 91 region of De Rijk et al. (1994), Kjer (1995), and Larsen (1992)) nuclear ribosomal DNA (rDNA) genes were used for the study. These genes have been very well-documented (Black et ai, 1997; Giribet et ai, 1999; Hillis and Dixon, 1991; Hwang and Kim, 1999; Klompen et ai, 2000; Larsen, 1992; Van de Peer et ai, 1994), and are present as multiple copies (up to ca. 5000 copies) within a genome that can offer higher success rate in amplification and sequencing than those in the single copy genes. Furthermore, molecular phylogenetic studies from ticks (Black et ai, 1997; Dobson and Barker, 1999; Klompen et ai, 2000) imply the fact that these genes can be useful for inferring phylogenetic relationships of higher-level systematics in Mesostigmata, including Trigynaspida. Both of these genes retain conserved regions along with fast evolving regions, allowing sorting of the taxa ranked at the Class to the Family level (Hwang and Kim, 1999).
Using at least two genes, rather than one gene, in phylogenetic study helps to keep systematists from falling into the pitfall of generating, so-called, gene trees rather than species trees (Doyle, 1992; Pamilo and Nei, 1988). Hence, it is suggested to use more than one gene in molecular phylogenetics as the
115 species tree is thought to be the tree h a x ti the sum total of the information of gene trees (Crozier, 1993).
Molecular procedures
DNA from 1 to maximum 6 individuals of fresh mite(s) from each taxon was extracted by the modified methods of Gustincich et al. (1991) and Black et al. (1997), using CTAB extraction protocol. Among the extracted genomic DNA, the specific target sequences of the gene, which were eventually sequenced and analyzed {i.e., NS5-NS8 region of 18S; and V-X region of 28S) were amplified with the set of primers (see Table 4.2) and Taq polymerase by following the protocols of Black and Piesman (1994) of polymerase chain reaction (PCR) (Saiki et al., 1988).
Gene Region Base Composition 185 N55 5-AACTTAAAGGAATTGACGGAAG-3' N58 5-TCCGCAGGTTCACCTACGGA-3 285 V 5-GTAGCCAAATGCCTCGTCA-3 X 5-CACAATGATAGGAAGAGCC-3’
Table 4.2: Sequences of primers used for the amplification of 18S (NS5 and NS8; from Black et ai, 1997) and 285 (V and X; from Hillis and Dixon, 1991) nuclear rDNA.
116 The PCR products were then purified by Wizard* PCR preps DNA purification kit (Promega Biotech Inc., Madison, Wisconsin, USA) and were sequenced by the ABI Prism 100 automated sequencer at the automatic sequencing facilities in the University of Georgia. The primers used for the PCR were also used for the sequencing. The two complementary sequences of opposite direction (5 - and 3'-) from a single taxon were compared by SeqAid n 3.6 (Rhoads and Roufa, 1989), to get a single 5'- sequence for each taxon.
Alignment
Although it is generally considered that DNA sequence data are more objective than morphological data because of the arbitrary and often controversial interpretations on morphological characters, it needs to be mentioned that molecular data too are not free from this argument. In molecular systematics based on nucleotide sequences, assessment of homology among the bases {i.e., characters) is summarized by the alignments among the bases of the sequences included. In other words, the scientific objectivity of molecular phylogenetics depends largely on the criteria applied to the alignments of nucleotide sequences.
In this study, the criteria of positional similarity of the base, model of secondary structure of rRNA, and compensatory base pair change have been applied for the alignment (Figure 4.4). All the DNA sequences were transcribed to RNA sequences, first aligned by ClustalX (Thompson et al., 1997), and then corrections of alignments were carried out with the aid of proposed model of secondary structures of rRNA. The model of secondary structure was followed from Black et al. (1997) and Van de Peer et al. (1994) for the IBS, and from De Rijk et al. (1994), Kjer (1995), and Larsen (1992) for the 28S.
117 Aligned sequences for the NS5-NS8 region of 18S rDNA gene in this study include 714 characters {i.e., aligned positions, including bases and gaps). Among these, 219 characters (30.7%) are parsimony-informative. The 28S data for this study include 738 characters, of which 191 characters (25.9%) are parsimony-informative. Each sequence of DNA, combined from these two 18S and 28S genes, is composed of a total of 1452 positions of aligned bases and gaps per taxon, of which 410 positions (28.2%) were parsimony informative in deducing evolutionary relationships. Aligned sequences of each gene is shown in the Appendices A and B.
As the different and often unreliable alignments can lead to different or incorrect inference of phylogeny (Winnepenninckx and Backeljau, 1996), variable regions of rDNA are often practiced to be removed from the analyses (Giribet et ai, 1999). However, as the variable regions contain crucial phylogenetic information (Hwang et al., 2000), none of the data at the loop regions has been removed from this study (see Gatesy et ai, 1993).
Analyses
Two separate analyses for the data from each 18S and 28S genes along with combined analyses for these two genes were performed. In all the analyses, all the gaps in the sequences, derived by indels (insertions or deletions), were coded as 'missing' as implemented in PAUP* 4.0b8 (Swofford, 2001). All the characters were equally weighted and treated as unordered. Heuristic searches with random addition of the taxa with 100 replicates were performed. Maximum parsimony, which seeks the minimum number of steps of evolutionary change within the data, implemented in PAUP* was used as the optimality criterion. Branches were collapsed if the maximum branch length is zero. 118 To assess the confidence level on the deduced phylogeny, ^branch support' (Bremer, 1988) (= 'decay index' sensu Donoghue et al, 1992; 'Bremer support' sensu Kâllersjô et al, 1992) estimated by AutoDecay 4.0.2 (Eriksson, 1999) using global reverse constraints of topologies. For the combined analyses, the percentage of incongruence length differences (ILD; Farris et al, 1995; Mickevich and Farris, 1981) among each partitioned data set were calculated. Partition-homogeneity tests (PHT; Huelsenbeck e ta l, 1996) with 100 homogeneity repetitions along with 20 sequence replicates incorporated in PAUP* were also performed.
119 RESULTS AND DISCUSSION
Comparison of Topologies
In the separate analyses of 18S and 28S sequence data, the portion of 18S generated three equally parsimonious trees of 780 steps (Table 4.3). A topology of strict consensus tree out of these three trees was largely unresolved, even failing to recognize ingroup monophyly (Figure 4.1). Although this would reveal the lack of phylogenetic information in the NS5-NS8 region of 18S sequences because of the highly conserved nature of this gene (Klompen et ai, 2000), this may also be explained by the poor sampling of the ingroup taxa (i.e., systematic error) that may help better internal resolutions (Swofford et ai, 1996).
From the 28S data, nine trees, each having a length of 767 steps, were discovered. A strict consensus tree recovered monophyletic Trigynaspida, carrying the bifurcating lineages of Cercomegistina and Antennophorina (Figure 4.2). Although antennophorine lineages were not resolved, the tree implied the recognition of superfamilial groupings, such as Parantennuloidea, Fedrizzioidea, and Celaenopsoidea. It is noted that Promegistus (Promegistidae) is not the sister to Paramegistus (Paramegistidae) but the sister to Micromegistus (Parantennulidae), supporting the same result discovered from the morphological study (Chapter 3).
120 Gene # of # o f # o f # of Tree a RI Taxa Positions Informative Trees Length Positions 18S 18 714 219 3 780 0.51 0.45 28S 21 738 191 9 767 0.48 0.45 18S + 22 1452 410 3 1551 0.49 0.44 28S
Table 4.3: Tree statistics.
The analyses from the combined 18S and 28S data yielded three equally parsimonious trees of 1551 steps. Despite the lack of either one set of 18S or 28S data in some taxa in combined analyses (see Table 4.1), strict consensus tree obtained from these 3 trees was well-resolved, leaving only a trichotomy of the three genera Cryptometastemum, Trichodiplogynium, and Passalacarus of monophyletic Diplogyniidae (Figure 4.3).
For the question of the monophyly of Trigynaspida and higher level relationships within it, as proposed by Kethley (1977b), the monophyly of Trigynaspida, containing bifurcating lineages of the suborder Cercomegistina and Antennophorina, was re-discovered from the combined molecular data. As to the higher relationships within Trigynaspida, Kethley's scheme of Fedrizzioidea, which contains Fedrizziidae, Klinckowstroemiidae, Promegistidae, and Paramegistidae, was not supported as this superfamily is not monophyletic. That is, while two sister families of Fedrizziidae and Klinckowstroemiidae are more or less basal in the lineages of Antennophorina of Trigynaspida, Promegistidae and Paramegistidae show closer relationships to Parantennuloidea and Antennophoroidea. That is, as
121 Astemoseius I Cercomegistoidea Cercoleipus Micromegistus I Parantennuloidea Paramegistus I Paramegistoidea Antennophorus I Antennophoroidea Klinckowstroemia I Fedrizzioidea Neofedrizzia
Megisthanus I Megisthanoidea Cryptometastemum Passalacarus Celaenopsoidea 361 Euzercon Neoeuzercon
Polyaspis I Uropodina Oplitis
Heterozerconidae I Heterozerconina Astemolaelaps
Sejus Sejina
Epicroseius
Figure 4.1: Strict consensus of 3 equally parsimonious trees deduced from NS5-NS8 region of 18S rDNA sequences under no topological constraints. Gaps were treated as missing. Numbers on the branches indicate branch support sensu Bremer (1988).
122 I Asternoseius Cercomegistoidea Cercoleipus Micromegistus Parantennuloidea Promegistus
Paramegistus Paramegistoidea 1 * ^ Antennophorus Antennophoroidea Kiinckowstroemla 10 Fedrizzia Fedrizzioidea Neofedrizzia 9) Megisthanus I Megisthanoidea
HI Trichodiplogynium Passalacarus Celaenopsoidea Euzercon Neoeuzercon
Polyaspis Uropodina Oplitis
Discozercon Heterozerconina Heterozerconidae
Astemolaelaps
Sejus Sejina Epicroseius
Figure 4.2: Strict consensus of 9 equally parsimonious trees inferred from the V-X region of 28S rDNA sequences under no topological constraints. Gaps were treated as missing. Numbers on the branches indicate branch support senstt Bremer (1988). 123 Astemoseius I Cercomegistoidea Cercoleipus Micromegistus I Parantennuloidea Promegistus Paramegistus I 'Paramegistoidea' s — Antennophorus I Antennophoroidea Klinckowstroemia Fedrizzia Fedrizzioidea Neofedrizzia Megisthanus I Megisthanoidea Trichodiplogynium Cryptometastemum Passalacarus Celaenopsoidea Euzercon Neoeuzercon Polyaspis OpIItls Uropodina Sejus Epicroseius Sejina Astemolaelaps Discozercon Heterozerconina Heterozercon
Figure 4.3: Strict consensus of 3 equally parsimonious trees deduced from 18S+28S sequence data under no topological constraints. Gaps were treated as missing. Numbers on the branches indicate branch support sensu Bremer (1988).
124 discovered in the separate 28S analyses, Promegistus (Promegistidae) becomes the sister to Micromegistus (Parantennulidae), which suggest the revision of the superfamily Parantennuloidea. This newly expanded Parantennuloidea is associated with carabid beetles or myriapods. The remaining family of Paramegistidae, well-known associates of millipedes or squamate reptiles, is raised alone as Paramegistoidea. These designations of the superfamilial groupings support the results of morphological analyses shown in Chapter 3.
The consensus tree from the combined analyses, however, differs from the tree inferred from morphology in terms of the phylogenetic position {i.e., branching order) of Parantennuloidea. That is, while the Parantennuloidea branched at the base of Antennophorina in the tree of morphology, it branched at the tip of Antennophorina, becoming the sister to Paramegistidae in the combined molecular analyses. Along with the fact that the molecular data of Aenictequoidea are missing in this study, this topological difference might be ascribed to less complete sampling of the ingroup taxa compared to the taxon sampling in morphology. However, despite the discordance in phylogenetic position of Parantennuloidea in these two trees, the clade-based taxonomic groupings {i.e., superfamilies) of Trigynaspida were identical in both morphological and combined molecular analyses.
When the monophyletic Fedrizzioidea sensu Kethley (1977b) was enforced from the combined 18S and 28S data set, 14 more steps {i.e., 1565 steps; 3 trees) were required. A strict consensus tree from these enforced trees, however, showed the sister group relationships between Micromegistus (Parantennulidae) and Celaenopsoidea.
125 5'- [ 77 ] [ 77' ] - 3 Asternoseius [GGÜGG] [CCACC] Cercoleipus [GGÜGG] [CCACC] Mi cromegistus [GGUGG] [CCACC] Promegistus [GGÜGG] [CCACC] Paramegistus [GGUGG] [CCACC] Antennophorus [GGUGG] [CCACC] Klinkowstroemia [GGUGG] [CCACC] F edrizzia [GGUGG] [CCACC] Neofedrizzia [GGUGG] [CCACC] Megisthanus [GGUGG] [CCACC] Trichodiplogyni urn [GGAGG] [CCUCC] Passalacarus [GGAGG] [CCUCC] Euzercon [GGCGG] [CCGCC] Neoeuzercon [GGCGG] [CCGCC] P olyaspis [AGUGG] [CCACU] O p litis [AGGGG] [CCCCU] Asternolaelaps [AGUGG] [CCACU] Epicroseius [AGUGG] [CCACU] Sejus [CGUGG] [CCACG] Discozercon [AGUGG] [CCACU] Heterozercon [GGUGG] [CCACU]
Figure 4.4: Sequence alignment of 77 and matching 77' regions of 288 rRNA, showing compensatory base pair changes. The first position (base) of 5'- of 77 stem matches to the last position (base) of 3'- in 77' stem. Note that the group Trigynaspida is uniquely distinguished from the outgroups. The region between 77 and 77' is omitted. Brackets are used to define the stem region.
*****
126 CHAPTERS
COMBINED ANALYSES
INTRODUCTION
A phylogeny inferred from a single set of characters, such as
morphological or molecular data, is a mere hypothesis that requires test confirmation). Although obtaining a single topology with identical branching order obtained from different data sets would be regarded as 'passing the test', very often, an identical topology from different studies is not obtained. Presumably, this resulted from the different number of taxa involved in each separate analysis, different attributes of characters, or, for the molecular analyses, (accidentally) choosing less informative gene or the region of the genes. The possible options to cope with this practical dilemma are enforcing a taxonomic / topological congruence, building a supertree, or performing a combined analysis.
Although the solution of taxonomic congruence, similar to seeking a consensus tree from multiple equally parsimonious trees, would be understood as a safe way, rendering a conservative hypothesis (Miyamoto and Fitch, 1995; Swofford, 1991), the resultant phylogeny may not be the phylogenetic truth as it fails the discovery of natural hierarchy by allowing
127 unresolved polytomies. Irt fact^ it retains very high, likelihood to generate completely unresolved relationships among the taxa especially when the branching orders are not identical at the base of each tree.
Inference of a supertree is based not on the characters but on the branches and ancestors deduced from the characters (Baum, 1992; Ragan, 1992; Sanderson et al., 1998). This method is therefore another form of taxonomic congruence. In this method, all the ancestors {i.e., nodes) of every clade within an input tree become the characters of a new matrix for inferring a supertree, and the total number of characters equals to the number of ancestors in all the original input trees. Although this relatively a simple methodology offers a solution to deal with multiple trees inferred from different methodologies, it treats both well and poorly supported clades {i.e., branches) equally, causing weighting problem in the new matrix. In addition, as this methodology is based on tree re-coding', regardless of the total number of characters in each data set used for each phylogenetic inference, when one tree has far more nodes than the others, the new matrix can be governed by the tree having more nodes. Finally, when the taxa included in different trees are not the same, the new matrix contains many characters that are 'artificially' unavailable for these non-matching taxa. All of these biases are eventually incorporated into the inference of a new phylogeny {i.e., supertree).
Strictly speaking, the final solution, inferring a tree from a combined data set to obtain a global optimal tree {i.e., combined analyses) (Kluge, 1998), is not the ultimate but a compromised solution, creating another hypothesis hrom that combined data set. This approach may also be vulnerable to deal with the cases where the hierarchical information in one data set overwhelmingly exceeds the other (e.g., morphology vs. molecules), or where the evolutionary history behind the two data sets is not parallel {e.g., nuclear 128 genome vs. oiganellai genome). Despite these philosophical and factual shortcomings, pragmatically, the approach of combined analyses is perhaps the best strategy available at present until one finds a better solution that resolves the differences in topologies in different analyses. It should also be noted that, when two or more different data sets {i.e., data partitions) are not heterogenous, often, locally unresolved branches within a tree from one data set can obtain a better (or full) resolution from the compensating different data set in the combined analyses (Chippindale and Wiens, 1994; Kluge and Wolf, 1993).
As to the phylogenetics of Trigynaspida based on morphological (Chapter 3) and molecular (Chapter 4) characters, although each superfamilial grouping was well-conserved in both strict consensus trees from two different analyses, the topologies {i.e., branching order) from these two analyses were not compatible at the position of Parantennuloidea, now including Parantennulidae, Philodanidae, and Promegistidae. In other words, in morphology, Paranteimuloidea was branched at the base of Antennophorina, while the molecules showed that Paranteimuloidea, represented by Micromegistus (Parantennulidae) and Promegistus (Promegistidae), is the sister to Paramegistidae (Paramegistoidea), disclosing incongruent topologies inferred from two different analyses.
To solve this problem of disparity between topologies inferred from two difrerent sets of data, among others, combined analyses, treating two data sets into one, were performed. Although the purpose of this chapter is to seek a global optimum tree from all the combined data set, it is also intended to test the cladogram of Trigynaspida proposed by Kethley (1977b), which has been rejected in both morphology and molecular analyses.
129 MATERIALS AND METHODS
laxQD. Sampling
The purpose of this study is to examine the higher level relationships of trigynaspid mites based on all combined data, and therefore, all the taxa used for both of morphological and molecular analyses were included. This includes a total of 51 species, which are composed of 40 species in 23 families from all the currently known superfamilies of Trigynaspida along with 11 species in 9 families of their 'monogynaspid' outgroup taxa (Table 5.1). Selection of outgroup taxa was based on the proposed phylogeny of Johnston in Norton et al. (1993, Figure 1.2). Outgroups are used for rooting the trees, and therefore, assuming the relationships among the outgroups is beyond the scope of this study.
To test the Kethley's relationships between two superfamilies of ant associates, i.e., Antennophoroidea and Aenictequoidea, three species, each from the families of Antennophoridae, Aenictequidae, and Ptochacaridae, were included. For the examination of the monophyly of Paramegistidae, species from all the known genera {i.e., Antennomegistus, Echinomegistus, Neomegistus, Ophiomegistus, and Paramegistus) along with Meristomegistus, a new genus recently described (Kim and Klompen, 2001), were included. Finally, to examine the relationships among the species that exhibit the association with carabid beetles, Micromegistus and Promegistus were included. 130 la additioa to the above sampling scheme focused directly on the testing of Kethley's cladogram, the following strategies were also considered. That is, to examine the relationships among the species within a same family from the same continent, Fedrizzia and Neofedrizzia (Fedrizziidae), passalid- associates from Australia, were included. As a possible back up of investigating this relationship, two species of Klinckowstroemiidae (Klinckowstroemia), another passalid-associated sister family to Fedrizziidae from Central America, were also included. Although these two families are very closely related as seen in the results from morphology (Chapter 3) and molecules (Chapter 4), their biogeographic distribution is quite peculiar, showing no overlap in the global distribution (Hunter, 1993a).
In an expansion of this sampling scheme, two carabid associates, Micromegistus bakeri (Parantennulidae) from North America and M. gourlayi from Australia were included to investigate the relationships between the taxa of the same genus (family) that distribute in the different continents. In addition, to examine the relationships between two closely related species (genera) that are associated with two different host groups, passalid-associated Euzercon latus (Euzerconidae) and millipede-associated Neoeuzercon sp. (Euzerconidae) were included to the study.
Morphological study was performed based on the slide-mounted specimens prepared hrom hresh materials and those h*om the collections housed at the Acarology Laboratory (OSAL), Museum of Biological Diversity, Department of Entomology, The Ohio State University. For the character selection, coding, and description, see the Chapter 3 for the details. Molecular procedures and the voucher numbers (see Table 4.1) of the taxa used for the molecular study were described in the Chapter 4.
131 Superfamily and Family Species Data SUBORDER CERCOMEGISTINA
S.F. Cercomegistoidea
Astemoseiidae Astemoseius sp. M, 18S, 28S Cercomegistidae Cercoleipus coelonotus M,18S, 28S Cercomegistus evonicus M Davacaridae Davacarus gressitti M Pyrosejidae Pyrosejm sp. M Pyrosejidae n. sp. M Saltiseiidae Saltiseius sp. M Seiodidae Seiodes sp. M SUBORDER ANTENNOPHORINA
S.F. Celaenopsoidea
Celaenopsidae Pleuronectocelaeno drymoecetesM Diplogyniidae Cryptometastemum sp. M,18S Ophiocelaeno sellnicki M Passalacarus sylvestris M, 18S,28S Trichodiplogynium sp. M, 28S Euzerconidae Euzercon latus M, 18S, 28S Neoeuzercon sp. M, 18S, 28S Megacelaenopsidae Megacelaenopsis oudemansi M Megacelaenopsis sp. M Schizogyniidae Paraschizogynium odontokeri M Triplogyniidae Funkotriplogynium vallei M Tripiogynium sp. M S.F. Parantennuloidea
Parantennulidae Micromegistus bakeri M Micromegistus gourlayi M, 18S, 28S Philodanidae Philodana johnstoni M S.F. Aenictequoidea
Aenictequidae Aenicteques chapmani M Ptochacaridae Ptochacarus sylvestrii M
(Continued)
Table 5.1: List of taxa used for both of morphological and molecular study. S.F. = Superfamily; M = Morphological study; 18S, 28S = Molecular study
132 Table 5.1 (Continued)
S.F. Antennophoroidea
Antennophoridae Antennophorus wasmanni M, 18S, 28S S.F. Fedrizzioidea
Fedrizziidae Fedrizzia sp. M, 28S Neofedrizzia leonilae M, 18S, 28S Klinckowstroemiidae Klinckowstroemia slarri M,18S,28S Klinckowstroemiella blumae M Paramegistidae Antennomegistus caputcarabi M Echinomegistus wheeleri M Meristomegistus vaztjuezus M Neomegistus julidicola M Ophiomegistus sp. M Paramegistus confrater M,18S,28S Promegistidae Promegistus armstrongi M, 28S S.F. Megisthanoidea
Hoplomegistidae Stenostemum truittae M Stenostemum sp. M Megisthanidae Megisthanus floridanus M, 18S, 28S SUBORDER UROPODINA
Thinozerconidae Thinozercon michaeli M Trachyuropodidae Oplitis sp. M, 18S, 28S Poiyaspididae Polyaspis lameliipes M, 18S, 28S Trachytes sp. M SUBORDER SEJINA
Sejidae Sejus sp. M,18S, 28S Epicroseius sp. M, 18S,28S Ichthyostomatogasteridae Astemolaelaps sp. M, 18S, 28S Uropodellidae Uropodella sp. M SUBORDER MICROGYNIINA
Microgyniidae Microgynium rectangulatum M SUBORDER HETEROZERCONINA
Discozerconidae Discozercon sp. H 28S Heterozerconidae Heterozerconidae n. sp. M, 18S, 28S
133 Combined Data
The combined data set includes 55 morphological characters used for Chapter 3, and a total of 1452 characters {i.e., positions of aligned bases and gaps) used for the analyses in Chapter 4. This new data set is composed of a total of 1507 characters, of which 465 positions are informative in parsimony analyses. For the details of each data set, see the Chapters 3 and 4.
Analyses
All of these characters were coded with equal weight. Multistate characters were coded as multistate. All the sequence characters were treated as unordered multistate. In all the analyses, all the gaps in the sequences, derived by indels (i.e., insertions or deletions), were coded as 'missing' as implemented in PAUP* 4.0b8 (Swofford, 2001). Heuristic searches with TBR branch swapping along with random addition of the taxa with 100 replicates were performed. Maximum parsimony, which seeks the minimum number of steps of evolutionary change within the data, implemented in PAUP* was used as the optimality criterion. Branches were collapsed if the maximum branch length is zero.
To assess the confidence level on the deduced phylogeny, 'branch support' (Bremer, 1988) (= 'decay index' sensu Donoghue et al., 1992; Bremer support' sensu Kàllersjô et ai, 1992) estimated by AutoDecay 4.0.2 (Eriksson, 1999) using global reverse constraints of topologies. For combined analyses, the percentage of incongruence length differences (ILD; Farris et al, 1995; Mickevich and Farris, 1981) among each partitioned data set were calculated. Partition-homogeneity tests (PHT; Huelsenbeck et al, 1996) w ith 100 homogeneity repetitions along with 20 sequence replicates incorporated in PAUP* were also performed. 134 RESULTS AND DISCUSSION
Tree Topology and Systematics
From a set of combined data (see Chapters 3 and 4), 180 equally most parsimonious trees (L = 1829 steps; Cl = 0.46; RI = 0.55) were deduced by PAUP* under no topological constraints. A strict consensus tree, allowing polytomies for all the unmatching branches, was obtained (Figure 5.1). The results from the incongruence length difference (ILD) test and partition- homogeneity test (PHT) performed by PAUP* showed no significant difference between morphological and molecular data (Table 5.2).
The topology of strict consensus tree from the combined analyses is simply a tree with molecular trunk with fine morphological branches. In other words, the tree retains the same branching order for each superfamily obtained from molecular analyses, and the additional taxa from morphological study are added onto this tree of molecular framework. As a result, this reinforced topology of 'molecular' tree recognized monophyletic clade of superfamilies as discovered in both separate analyses of previous chapters.
This result indicates that the phylogenetic position of Parantennuloidea, the only disagreement between two separate studies, concords with the results obtained from the molecular study. However, with regard to this result, it should be noted that displaying sister group 135 Partition Length Length ILD PHT Combined Difference Molecule 1551 4 0.26 ns Morphology, 1829 11 0.60 ns Molecule (Morphology, 1829 15 0.82 ns 18,28)
Table 5.2: Test for compatibility among the partitioned data sets, ns: not significant
relationships between Parantennuloidea and Paramegistoidea shown in both molecular and combined studies might have resulted from the 'swamp effect' caused by outnumbered characters in nucleotide sequence data. In fact, among the 9 unambiguously changing characters that support the clade of (Parantennuloidea, Paramegistoidea) in combined analyses, all the 9 characters were derived from the molecules. When the option of almost all possible changes' implemented in MacClade 3.08a (Maddison and Maddison, 1999) was chosen, among the 45 characters that support (Parantennuloidea, Paramegistoidea), 42 characters were derived from molecules.
From the morphological data that separated Parantennuloidea at the base of Antennophorina (see Chapter 3), 4 more steps (267 —> 271 steps) were required to enforce monophyletic (Parantennuloidea, Paramegistoidea), the result obtained from molecular and combined analyses. Conversely, from the combined molecular (18S+28S) data alone (see Chapter 4), 7 more steps
136 Astemoaeius ■ ' " I ' Seiodes Cercoleipus 3 ir~f Cercomegistus iM ju Pyosejus Cercomegistoidea Pyrosejidae n.sp. - a H _ ^ Davacerus Seiliseius Mkro_bekeri Micro_gouilayi P rom e^us Parantennuloidea Philodana Antennomegislus Echinomegistus Ophiomegistus Neomegistus 5 — Paramegistus « Meristomegistus Aenicteques I Aenictequoidea S Ptochacarus Antennophorus I Antennophoroidea Klinckovstroemia KHnckjvictoriae Fedrizzia I Fedrizzioidea Neofedrizzia Megisthanus Stenostemum Megisthenoidea StenoJruRtae Trichodiplogynium Cryptometastemum Passelacerus Ophiocelaeno Pleuronectocelaeno Euzercon Neoeuzercon Celaenopsoidea Megacelaenopsis I Megaceiaenopsis 2 Paraschizogynium Tripiogynium Funkotriplogynium Thinozercon Polyaspis Oplitis Vropodina Trachytes Mkfogynium tMterogyrina Discozercon Heterozerconidae Neterozerconina Astemolaelaps Sejus Epicroseius Sejkw Vropodeila
Figure 5.1: Strict consensus tree from 180 equally most parsimonious trees with no topological constraints. Numbers shown are the values of branch support sensu Bremer (1988).
137 (1551 —> 1558 steps) were required to enforce the basal Parantennuloidea inferred from morphology. Although the numbers of these additional steps are relative and not absolute, empirically, these numbers of additional steps are high enough to well-support each tree discovered from each separate analysis.
The result from the combined analyses recovered the monophyly of Trigynaspida within Mesostigmata, supporting the bifurcating lineages of Cercomegistina and Antennophorina assumed by Kethley (1977b). In overall, however, this result is different from his in terms of two facts: recognition of the monophyletic grouping of superfamilies and the relationships among them. As to the recognition of monophyletic superfamilies, Kethley's superfamily of Fedrizzioidea, which he included Fedrizziidae, Klinckowstroemiidae, Promegistidae, and Paramegistidae, was no longer supported as these families are not monophyletic (Figure 5.1). That is, while the two sister families of Fedrizziidae and Klinckowstroemiidae, passalid associates, can be remained in this superfamily, Promegistidae and Paramegistidae require the shift of the superfamilies to which they belong.
Promegistidae, represented by a single known species of Promegistus armstrongi Womersley that is associated with Pamborus (Coleoptera: Carabidae) in Australia, become the sister to the two species of Micromegistus (Parantennuloidea: Parantennulidae), carabid-associates from North America and Australia. Along with another known parantennuloid, Philodana johnstoni Kethley (Philodanidae), an associate of Neatus tenebrioides (Beauvois) (Coleoptera: Tenebrionidae), this new grouping forms a clade of extended superfamily of Parantennuloidea. Unlike Fedrizziidae and Klinckowstroemiidae that retain an entire (= impaired) stemogynial shield, heavily sclerotized comiculi, and robust dentate cheliceral digits, Promegistidae retain no stemogynial shield, more or less membranous base 138 of the comiculi, and tapered edentate chelicerae, which are characteristic of the members of Parantennuloidea.
The remaining family of Kethley's Fedrizzioidea, i.e., Paramegistidae, formerly treated as paraphyletic in his study, stands out as a monophyletic clade, sister to the new Parantennuloidea. Unlike Fedrizziidae, Klinckowstroemiidae, or Promegistidae, all of which exhibit the associations with the beetles, most of the species of Paramegistidae are associated with diplopods or squamates (see Table 2.1).
The discovery of the non-monophyly of Fedrizzioidea sensu Kethley (1977b) and the suggested relocation of Promegistidae and Paramegistidae from the combined analyses were also supported by the separate morphological and combined molecular analyses described in previous chapters. When the Kethley's Fedrizzioidea was enforced from the combined data set, 22 additional steps (i.e., 1851 steps), carrying 330 equally most parsimonious trees, were required. A strict consensus tree from these enforced trees showed trichotomy within Trigynaspida, carrying Cercomegistina, Parantennuloidea, and the remaining Antennophorina, rendering phylogenetic uniqueness of Parantennuloidea.
With regard to the relationships between Parantennuloidea and Paramegistidae, although these two groups can be combined together to form a single superfamily of, possibly, Parantennuloidea sensu lato, I suggest to recognize two separate superfamilies by raising Paramegistidae to Paramegistoidea (see Chapter 3). This proposal reflects the unique phylogenetic position of Parantennuloidea discovered in morphological analyses, in which this superfamily was unique at the base of Antennophorina, while Paramegistoidea becomes the sister to the monophyletic clade of (Aenictequoidea, Antennophoroidea). A few 139 morphological characters supporting^ Paramegistidae (Paramegistoidea), such as paired stemogynial shields or male eugenital setae, are not present in Parantennuloidea, including Promegistidae. In addition, as stated above, each group of Parantennuloidea and Paramegistoidea stands alone as a monophyletic clade in all the analyses, allowing separate treatment of each unique superfamily (see Table 5.3).
Apart from the discrepancy in the relationships of Parantennuloidea and Paramegistoidea between this study and the Kethley (1977b) discussed above, another difference comes from the two groups of strict ant-associates, Antennophoroidea and Aenictequoidea. While these two monophyletic groups were sisters in both morphological and combined studies, they were treated as disparate groups in Kethley (1977b), not sharing their immediate ancestor. In fact, these ant-associates are not that different in terms of morphology, as they share the characters of short peritremes, eugenital setae on male genital opening, and comiculi with small protuberance(s) or serration. It should be mentioned that, however, the sister group relationship between Antennophoroidea and Aenictequoidea shown in this combined study is entirely derived from such morphological characters as no molecular data from Aenictequoidea were involved in the analyses. Retaining the name of two current superfamilies is, therefore, suggested.
In overall, the results from this study suggest a new systematic paradigm of monophyletic Trigynaspida, containing two suborders of Cercomegistina and Antennophorina. While the former is composed of a single superfamily Cercomegistoidea, containing 6 families, the latter is composed of 7 superfamilies, i.e., Parantennuloidea, Paramegistoidea, Antennophoroidea, Aenictequoidea, Fedrizzioidea, Megisthanoidea, and Celaenopsoidea, containing a total of 21 families.
140 Systematics and Host Associations
Studies on the evolutionary relationships among the mites and their hosts are another aspect of systematics. Although the details may require the inputs from other disciplines, such as chemical ecology, behavioral biology, or physiology, a phylogenetic tree can serve as a useful framework for understanding the evolutionary patterns of associations. In fact, nothing is known about the chemical ecology of trigynaspid mites and their hosts, and little detail is available on the behavior and physiology of the hosts and mites.
Trigynaspida exhibits peculiar biological features, in which all the immatures, excluding Micromegistus (Parantennulidae), are free-living, while most of the adults, especially in Antennophorina, are associated with their hosts, such as passalids, scolytids, carabids, formicids, diplopods, or squamates (Hunter and Rosario, 1988; Kinn, 1971; Seeman and Walter, 1997). Immatures are commonly found in moist and decaying vegetation around the hosts, and they are saprophagous to predatory in feeding (Butler and Hunter, 1968; Hunter and Davis, 1965; Hunter and Rosario, 1988). Based on the information from the ontogeny and phylogeny, 1 assume that the ancestral Trigynaspida was a free-living predator, dwelling in moist and decaying environment feeding on nematodes, coUembolans, or other insects' eggs. From this environment, evolution of phoretic associations, allowing dispersal, is thought to be established.
Among the 8 superfamilies proposed in this study, nearly all the members of Cercomegistoidea are free-living in both immature and adult stages. Exceptions are found in Cercomegistidae, in which the adults show phoretic associations with the bark beetles (Kinn, 1971), social spiders (Evans,
141 1958), or pagurid crabs (André, 1937). All the cercomegistoids have robust and dentate chelicerae, supporting their predatory feeding behavior.
The patterns of host association become more diverse in the remaining 7 superfamilies that collectively constitute Antennophorina. Except for a few free-living groups {i.e., Triplogyniidae, Megacelaenopsidae, and some Schizogyniidae), nearly all the adults of Celaenopsoidea display the associations with their hosts, mostly with the passalid beetles. Deviations from this passalid association are found in some Celaenopsidae and Schizogyniidae (scolytid associations); and in a few Diplogyniidae, Euzerconidae, and Costacaridae (diplopod associations). Although most members of Diplogyniidae, the most diverse family in terms of known genera and species, are associated with passalids as adults, a few also show the associations with scarabaeids, histerids, or cockroaches. Despite these deviations within Celaenopsoidea, passalids still serve as the major hosts in terms of the number of genera and species of trigynaspid mites (Hunter, 1993a). In fact, among the 123 known species of Celaenopsoidea, about 90 species show the associations with passalid beetles at the adult stage. From the basal free-living and passalid associations, multiple independent associations toward non-passalids are thought to be evolved within Celaenopsoidea. In fact, the highest degree of diversification of trigynaspids appearing in Celaenopsoidea is thought to be triggered by the shifts of their phoretic host, dwelling in moist and decaying habitats.
Two super families of Megisthanoidea (containing Megisthanidae and Hoplomegistidae) and Fedrizzioidea (now only containing Fedrizziidae and Klinckowstroemiidae) are also associated with passalids. Along with Celaenopsoidea, these two superfamilies display basal passalid associations within the Antennophorina in the phylogeny (Figure 5.2).
142 Astemoseius Seiodes Cercoleipus Cercomegistus FreeHMngor Pvrosejus Scolytids Pyrosejidae n.sp. Davacarus Saltiseius Micro_bakeri Micro_gout1avi Promegistus Carabids Philodana Antennomegistus Echinomegistus Diplopods, Ophiomegistus Neomegistus Reptles, Paramegistus Carabids Meristomegistus Aenicteques I Formicids Ptochacarus Antennophorus I Formicids Klinckovstroemia Klinck_victoriae Fedrizzia I Passalids Neofedrizzia Megisthanus Stenostemum Passalids Stenojruittae Trichodiplogynium Cryptometastemum Passalacarus Ophiocelaeno Pleuronectocelaeno mostly Passalids, Euzercon Scolytids, Neoeuzercon FreeHMng Megacelaenopsis 1 Megacelaenopsis 2 Paraschizogynium Tripiogynium Funkotriplogynium Thinozercon Polyaspis Oplitis Trachytes Microgynium Discozercon Heterozerconidae Astemolaelaps Sejus Epicroseius Uropodella
Figure 5.2: Strict consensus tree from combined morphology and molecules with host information.
143 Current Fedrizzioidea Host Proposed Superfamily Fedrizziidae Passalids Fedrizzioidea Klinckowstroemiidae Passalids Fedrizzioidea Paramegistidae Millipedes, Reptiles, Paramegistoidea, NEW Carabids Promegistidae Carabids Parantennuloidea
Table 5.3: Proposed classification for currently known Fedrizzioidea. This proposal is corroborated by the morphological, molecular, and combined analyses.
The pattern of strict association applies to Antennophoroidea and Aenictequoidea, which are associated with ants. The associations in the remaining superfamilies, i.e., Paramegistoidea and Parantennuloidea, were discussed above and summarized in Table 2.1 and 5.3. Although there are minor exceptions of Echinomegistus and its close sister Antennomegistus (Paramegistidae) that are associated with carabid beetles, it needs to be noted that the members of Paramegistoidea do not share the same host range with those of Parantennuloidea in general.
Biogeography and Origin
The facts that most trigynaspids are found in the continents of former Gondwanaland and that they are either free-living (as in most Cercomegistoidea, and some Celaenopsoidea) or associated with mostly non volant gregarious hosts indicate that the origin (age) of trigynaspids is linked to the age of the southern continents and global distribution of the hosts,
144 which are primarily passalids» Passalids^ which serve as the major hosts for basal antennophorine lineages in the phylogeny (Figure 5.2), are also thought to be former Gondwanaland in origin (Pedro Reyes-Castillo; Alan Gilogly; Chris Marshall; pers. comm.).
Accordingly, without having a direct evidence of fossil record of Trigynaspida, it is possible to estimate the age of these mites based on the information from the plate tectonics (vicariance) and the biogeography of trigynaspids and their hosts. Based on the information and the phylogeny, it is assumed that the origin of free-living Cercomegistina is in, at the latest. Upper Triassic period of Mesozoic era (ca. 225-200 million years ago) when the supercontinent Pangaea broke into northern Laurasia and southern Gondwanaland (Kious and Tilling, 1994). By Jurassic, when this former Gondwanaland became separated into South America, Africa, India, and Antarctica-Australia complex, the evolution of the major groups in Cercomegistina and Antennophorina that display the host associations is thought to be already established. Although it is almost certain that the free- living trigynaspids appeared earlier than those showing the associations, there is no direct evidence on the starting point of the acquisition of host association or the evolution of phoresy found in most Antennophorina. However, as the host associations occur in the adult stase and are thought to be obligatory, the appearance of phoretic trigynaspid groups should address the initiation of host association.
Although the only known passalid fossil is from the Cenozoic Oligocene deposits (Reyes-Castillo, 1977), the paleogeography of this period shows the complete separation of the Gondwanaland (Kious and Tilling, 1994; Mintz, 1977). Accordingly, assuming the acquisition of passalid association at this period requires multiple independent associations toward passalids on these already separated continents. In other words, it is more 145 plausible to speculate the starting, point of the evolution of phoresy in, at the latest, the Jurassic, when the continents and these flightless animals are still combined together. Furthermore, the passalids are also thought to be at the latest Mesozoic (Pedro Reyes-Castillo, pers. comm.) or even Paleozoic (Alan Gilogly, pers. comm.) in origin.
Another support to the speculation of the Mesozoic origin of Antennophorina comes from the stenoxenic ant associates, Antennophoroidea and Aenictequoidea. Although these two superfamilies are closely related in the phylogeny (Figure 5.2), they do not share the same distribution area in the globe. While the members of Aenictequoidea appear in the eastern Australia, Papua New Guinea, and the Philippines, those of Antennophoroidea appear in western Europe and North America, better supporting vicariance than dispersal. Information on the Jurassic (Crozier et ai, 1997) or the Cretaceous (Grimaldi et al., 1997) origin of ants along with the time of separation of Europe and North America (ca. 135-65 mya) also supports, at the latest, the Mesozoic origin of these mites.
Current biogeographic distributions of the three species of Micromegistus (Parantennuloidea: Parantennulidae) that exhibit the association with carabids, i.e., M. bakeri from North America, M. viduus from South America (not included in this study), and M. gourlayi from Australia, may also support the above speculation on the origin. Although carabids are more active than passalids, the peculiar global distribution patterns of these mites on carabids in three different continents still prefers passive vicariance over active dispersal, allowing the estimation of the age of these mite at the latest to the Mesozoic. Dispersal of non-volant animals in the terrestrial environment can be better supported within a continent or within a supercontinent (i.e., Pangaea).
146 This effort to estimate the origin and the age of trigynaspid mites is important as this estimation reduces the significant gap of the origin (age) between two different acarine lineages; Acariformes and Parasitiformes. Although the formation, preservation, and discovery of fossils are rare events which involve in chance, and although this is especially true for the tiny mites, there is a gap of time in terms of the oldest fossil records between Acariformes and Parasitiformes. That is, while there are several fossil records of acariform mites beginning from the Devonian {ca. 405-400 mya) (Bernini, 1991; Hirst, 1923; Norton et al., 1988; Selden, 1993; Trewin, 1994), the earliest known fossil of Parasitiformes is only from the Upper Cretaceous (ca. 94-90 mya) (Klompen and Grimaldi, 2001), leaving the gap of time of about 315-306 million years. It would be natural to have the same or close time of origin between Acariformes and Parasitiformes, if the Acari is monophyletic. Accordingly, if the estimation of the age of Trigynaspida proposed herein, based on the information from the biogeography of mites and their hosts, phylogeny, and the plate tectonics, is legitimate, the gap of time between the two acarine lineages is much narrowed by only about 175-212 million years.
147 APPENDIX A. Aligned NS5-NS8 region of the 18S rRNA sequences.
31 ) [ 2' ] [ 32 ] [ 33 ] [ 3 4 Astemoseius AACUU)AA[AGGA]A U [UGACGGAAGGGCA]CCACCA- [GGAGU]GG[ -GCCUG Cercoleipus AACUU ) AA [AGGA] AU [UGACGGAAGGGCA] CCACCA- [GGNGU] GG[AGCNUG Micromegistus AACUU ) AA [AGGA] AU [UGACGGAAGGGCA] CCACAA- [AGCGU1GG[AGCUUG Paramegistus AACUU)AA[AGGA]A U [UGACGGAAGGGCA]CCACCA- [GGAGU]GG[AGCCUG Antennophorus AACUU ) AA [ AGGA ] AU ( UGACGGAAGGGCA ) CCACCA- [GGCAU]GG[AGCCUG Klinckowstro AACUU ) AA [ AGGA ] AU [ UGACGGAAGGGCA ] CCACCA- [GGAGU]GG[AGCCUG Neofedrizzia AACUU ) AA [ AGGA ] AU [ UGACGGAAGGGCA ] CCACCA- [GGAAU]GG[AGCCUG Megisthanus AACUU) A A [AGGA] A U [UGACGGAAGGGCA] CCACCA- [GGAGU]GG[AGUCUG Cryptometast AACUU ) AA [ AGGA ] AU [ UGACGGAAGGGCA ] CCACAAU [GGAGU]GG[AGCCUG Passalacarus AACUU) AA [AGGA] AU [UGACGGAAGGGCA] CCACAA- [CGGAG]UG[GGCCUG Euzercon AACUU) AA [AGGA] AU [UGACGGAAGGGCA] CCACAA- [GAAGU]GG[AGC-UG Neoeuzercon AACUU) AA [AGGA] AU [UGACGGAAGGGCA] CCACAA- [GAAGU]GG[AGC-UG P o lya sp is AACUU) AA[AGGA] A U [UGACGGAAGGGCA] CCACCA- [GGAGU]GG[AGCCUG O p litis AACUU) A A [AGGA] A U [UGACGGAAGGGCA] ACACAA- [GGAGU]GG[AGCUUG Asternolaelap AACUU ) AA [ AGGA ] AU [ UGACGGAAGGGCA ] CCACCA- [GGAGU]GG[AGCCUG Epicroseius AACUU) A A [AGGA] A U [UNACGGAAGGGCA] CUACCA- [GGAGU]GG[AUUGUG S eju s AACUU) AA[AGGA]A U [UGACGGAAGGGCA]CCACCA- [GGAGU] GG[AGCAUG Heterozerconid AACUU ) AA [ AGGA ] AU [ UGACGGAAGGGCA ] CCACCA- [GGAGU]GG[AGCCUG
1 ( 35 ) [ 36 ] ( Astemoseius CGGC]UUAA(UUUGACUCAA-CA-CGGA)GAAACUUA[CUCGGCC]CGGA(CUC Cercoleipus CGGC ] UNAA ( UUUGANUCAA-CN-NGGG ) AAA-CUUA [CCCGGCC ] CGGA ( CAC Mi cromegistus CGGC ] UUAA ( UUUGACUCAA-CA-CGCG ) AAAUCUUA [CCAGUGC ] AAGA (CAC Paramegistus CAGC] UUAU(UUUGACUCAA-CA-CUAG)AAAACCUA[CCCGGUC]CGGA(CAC Antennophorus CGGC ] UUAA ( UUUGACUCAA-CA-CGGG ) AAAACUUA [CCCGGCC ] CGGA ( CAC K1inckowstro CGGC I UUAA (UUUGACUCAA-CA-CGGG ) AAAACUUA [CCCGGCC ] CGGA ( CAC Neofedrizzia CGGC] UUAA (UUUGACUCAA-CA-CGGG) GAAACUUA [CCCGGCC] CGGA (CAC Megisthanus CGGC ] UUAA ( UUUGACUCAA-CA-CGGG ) AAAACUUA (CCCGGCC] C(3GA (CAC Cryptometast CGGC] UUAA (UUUGACUCAA-CG-CGGG) AAACCUUU [CCCGGCC] CGGA(CAU Passalacarus CGGC] UUAA (UUUGACUCAA-CA-CGGG) AAAACUUU [CCCAGGC] CGGA (CAC Euzercon CGGC ] UUAA (UUUGACUCAA-CA-CUGG) GCACCUCA [CCUGGUC ] CUGA (CAC Neoeuzercon CGGC ] UUAA ( UUUGAUUCAA-CAUUGGG ) CAA-CUCA [ CCUGUUC ] CUGA ( CAC P o lya sp is CGGC ] UUAA ( UUUGACUCAA-CA-CGGG ) GAAACUCA [CCCGGCC ] CGGA ( CAC O p lit is CNGC ] UUAA ( UUUGAUUCAAACAACGGG ) AAAUUUUG [ CCCAGCC ] GAGA ( CAC Astemolaelap CGGC ] UUAA ( UUUGACUCAA-CA-CGGG ) AAAACUUA [ CCCGGCC ] CGGA ( CAC Epicroseius CGGC] UUAA(UUUGACUCAA-CA-CGGG) AAAACUUA[CCCGGCC] CGGA(CAC S eju s CGGC ] UUAA ( UUUGACUCAA-CA-CGGG ) AAACCUCA [ CCCGGCC ] CGGA ( CAC Heterozerconid CGGC] UUAA (UUUGACUCAA-CA-CGGG) AAAACUUA [CCCGGCC] CGGA (CAC 148 37 ) [ 38 Astemoseius CGGGAGGAÜUGACAGAUUGAGAGCUCUUUCAUGAUUCGGCG)GAU[GGUGGUGC Cercoleipus UGUNNNGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCAAUG)GAU[GGUGGUGC Micromegistus UCGAAGGAUUGACAGAUUAAGAGCUCUUUCUUGAUUGAGUG) GAA[GGUGGUGC Paramegistus UGGGAGGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCGGUG) GAU[GGUGGUGC Antennophorus UGGGAGGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCGGUG)GAG[GGUGGUGC Klinckowstro UGGGAGGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCGGUG) GAU[GGUGGUGC Neofedrizzia UGGGAGGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCGGUG)GAU[GGUGGUGC Megisthcinus UGGGAGGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCGGUG ) GAU [GGUGGUGC Cryptometast UAGGACGAUUGACGGAUUAAUAGCUCUUUCUAGAUUNGGUG) GAU [AGUGGUGC Passalacarus UGGAAGGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCUGUG)GAU[UGUGGUGC Euzercon UGGGAGGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCGGUG)CAU[AGUGGUGC Neoeuzercon UGGGAGGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCGGUG) UAU [AGUGGUGC P o ly a sp is UGAGAGGAUUGACAGAUUGAGAGCUCUUUCUUGAUUCGGUG) GAU[GGUGGUGC O p litis UGCAUGGAUUGACAGAUUAAGAGCUCUUUCUUGAUUCGGUG) GUU[GGUGGUGC Astemolaelap UGGAAGGAUUGACAGAUUGAUAGCUCUUUCUUGAUUCGGUG) GAU [GGUGGUGC Epicroseius UGGAAGGAUUGACAGAUUGACAGCUCUUUCUUGAUUCAGUG)GAU[GGUGGUGC S ejus UGGAAGGAUUGACAGAUUGACAGCUCUUUCUUGAUUCGGUG)GAU[GGUGGUGC Heterozerconid UGGAAGGAUUGACAGAUUGAUAGCUCUUUCUUGAUUCGGUG)GAU[GGUGGUGC
38 ] [39 ] ( 40 ) ( 41 ) Astemoseius AUGGCCGUUCAU] A [GUU] (CGUGGAAUUGAUCCGUC) (UGGUUA--UCCG)AC Cercoleipus AUGGCCGUUCUU]C[GUU] (CGUGGAGU-AAUCCGGC) (UGGUUAAUUCCG) AU Mi cromegistus AUGGCCGUCUCU]A[GUU] (GGUGAACUUGAUUUGUC) (UGCUUGAUUGCG) GU Paramegistus AUGGUCGUUCAU] A[GUU] (GGUGGAGC-GAUUUGUC) (UGGUUUAUUCCG) AU Antennophorus AUGGCCGUUCUU]A[GUU] (GGUGGAGC-GAUCUGUC) (UGGUUUAUUCCG) AU K1 inckowstro AUGGCCGUUCUU] A[GUU] (GGUGGAGC-GAUCUGUC) (AGGUUCAUUCCG) AU Neofedrizzia AUGGCCGUUCUU] A[GUU] (GGUGGAGC-GAUCUGUC) (UGGUUAAUUCCG) AU Megisthanus AUGGCCGUUCUU] A[GUU] (GGUGGAGC-GAUCUGUC) (UGGUUAAUUCCG) AU Cryptometast AUGGCCGUUCUU1A [GUU] C CGUGGAGU-GAUUUCUC) ( AGGUCAAUUCCG)AU Passalacarus AUGGCCGUUCUU] A[GUU] (GGUGGAGC-GAUCUGUC) (UGGUUAAUUCCG) AU Euzercon AUGGCCGUUCUU]A[GUU] (CGUGGAGC-GAUCCGUC) (UGGUUAAUUCCG) AU Neoeuzercon AUGGCCGUUCUU] A [GUU] (CGUGGAGU-GAUCCGUC) (UGGUUAAUUCCG) AU P o lya sp is AUGGCCGUUCUU] A[GUU] (GGUGGAGC-GAUCUGUC) (UGGUUAAUUCCG) AU O p litis AUGGCCGUUCUU] A[GUC] (GGUGGAGU-GAUUUGUC) (UGGUUUAUUCCG) UU Astemolaelap AUGGCCGUUCUU]A[GUU] (GGUGGAGC-GAUCUGUC) (UGGUUAAUUCCG) AU Epicroseius AUGGCCGUUCUU] A [GUU] (GGUGGAGC-GAUCUGUC) (UGGUUCAUUCCG) AU S eju s AUGGCCGUUCUU] A[GUU] (GGUGGAGC-GAUCUGUC) (UGGUUAAUUCCG) GU Heterozerconid AUGGCCGUUCUU] A [GUU] (GGUGGAGC-GAUCUGUC) (UGGUUAAUUCCG) AU
149 [39'] [ 42 ] ( 43 As tem osei us AACG [AAC ] CAGA [CUCU] AG ( CCUAUUAAAUAGUCGCGGUGUC------GAAAGC- Cercoleipus AACG [AAC ] GAGA [CUCU] AN ( CCUACUAACUAGUCGUCA-GUGU-GGAACA— Micromegistus AACG [AAC ] GAGU [UCCC] AG (CCUAUUAAAUAGUCGCGA-AUGU— GGAUAU- Paramegistus AACG [AAC] GAGA [CUCU] AG (CCUAUUAAAUAGUCGCCA-GUAUUACGCAAC- Antennophorus AACG [AAC] GAGA [CUCU] AG (CCUAUUAAAUAGUCGCCA-GGGUCACGCAGC- K1 inckowstro AACG[AAC] GAGA[CUCU]AG( CCUAUUAAAUAGUCUCUC-GCGUUACGCAGC- Neofedrizzia AACG[AAC]GAGA[CUCU]AG(CCUAUUAAAUAGUCUCUC-GCGUUACGCAGC- Megisthanus AACG [AAC ] GAGA [CUCU] AG ( CCUAUUAAAUAGUCGCGU-GGGUCACGCAGC- Cryptometast AACG [ GAC ] GAGA [ UCCU ] AG ( CCUACUAACUAGGGGCUU-GCCC— UU------Passalacarus AACG [AAC] GAGA [CCCU] AG ( CCUAUUAAAUAGACUCGC-ACGU-AUUCUGC- Euzercon AACG [AAC ] GAGA [CUCU] AU ( CUUACUAACUAGUCU-GC-GCGU-AU-CAGC- Neoeuzercon AACG [AAC ] GAGA [CUCU] AU ( CUUACUAACUAGUCUGCG-CGUA--C-CNVGCC P o lya sp is AACG [ AAC ] GAGA [CUCU ] AG ( CCUAUUAAAUAGACAAGA-UGUU— UAUGAC- O p litis AACG [GAC] GAGA [UUCU] AA (CCUACUAAAUAGACGUGG-UGUU-GAUUUGC- Asternolaelap AACG [AAC ] GAGA [CUCU] AG ( CCUAUUAAAUAGACGUGA-UGGU-CUUUUGC- Epicroseius AACG [AGC ] GUGA [CUCU] AG (CCUAUUAAAUAGACGGAG-GGUU-UGUACAC- S ej us AACG [AAC ] GAGA [CUCU] AG ( CCUGCUAAAUAGGGGCGG-AGAUAUUUAUUC- Heterozerconid AACG [AAC ] GAGA [UUCU] AG ( CCUAUUAAAUAGGCGCGA-GCCU— CUUCGC-
43 ) ( 44 ) Astem oseius CACC - — CCGUGU ACUUCUUAGAGG ) - ACAA ( UGUGACUCUAGCACA ) AUG Cercoleipus CACCC - - ACGUGUACGACUUCUUAGAGG ) GACAG ( GGUGCUUUUAGCACC ) AGG Mi cromegistus CCUCCAAUCGCGC ACUUCUUAGAGG ) GACGA (GGUGCUUCCAGCAUC ) GCG Paramegistus GAC------UGGCGU------ACUUCUUAGAGG ) GACAA (GGUGCUUUCAGCACC ) ACG Antennophorus CCC------UGGCGU------ACUUCUUAGAGG ) GACAA (GGUGCUUUCAGCACC ) GCG Klinckowstro CGC------GAGCGU------ACUGCUUAGAGG ) GACAA ( GGUGCUUUCAGCACC ) GCG Neofedrizzia CGC------GAGCGU------ACUGCUUAGAGG ) GACAA (GGUGCUUUCAGCACC ) GCG Megisthanus c u e ------ACGCGC------ACUUCUUAGAGG ) GACAA ( GGUGCUUUCAC3CACC ) GCG Cryptometast -GC ------GGCUCGCC -CCUUCUUAGAGG ) GACAA (GGUGCUUCUAGUACC ) GUG Passalacarus CAA------GCGAGU------UCUUCUUAGAGG ) GACAA (GGUGCUUUCAGCACC ) GUG Euzercon CGC------GUGCAG------ACUUCUUAGAGG ) GACGA (GGUGCUUCCAC3CACC ) GCG Neoeuzercon AGC------GUGCAG------ACUUCUUAGAGG ) GACGA (GGUGCUUCCAGCACC ) GCG P o lya sp is ACA------UCUUGCA- -UCUUCUUAGAGG ) GACAA ( GGUGCUUUCAGCACC ) ACG O p litis ACA------UCACG-AUUCCUUCUUAGAGG ) GACAC (GUUGCUUAUAGCUAC ) ACG Asternolaelap CCG------UCACGU------CCUUCUUAGAGG ) GACAA (GGUGCUUUUAGCACC ) GCG Epicroseius GCC------CUACGU------UCUUCUUAGCGG ) AACAA (GGUGCUUUUAGCACC ) ACG S eju s UCU------CCGUGC------CCUUCUUAGAGG ) GACAA (GGUGCUUUUAGCACC ) GCG Heterozerconid GGC------UCGCGC------UCUCCUUAGAGG ) GACAA (GGUGCAAUUAGCACC ) ACG
150 [ 42'] [ 38' ] [ 3 6 ' ] Astemoseius AAAG[AGAG]C[UAUAACAGGUCUGUGAUGCCCU] UAGAU[GUUCGGG]GCC Cercoleipus AAGA[AGAG]C[AAUAACAGGUCUGUGAUGCCCU] ACGAU[GUUCGGG]GCG Mi cromegistus AAAC[GGGG] U [CACGACAGGUCUGUGAUGCCCC] AAGAU[GUACUGG]GCC Paremegistus AAAC[AGAG]C[AACAACAGGUCUGUGAUGCCCU] AAGAU[GUCCGGG]ACC Antennophorus AAAA[AGAG] C (AAUAACAGGUCUGUGAUGCCCU] UAGAU[GUCCGGG]GCC Klinckowstro AAAC[AGAG]C[AAUAACAGGUCUGUGAUGCCCU] UAGAU[GUCCGGG]GCC Neofedrizzia AAAC[AGAG]C[AAUAACAGGUCUGUGAUGCCCU] UAGAU [ GUCCGGG ] GCC Megisthanus AAAC[AGAG]C[AAUAACAGGUCUGUGAUGCCCU] UAGAU[GUUCGGG]GCC Cryptometast AGAA[AGAG]C[AAUAACAGGUCUGUGAUGCCCU] CAGAU[GUCCGGG]GCC Passalacarus AAAC[AGAG]C[AAUAACAGGUCUGUAAUGCCCU] ACGAU[ACCUGGG]GCC Euzercon AAAA[AGAG]C[GAUAACAGGUCUGUGAUGCCCU] UAGAU[GUUCAGG]GCC Neoeuzercon AAAA[AGAG]C[GAUAACAGGUCUGUUAUGCCCU] UAGAU[GUUCAGG]GCC P o lya sp is AAAC[AGAG]C[AAUAACAGGUCUGUGAUGCCCU] UAGAU[GUCCGGG]GCC O p litis AAAA[AGAG]C[AAUAACAGGUCUGUGAUGCCCU] UAGAU[GUCUGGG] ACC Astemolaelap AAAC[AGAG]C[AAUAACAGGUCUGUGAUGCCCU] UAGAU[GUCCGGG] GCC Epicroseius ACAA[AGAG]C[AAUAACAGGUCUGUAAUGCCCU] UAGAU[GUCCGGG]GCC S eju s AAAC[AGAG]C[AAUAGCAGGUCUGUGAUGCCCU] UAGAU[GUCCGGG] GCC Heterozerconid AAAC[AGAG]C[AAUAACAGGUCUGUGAUGCCCU] UAGAU[GUCCGGG]GCC
[ 34' ] [ 45 ] ( Astemoseius [GCACGCGCGCC] ACAA[UGCUGGA]GUAAGC-GUGU--UGAGAU(CCUUGCUC Cercoleipus [GCACGCGUGCU] ACAC [UGCGAGA] GGAAACAGGGUA------{ACUUGUUC Micromegistus [GCACGCGCGUU] ACAA[UGAAGGA] AUCAGC-GUGU-GUGCAA-(CCUAGUUC Paramegistus [GCAUGUGCGCU]ACAC[UGAUGGA]AGAAGC- GUCUCGUGUU-- (CCUAGUUC Antennophorus [GCACGCGCGCU]ACAC[UGAUGGA]AAAAGC-GUGUCGUGUU— (CCUAGUUC Klinckowstro [GCACGCGCGCU] ACAC [UGAUGGA] AAAAGC-GUGUCGUGUU— (CCUAGUUC Neofedrizzia [GCACGCGCGCU] ACAC [UGAUGGA] AAAAGC -GUGUCGUGUC - - (CCUUGUUC Megisthanus [GCACGCGCGCU] ACAC [UGCUGGA] AUAAGC-GUGUCGUGUUU- (CCUAGUUC Cryptometast [ GCACGCGGGCU1 ACAC [ UGAAGGG] GGGAAC-GAGACGG-ÜUU- ( CCUGGCUC Passalacarus [GCACGCGCGCC] ACAC [UGAGGGG] ACAAAC-GAGUCGGCUG— (CCUUGCUC Euzercon [GCACGCGCGCU] ACAA [UGGGGCA] AGCAGC-GUGUCGGUGAG- (CCUAGCCC Neoeuzercon [GCACGCGCGCU] ACAA [UGGGGCA] AACAGC-GUGUCGGUGAG- (CCUAGCCC P o lya sp is [GCACGCGCGCU]ACAC[UGAAGAA]AGAAGC-GUGU--GUUUUC(CCUUGUUC O p litis [GCACGCGUGCU] ACAC [UGAAGGA] AGCAGC- GUGUA-GAAU-- (CCUGGUUC Asternolaelap [GCACGCGCGCU] ACAC [UGAAGGA] AGCAGC-GUGUGAUUUU— (CCUUGACC Epicroseius [GCACGCGCGAU] ACAC [UGAAGGA] AUCAGC-GUGUAAUAAAC- (CCUGGUUU S eju s [GCACGCGCGCU] ACAA [UGAAGGA] ACGAAC-GUGGCACCGAC- (CCUCGCAU Heterozerconid [GCACGCGCGCU] ACAC [UGAAAGA] AGCAGC-GUGUCGUUUU— (CCUCGUCC
151 4 6 ) [45' ] ( AsCernoseius GAGGGAGCAGGG) CAACCCCUUGAACC [UCUCUCA] UGAUUGGNAU (AGGGGCU Cercoleipus GACAGAACUGGU) GAAACCUGUUAAAC [UCUCGCA] CGAUUGGGAU ( UGGGAUU Mi cromegistus GAAUGAACUGGG) UAACCCAGUGAGUG [UUCUUCA] CGAUUGGGAU (AGUGGAU Paremegistus GAACGAACUGGG ) AGCCCCA- UGAAGU [ UCCUUCA ] UGAUUGGG-U (? ? ? ? ? ? ? Antennophorus GAAGGAACUGGG) UAACCCAAUGAAGC [UCCUUCA] UGAUUGGGAU (AGGGGCU Klinckowstro GAACGAACUGGG ) UAACCCCAUGAAGU [UCCUUCA] UGAUUGGGAU (AGGGGCU Neofedrizzia GAACGAACUGGG) UAACCCCAUGAAGU [UCCUUCA] UGAUUGGGAU (AGGGGCU Megisthanus GAACGAACUGGG ) UAACCCAAUCAAGU [ UCCGGCA ] UGAUUGGGAU ( AGGGGUU Cryptometast CUCGGGGCUGGG) CGACCUCUUCAACC [CCCUUCU] UAGUGGGAAC (AGGGGCU Passalacarus UUGCGAGCUGGG) UGACCUCUUGAAGU [UCCCUCA] UGGUGGGGAU (ANGNGCU Euzercon GAAGGGGCCGGG) UAACCCCG-GAACC [UCCCUCA] UGCUCGGGAU ( AGAGGAU Neoeuzercon GAAGGGGCCGGG ) UAACCCCG-GAACC [UCCCUCA] UGCUCGGGAU (AGAGGAU P o lya sp is GAGAGAACUGGG ) CAACCCCAUAAAAU [ UUCUUCA ] UGAUUGGGAU ( UGGGGCU O p litis GAGAGAACUGGG) UAACCGCAUGAAAC [UCCUUCA] UGAUUGGGAU (UGAGGCU Asternolaelap UAACGGUCUGGG) UAACCCGAGGAACU [UCCUUCA] UGAUUGGGAU ( AGGGGAU Epicroseius UUGCAAACCGGG ) UAACCCACUGAAAU [ UCCCUCA ] UGAUUGGAAU ( UGGGGCU S eju s UUGCUUGCGGGG) UAACCGGGUCAAAC [UCCUUCU] UGAUUGGGAC (AGGGGUC Heterozerconid UGCGGGACUGGG) CAACCCGAGGAACU [UCUUUCA] UGAUUGGGAU (AGGGGCU
47 ) [ 33' ] ( 48 Asternoseius UGCAACUGUUCCCCU) UGAACGAGGA [AUUCC ] CAGUA ( AGCACUAGUCAUCAG Cercoleipus UGCCCACCUCUCCCA) UGAACGCGGA [AUUCC ] CAGUA (AACGCUGCUCAUCAC Micromegistus UGCAAUUGUUCCACU) UGAACGAGGA [AUGCU] UAGUA ( AACGUUAGUCAUCAA Paramegistus ???????????????) UGAAC?????[?????]?????(??????????????? Antennophorus UGCAAUUGUACCCCU) UGAACGAGGA [ AUGCC ] CAGUA ( AACACUAGUCAUCAG Klinckowstro UGAAAUUGUACCCCU) UGAACGAGGA [AUUCC ] CAGUA (AACACCAGUCAUCAC Neofedrizzia UGAAAUUGUACCCCU) UGAACGAGGA [ACUCC ] CAGUA ( AACGCCAGUCAUCAC Megisthanus UGCAAUUGUACCCCU) UGAACGAGGA (AUUCC ] CAGUA (AACGCCAGUCAUCAA Cryptometast UGCAACUGUUUCCCU) CÂÂACCAGGA [ AUUCC ] UAGUA ( GACGCUGGUCACCAG Passalacarus NGCAAUUGUUCCCCN) UGAACCANNA [AUUCC ] UAGUA (AANGCUGGUCAUCAG Euzercon UGCAAUUGCGCCUCU) UGAACGAGGA [ AUUUC ] UCGUA ( AACGUAAUUCACCAG Neoeuzercon UGCAAUUGCGCCUCU) UGAACGAGGA [AUUUC ] UCGUA (AACAUAAUUCACCAG P o ly a sp is UGCAAUUGUUCCCCU) UGAACGAGGA [AUUCC ] CAGUA (AACGCUAGUCAUCAG O p lit is UGUAAUUUUACCUCA) UGAACGAGGA [AUUCC] UUGUA (AGCACAUGUCAUUAG Astemolaelap UGCAAUUGUCCCCCU) UGAACGAGGA [AUUCC ] CAGUA ( AACACCUGUCAUCAG Epicroseius UGCAAUUGUCCCCCA) UGAACGAGGA [AUUCC ] CAGUA (AACACCUGUCAUÜAA S eju s UGCAAUUGUCCCCCU) UGAACGAGGA [AUUCC ] UAGUA (AGCGCCAGUCAUCAC Heterozerconid UGCAAUUGUACCCCU) UGAACGAGGA [AUUCC ] CAGUA (AACACUAGUCAUCAG
152 ) [ 32' ] (< Astemoseius CUAGUGCU) GAUUACGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA Cercoleipus GCAGUGUU)GAUUGAGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA Micromegistus CUAGCGUU)GACUAUGUCCC[UGCCCUUUGUCCA]CACCGCCCGUCGC(UGUUA Paramegistus Antezmophorus CUAGUGUU)GAUUACGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA K1 inckowstro CUGGUGUU)GAUUACGUCCC[UGCCCUUUGUGCA]CACCGCCCGUCGC(UACUA Neofedrizzia CUGGCGUU) GAUUACGUCCC [UGCCCUUUGUGCA] CACCGCCCGUCGC (UACUA Megisthanus CUGGUGUU)GAUUACGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA Cryptometast CCAGCGUC)GAAUAGGUCCC[UGCCCUUUGUCCA]CACCGCCCGUCGC(UACAU Passalacarus CCANNGNN)NACUAUGUNCC[UG???????????]?????????????(????? Euzercon AUUGUGUU)GAUUAGGUCCC[UGCCCUUUGUCCA]CACCGCCCGUCGC(UGCUA Neoeuzercon AUUGUGUU)GAUUAGGUCCC[UGCCCUUUGUCCA] CACCGCCCGUCGC(UGCUA P o ly a sp is CUAGUGUU) GAUUACGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA O p lit is CGUGUGCU)GAUUAAGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA Asternolaelap CAGGUGCU)GAUUACGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA Epicroseius CAGGUGCU)GAUUACGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA S ej us CUGGCaCU)GAUUAUGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA Heterozerconid CUAGUGCU)GAUUACGUCCC[UGCCCUUUGUACA]CACCGCCCGUCGC(UACUA
49.1 X Astemoseius CCGAUCGA-UGGUUU-AGUAAGGUCAUCGGACUGGUCCUCUA-CU CU-GGU Cercoleipus CCGAUUGGGUG-UUUAAGUGAGGUCAGCUGACUGAUCCUCCG— UAG-A— GGC Micromegistus CCGAUUGAGUGAUUC-AGUGAGGUCAUCCGACGG--CGUCCCCGUAC-CAAACC Paramegri s t us Antennophorus CCGAUUGAGUGAUUU-AGUGAGGUCAUCUGACUGAGCCCCCG AG-C--ACC K1 inckowstro CCGAUUGAGUGAUUU-AGUGAGGUCAUCUGACUGAGCCCCCG GG-C--GGC Neofedrizzia CCGAUUGAGUGAUUU-AGUGAGGUCAUCUGACUGAGCCCCCG------GG-C— GGC Megisthanus CCGAUUGAGUGAUUU-AGUGAGGUCAUCUGACUGAGCCCCCG------AG-C— CGC Cryptometast CCGAUUGAGUGACNU-AAUGAGACCCUCGGACCGGUCUCCCG CG-C--GGC Passalacarus Euzercon CCGAUUGAGUGAACU-AGUGAGGCCAUCCGACCGGGCCCCCG------GG-C— GUC Neoeuzercon CCGAUUGAGUGAACU-AGUGAGGCCAUCCGACCGGGCCCCCG------GG-C— GUC P o ly a sp is CCGAUUGAAUGAUUU-AGUGAGGUCUUCCGAUCGACUACUCA------C - -GUU O p lit is CCGAUUCAAUGAUUU-AGUGAGGGUUUCUGAUUGGUUUUUGA GG GGC Astemolaelap CCGAUUGAAUGGUUU-AGUGAGGUUUUCCGACCGGUACUCUG------AG-C— GGC Epicroseius CCGAUUGAAUGGUUU-AGUGAGGCCGUCUGACUGGUGUUCUG-CGAGUC— AUC S eju s CCGAUUGAAUGGUUU-AGUGAGGCCAUCCGACCGGCUCUCUG-CG— U------GAA Heterozerconid CCGAUUGAAUGGUUU-AGUGAGGUCUUCGUAGCGGUCCUCUG-CUUAU GGC
153 49.2 X Astemoseius c u e ------CG------GAG-UAA—A-AGG—AA~—CGGGAA—GACGA Cercoleipus CUU------CGGGCCAU-CGA— UGG-A-UGG-AG— CGGAAC-GCCAG Micromegistus UUCU------GGAGCG-GAAC GGG-AGAUG CAGGAA-GAAGA Parêunegistus Antennophorus UCC------GGGUC“G“ CG------CGGCG— GG—AG— —CGGAAA—GACGA Klinckowstro CCCC------GGUC--ÜCG------CGG-G-CGG-AG— CGGAAA-GACGA Neofedrizzia CUUC------GGUC— UCG“ — — CGG—G“UGG~AG— CGGAAA—GACGA Megisthanus CUUC------GGGUG-G-CG------CGG-G-AGG-AG— CGGGAA-GACGA CrypCometast AUCC------GUGUC-G-CG------UGG-G-AAGCAC— UGGGAAGGA-GG Passalacarus Euzercon CUUC------GGGGAUCG-CG CGG-G-UGG-AG— CGGGAA-GACGG Neoeuzercon CCUCC------GGGGACCG-CG CGG-G-AGG-AA— CGGGAA-GACGG P o ly a sp is ACCUCC------GGUAAC U CGA-G-A-A-GU— UGAAAA-GAUGC O p liC is CUUC------GGUC------UCG-A— AA-AUUCUGAGAU-UAAGC Asternolaelap c u e ------GUGCC-G-UGA— UGU-U— GG-AC— UGGGAA-GAUGA Epicroseius CUUUC------GGGGUACGAC-ACGCUU-U--GC-GU- -UAGAAU-GACAG Se ju s CUUC------GUUC—G—GG------CUU—U— GA—GC——UGGAAA—GACAG Heterozerconid CUCCUCCUCACGGAGGGGUC-G-UG------CUU-A-CGG-AC— UGUGAA-GAUAG
49.3 >) ( 50 Astemoseius CCGAACUUGACC-UUUGAAAGGAAGUA)AAAGUCGUAACAAGG(UUUCCGUAGG Cercoleipus CCGAACUUGAUUACCUA-GAGGAAGUA)AAAGUCGUAACAAGG(UUUCCGUAGG Mi cromegistus CCGAACUCGAUUGCUUA-GAGGAAACA) AAAGUCGUAACAAGG (UUUCCGUAGG Paramegistus ?????????????????-?????????)????????AACAAGG(UUUCCGUAGG Antennophorus CCAAACUUGAUUACUUA-GAGGAAGUA ) AAAGUCGUAANAAGG ( UUUCCGUAGG K1inckowstro CCGAACUUGACCACUUA-GAGGAAGUA)AAAGUCGUAACAAGG(UUUCCGUAGG Neofedrizzia CCAAACUUGAUUACUUA-GAGGAAGUA)AAAGUCGUAACAAGG(UUUCCGUAGG M egisth m u s CCAAACUUGAUUACUUA-GAGGAAGUA)AAAGUCGUAACAAGG(UUUCCGUAGG Cryptome cast UCGAAUUUGGUCUCUUÀ-GAGGAAGUA) AAAGUCGUAACAAGG (UCUCCGUÀGG Passalacarus ?????????????????-?????????)???????????????(?????????? Euzercon CCGAACUUGUGCACUUA-GAGGAAGCA)AAAGUCGUAACAAGG(UUUCCGUAGG Neoeuzercon CCGAACUUGUGCCCUUA-AAGGAAGCA) AAANUCNUAACAAGG (UUUCCGUAGG P o ly a sp is CCUAACUUGAUCAUUUA-GAGGAAGUA)AAAGUCGUAACAAGG(UUUCCGUAGG O p lit is CCAAACUUGAUCAUUAA-GAGGAAGUU) AAAGUCGUAACAAGG (UUUCCGUAGG Asternolaelap CCGAACUUGAUCACUUA-GAGGAAGUA) AAAGUCGUAACAAGG (UUUCCGUAGG Epicroseius CCAAACUUGAUCAUUUA-GAGGAAGUA) AAAGUCGUAACAAGG (UUUCCGUAGG Se ju s CCAAACUUGAUCAUUUA-GAGGAAGUA) AAAGUCGUAACAAGG (UUUCCGUAGG Heterozerconid CCGAACUUGAUCAUUUA-GAGGAAGUA) AAAGUCGUAACAAGG ( UUUCCGUAGG
154 50 Asternoseius UGAACCUGCGGA Cercoleipus UGAACCUGCGGA Mi cromegistus UGAACCUGCGGA Paramegistus UGAACCUGCGGA Antennophorus UGAACCUGCGGA Klinckowstro UGAACCUGCGGA Neofedrizzia UGAACCUGCGGA Megisthanus UGAACCUGCGGA Cryptometast UGAACCUGCGGA Passalacarus Euzercon UGAACCUGCGGA Neoeuzercon UGAACCUGCGGA P o lyaspis UGAACCUGCGGA O p litis UGAACCUGCGGA Asternolaelap UGAACCUGCGGA Epicroseius UGAACCUGCGGA Se ju s UGAACCUGCGGA Heterozerconid UGAACCUGCGGA
155 APPENDIX B. Aligned V-X region of the 28S rRNA sequences.
7 0 ) ( 71 ) [67'] Astemoseius GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA Cercoleipus GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU]UAA Mi cromegistus GUAGCCAAAUGCCU) (CGUCAUAUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA Promegistus GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA Paramegistus GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG)CGCAUGAAU[GGAU]UAA Antennophorus GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU]UAA Klinkowstro GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGA?] UAA F ed rizzia GUAGCCAAAUGCCU) {CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA Neofedrizzia GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG)CGCAUGAAU[GGAU]UAA Megisthanus GUAGCCAAAUGCCU) (CGUCAU ?UAAUUAGUGACG) CGCAUGAAU[GGAU]UAA Trichodiplog GUAGCCAAAUGCCU) (CGUCAUUUAAUUAGUGACG)CGCAUGAAU[GGAU]UAA Passalacarus GUAGCCAAAUGCCU) ( CGUCAUUUAAUUAGUGACG ) CGCAUGAAU [GGAU ] UAA Euzercon GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA Neoeuzercon GUAGCCAAAUGCCU) (CGUCAUCU?AUUAGUUACG) GGCAU?AAU[GGAU] UAA P o lya sp is GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG ) CGCAUGAAU [GGAU] UAA O p litis GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA As tem ol aelap GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA Epicroseius GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA Se ju s GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA Discozercon GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA Heterozerconid GUAGCCAAAUGCCU) (CGUCAUCUAAUUAGUGACG) CGCAUGAAU[GGAU] UAA
156 [ 6 4 ' ] [ 6 1 ' ] ( Astemoseius [CGAGAUUCCC]AC [UGUCCCUACCUACUAUCUA]GCGAAACCACUG(CCAAGGG Cercoleipus [CGAGAAUCCCIAC [UGUCCCUACUUACUAUCUA]GCGAACCCACAG(CCAAGGG Mi cromegistus [CAAGAUUCCC]GC [UGUCCCAACCUACUAUCUA]GCGAAACCACAG(CCAAGGG Promegistus [CGAGAUUCCC]GC [UGUCCCUACUUGCUAUCCA]GCGAAACCACAG(CCAAGGG Paramegistus tCGAGAUUCCC]AC [UGUCCCUACCUACUAUCUA]GCGAAACCACAG(CCAAGGG Antennophorus [CGAAAUUCCU]AC [UGUCCCUACCUACUAUCUA]GCGAAACCACUG(CCAAGGG Klinkowstro [C?????UCCC]AC [UGUCCC? ??????? ?UCUA]GCGAAAUCACAG(CCAGGGG F e d rizzia [CGAGAUUCCCJAC [UGUCCCUACCUACUAUCUA]GCGAAACCACAG(CCAAGGG Neofedrizzia [CGAGAUUCCC]AC [UGUCCCUACCUACUAUCUA]GCGAAACCACAG(CCAAGGG Megisthanus [CGAGAUUCCC]AC [UGUCCCUACCUACUAUCCA]GCGAAACCACAG(CCAAGGG Trichodiplog [CGAGAUUCCC]AC [UGUCCCUAUCCGCUAUCUA]GCGAACCAACAG(CCAAGGG Passalacarus [CGAGAUUCCCIGC [UGUCCCUAUCCGCUAUCUA]GCGAAACCACUG(CAAAGGG Euzercon [CGAGAUUCCC]AC [UGUCCCUACCUACUAUCUA]GCGAAACUACAG(CCAAGGG Neoeuzercon [CGAGAUUCCC]AC [UGUCCCUACCUACUAUCUA]GCGAAACCACAG(CCAAGGG P o lya sp is [CGAAAUUCCC]AC [UGUCCCUAUCUACUAUCUA]GCGAAAUCACAG(CCAAGGG O p litis [CGAGAUUCCC]AC [UGUCCCUAUCUACUAUCUA]GCGAAACCACAG(CCAAGGG Asternolaelap [CGAGAUUCCC]AC [UGUCCCUAUCUACUAUCUA]GCGAAACCACAG(CCAAGGG Epicroseius [CGAGAUUCCC]UC [UGUCCCUAUCUACUAUCUA]GCGAAACCACAG(CCAAGGG Se ju s [CGAGA7UCUC]UC [UGUCCCUAUCUACUAUCUA]GCGAA7CCACAG(CCAAGGG Discozercon [CGAGAUUCCC]AC [UGUCCCUACUUACUAUCUA] GCGAACCCACAG (CCAA(3GG Heterozerconid [CGAGAUUCCC]AC [UGUCCCUACCUACUAUCUA]GCGAAACCACAG(CCAAGGG
72 ) 73 ] [ 74 ] Astemosei us AACGGGCUUGG)AAGAC- GU- UUGCGGGGAAA] GAAGA[CCCUGUUGAGCUU] G Cercoleipus AACGGGCUUGG)GAGG— UU-UAGCGGGGAAA] GAAGA[CCCCGUUGAGCUU] G Mi cromegistus AACGGGCUUGG)AACAA- UGUCGGCGGGGAAA]GAAGA[CCCCGUUGAGCUU]G Promegistus AACGGGCUUGG) AACAU- UGUCAGCGGGGAAA]GAAGA[CCCUGUUGAGCUU]G Parêunegistus AACGGGCUUGG)A— AU- UAUCGGCGGGGAAA] GAAGA[CCCUGUUGAGCUU] G Antennophorus AACGGGCUUGG) AAGC- - UAUCUGCGGGGAAA]GAAGA[CCCUUUUGAGCUU]G Klinkowstro AAC 7GGCCUGG) AAACGC AU? 7GGCGGGGAAA]GAA?A[CCCUGUUGAGCUU]G F e d rizzia AACGGGCUUGG) AAACAA CU GCGGGGAAA]GAAGA[CCCUGUUGAGCUU] G Neofedrizzia AACGGGCUUGG) AAACAC AU-CGGCGGGGAAA]GAAGA [CCCUGUUGAGCUU]G Megisthcuius AACGGGCUUGG) AAUC-- UA-CUGCGGGGAAA] GAAGA [CCCUGCUGAGCUU]G Trichodiplog AACGGGCUUGG) AACGAG [ AC-CU-CGGGGAAAJGAAGA [CCCUACUGAGCUU] G Passalacarus AACGGGCUUUG) AAAAUG [AC-CU-CGGGGAAA]GAAGA[CCCUACUGAGCUU]G Euzercon AACGGGCUUGG) GCCUUG [AC-CAGCGGGGAAA]GAAGA[CCCUGUUGAGCUU]G Neoeuzercon AACGGGCUUGG) ACCUGG [AC-CAGCGGGGAAA]GAAGA[CCCUGUUGAGCUU]G P o lya sp is AACGGGCUUGG) AAAAU- [AA-CGGCAAGGAAA]GAAGA[CCCUAUUGAGCUU]U O p litis AACGGGCUUGa) GAUG— [CA-CUGCGGGGAAA]GAAGA[CCCUAUUGAGCUU]G Asternolaelap AACGGGCUUGG) AACA-- [AA-CAGCGGGGAAA]GAAGA[CCCUGUUGAGCUU]G Epicroseius AACGGUCUUGG)UAACG- [AA-CAGCGGGGAAA] GAAGA[CCCUGUUAAGCUU] G Se ju s AACGGGCUUGG) UGAG— [A7-CGGCGGGGAAA]GAA?A[CCCUGUUGAGCUU]G Discozercon AACGGGCUUGG) AACG— [AA-UAGCGGGGAAA]GAAGA[CCCUGUUGAGCUU]G Heterozerconid AACGGGCUUGG) AACG— [AA-CCGCGGGGAAA]GAAGA[CCCUGUUGAGCUU]G
157 [ 75 ] [ 76 ] t 77 ] Astemoseius [ACUCUAGUCUGAA]UUU[GUGAAGAGACAUGAG-AG]GUGUAGCAUA[GGÜGG] Cercoleipus [ACUCUAAUCUGAC]AUU[GUGAGGAGACAUGAG-AG]GUGCAGUAUA[GGUGG] Micromegistus [ACUCUAGUUUGAC]UCU[GUGAGGAGACAUGAG-AG]GUGUAGAAUÀ[GGUGG] Promegistus [ACUGUAGUCUGAC]UCU[GUGAAGAGACAUGAG-AG]GUGUAGCAUA[GGUGG] Paramegistus [ACUCUAGUCUGAA]UCU[GUGAAGAGACAUGAG-AG]GUGUAGCAUA[GGUGG] A n tennophorus [ACUCUAGCUUGAC]UCU[GUGAAGAGACAUGAG-AG]GUGUAGCAUA[GGUGG] Klinkowstro [ACUGUAGUCUGAC]UCU[GUGAAGAGACAUGAG-AG]GUGUAGCAUA[GGUGG] F e d r iz z ia [ACUGUAGUCUGAC] UCU[GUGAAGAGACAUGAG-AG]GUGUAGCAUA[GGUGG] Neofedrizzia [ACUGUAGUCUGAC] UCU[GUGAAGAGACAUGAG-AG]GUGUAGCAUA[GGUGG] Megisthanus [ACUGUAGUCUGAC] UCU[GU?AAGAGACAUGA?-AG] GUGUAGCAUA[GGUGG] Trichodiplog [ACUCUAGUUUGAC] UCU[GUGAAGAGACACGAG-AG] GUGUAGAAUA[GGAGG] Passalacarus [ACUCUAGUUUGAC]UCU[GUGAAGAGACAUGAG-AG]GUGUAGAAUA[GGAGG] Euzercon [ACUUUAGUCUGCC]GUU[GUUAAGAGACAUGAG-AG]GUGUAGGAUA[GGCGG] Neoeuzercon [ACUUUAGUCUGCC]GUU[GUUAAGAGACAUGAG-AG]GUGUAGUAUA[GGCGG] P o ly a sp is [ACUUUAGUCUGAC]UCU[GUGAAGAGACAUGAG-AG]GUGUAGCAUA[AGUGG] O p litis [ACUCUAGUCUGAA]UCU[GUGUAGAGACAUAAG-AG]GUGUAGCAUA[AGGGG] Asternolaelap [ACUCUAGUCUGAC] UCU[GUGAAGAGACAUAAG-AG] GUGUAGCAUA[AGUGG] Epicroseius [ACUCUAGUCUAAC] UCU[GUAAGGAGACAUAAG-AG] GUGUAGCAUA[AGUGG] S e j us [ACUCUAGUCUGAC] GCU[GcGCAGAGACAUAAGCGG]GCGUAGAAUA[CGUGG] Discozercon [ACUCUAGUCUGAC]UCU[GUGAAGAGACAUAAG-AG]GUGUAGCAUA[AGUGG] Heterozerconid [A?U?UAGUC?GAC] UCU[GUGAAGAGACAUGAG-AG]GUGUAGCAUA[GGUGG]
( 78(D9) ) Astemoseius GA {GACGGCCU CGGCCGUC ) GUCCAUGAAAU Cercoleipus GA ( GUCGUGUGUCCUUU CGGGGAUGCGCGUC ) GCCAAUGAGAU Micromegistus GA {GGCUU AUGCC ) GUCCAUGAAAU Promegistus GA ( GGGAGGGCUGGCCU UUGGUCACCCUUUC ) GACAAUGAAAU Paramegistus GA ( GACGGUUCU CCGUC ) GCAAAUGAAAU Antennophorus GA ( GACGGUACU GCUGUC ) GCCAAUGAAAU Klinkowstro GA ( GACGGGGC AACCCGUC ) GCCAAUGAAAU F e d r iz z ia GA ( GAUGGGGC AACCCGUC ) AACAAUGAAAU Neofedrizzia GA ( GACGGGGA AACCCGUC ) GCCAAUGAAAU Megisthêinus GA ( GACGCCGC AAGGUGUC ) GCCAAUGAAAU Trichodiplog GA ( GGGUCGCGCGCCCAGC GAUGGGACGCGGCUC ) GCCAGUGAAAU Passalacarus GA (GGUGGGUCGCCCCGUU UCGGCGGGGUCCCGCC ) GCCAGUGAAAU Euzercon GA(GGCGUCCGAUUGGGGCCG AGAGGUUCCGCGGGCGCC) GCCGGUGAAAU Neoeuzercon GA ( GCGGUCCGGGCA ACCGGGUGCCGC ) GCCAGUGAAAU P o ly a sp is GA ( GUUAGCUU GCUGAC ) AAAAAUGAAAU O p litis GA {GACAGCUU CGGUUGUC ) GCCAAUGAAAU Asternolaelap GA ( GCCGGGCAGCCUU CGGGCAGCCGGC ) GUCAAUGAAAU Epicroseius GA {GCUCAAAGGCU UGCCGAUGGGC ) GUCAAUGAAAU S eju s GA (GACAAGGCGACCCUC------GCGGUA?CCUUGCC) GUCAAUGAAAU Discozercon GA {GCUU CGGC ) GCCAAUGAAAU Heterozerconid GA {GCAGGACCAAACGCCGGGACCCUCGGGCUCGGCGCCUGC ) GCCAAUGAAAU
158 [ 7 7 ' [ 7 6 ' ] ( As tem osei us A[CCACC A [CUCUCACUGUUUCUUUAC ] UUA ( CUCGGUGAAACGGAAAGAUGG Cercoleipus A[CCACC A[CUCUCACUGUUUCCUCAC]UUA(UUCGAUGACACGAAAG-ACGG Micromegistus A[CCACC A[CUCUCACUGUUUCCUUAC]UUA(CUCGGUGAAACGAAGG-ACGG Promegistus A[CCACC A [CUCUCACUGUUUCUUUAC] UUA(CCCGGUUAA-CGGAAAAAUGA Paramegistus A[CCACC A[CUCUCACUGUUUCUUUAC]UUA(CUCGGUUAAACGAAGA-AUGG Antennophorus A[CCACC A [CUCUCACUGUUUCUUUAC] UUA(CUCGGUAAAGCGAAGA-AUGG Klinkowstro A[CCACC A [CUCUCACUGUUUCUUUAC]UUA(CUCGGUGAUGaGAGGA-AUGG F e d rizzia A[CCACC A[CUCUCACUGCUUCUUUAC]UUA{CUCGGUCAAAAGAGGA-AUGG Neofedrizzia A[CCACC A[CUCUCACUGCUUCUUUAC] UUA(CUCGGUGAAAAGAGAA-AUGG Megistheuius A[CCACC A[CUCUCACUGUUUCUUUAC] UUA(CUCGGUUGAGCGAAGA-ACGG Trichodiplog A[CCUCC A[CUCUCGCUGUUUCUUUAC]UUA(CUCGGUUAAGCGAAGG-ACGG Passalacarus A[CCUCC A [CUCUCACUGUUUCUUUAC] UUA(CUCGGUUGAGCGAAGG-ACGG Euzercon A[CCGCC A[CUCUCACUGUUUCUUUAC]UUA(CACGGUAGAGCGGAGA-ACGG Neoeuzercon A[CCGCC A [CUCUCACUGUUUCUUUAC] UUA(CACGGUAAAGCGGAGA-ACGG P o lya sp is A[CCACU A[CUCUCAUUGUUUCUUUAC]UUA(CUCGGUAAAACGAAGA-GCGG O p litis A[CCCCU A[CUCUUAUCGUUUCUGUAC]UUA(CUCGGUUAAACGAAGAGUUGG Asternolaelap A[CCACU A [CUCUUAUUGUUUCUUUAC] UUA(CUCGGUAAUACGAAGA-GCGG Epicroseius A[CCACU A [CUCUUAUCGUUUCUUUAC] UUA(CUCGGUCAAACGAAGA-GUGG S ej us A[CCACG A [CCCUUAUCGUUUCUGCAC] UUA(CUCGGC- ACACGAGAA-ACGG Discozercon A[CCACU A [CUCUUAUCGUUUCUUUAC] UUA(CUCGGUUACACGAAGA-GUGa Heterozerconid A[CCACU A[CUCUCAUCGUUUCUaCAC]UUA(CUCGGUGGAGAGAGGG-GGGG
DIO Astemoseius CCUGC------CUGGCCUGUCUUC-UAGCGUCGAACCCUGAAGCGCCU------Cercoleipus CGACC--UGUUUGCCUGUUUUU-UGGUGAAG- - AGCGUGGUGCCCGCGUUGUGU Micromegistus GUUU------CACCCGA- -CUUU-UAGCGUU-AACUACAGCGCCUU------Promegistus UCUUUUUCAAAGUUCUGCUUUC -UAGC - UU-AAACCUUAGUUGAG------Parêunegistus GCCUU------GGUCCAGACUUC - UAGCGUU-AACCUCCGCGCCCC------Antennophorus ACUUC------GGUUCAGACUUC -UAGCGUU - AACCUCCGCGCCCC------Klinkowstro GCUC UUGGCCCUCAUUUU-UGGCUCU-AACCUCCGCGCCAC------F e d rizzia GCCC AUGGCCCUGACUUU-UGGCGUU-AACCUACGAGCCAU------Neofedrizzia GCCC AUGGCCCUGACUUU-UGGCGUU-AACCUCCGAGCCUU------Megisthanus GCUC------CGGUCCGGCCUUC - UGGCGUU-AACCCGGGCGCCUC------Trichodiplog GCUC------CGGCCCUUACUUUGUGGCGUU-AACCCGGGGGCCCUC------Passalacarus GCCUC---- ACGGCCCUUACUUUGUGGCGUU-AAUCGGGGCGCGGUUC------Euzercon GCCUC ACGGCUCGGUCUUUGUGGCGUU-AACGCGGUCGCCCC------Neoeuzercon GCCUC ACGGCCCGGACUUUGUGGCGUU-AACGCGGCCGCCCG------P o lya sp is ACGU------UUGUCCAUU7UUC-UAGCGUU-AA-CCGCAGUACAU------O p litis CCUU------GUGUCG AC - ACUUC - UAGCGUUUAAACCUUGCUCAG------Asternolaelap ACUU------CGGUCCACGCUUC-UAGAAUU-AA-CGGCGUAUCGCCC------Epicroseius AUUU------CGGUUCAAUCUUC-UGGCGUU-AAGCUUGGGACCCU------S eju s ACCU------CGGUCCGAUCUUG-UGGCGUU-AAGCUGGGACCCCCUA------Discozercon CCUC------CGGGCCACGCUUC-UGGUGUU-AAUCGCCUCUACUU------Heterozerconid CCUU------CGGGUCCCACUUC -UGGUGCC -AAGCCGCGUCGCGGGC------
159 DIO Astemoseius ------CCGCGCGCCGAGGGGAUCCGCU Cereal eipus GCCAGCGCCCCUUCCGGGGUGCGAAGCGCGCGGCGUGUGGAUGCGCGAUUCGCÜ Micromegistus ------CGUUGCGCAGUGGGAUUCGCG Promegistus CUCUUCGGGGUUCCUUUGGGAUUCGCG Paramegistus ------CGUGCGCAGAGGGAUUCGCA Antennophorus ------CGUGCGGGGAGGGAÜUCGCU Klinkowstro CGUGUGCGGAGGGACUCCAC F e d r iz z ia CGÜGUUCGGAGGAACUCCUG Neofedrizzia ------CGUGUUCGGAGGAACUCCUC Megisthanus CGCGCGCCUGGGGAUUCGCU Trichodiplog ------ACCGGUCACCGGGGAUUCGCG Passalacarus ------CGGCCGCGCCCCGAGAUUCGCG Euzercon UGCGCGACCGCGGAUUCGCU Neoeuzercon UGAGCGGCUGCGGAUUCGCU P o ly a spis GUGUACAGUGG-GAUUCGCU O p lit is CAUAGUGAGGUGAÜUCGCU Asternal aelap ------GACGGCGACGGCCUGAUUCGCÜ Epicroseius UGCGGCCCCGGGUGAUUCGCU S eju s ACCGGGGACCCGGCGAUUCGCG Discozercon GCUAGUGGCGUGAUUCGCU Heterozerconid ------AGUCGUGACGCGGCGACUCUGU
) [ 75' ] ( 80 ) ( 81 Astemoseius CCGAG ) GACAGU [ UUCAGAUGGGCAGU ] UC ( GACUGGGGCGGUAC ) ( AUCUGUU Cercoleipus UCGAA)GACAGU[GUCAGGCGGGGAGU] UC(GACUGGGGCGGUAC) ( AUCUGUC Mi cromegistus CCGAG) GACAGU [GUCAGACGGGGAGU] UC (AGCUGGGGCGGUAU) (AUCUGUC Promegistus CCGGG) GACAGU(GUUAGGUGGGCAGU] UU(GGCUGGGGCGGUAC) ( AUCUGUC Paramegistus CCGAG ) GACAGU [ UUCAGGUGGGGAGU ] UU ( GACUGGGGCGGUAC ) ( AUCUAUC Antennophorus CCGAG) GACAGU[GCCAGGUAGGCAGU] UU(GGCUGGGGCGGUAC) ( AUCUGUC Klinkowstro CCGAG)GACACU[GUCAGGUGGGGAGU] UU(GACUGGGGCGGUAC) ( AUCUGUC F e d r iz z ia CCGAG ) GACACU [ GUCAGGUGGAGAGU ] UU ( GACUGGGGCGGUAC ) ( AUCUGUC Neofedrizzia CCGAG ) GACACU [ GUCAGGUGGAGAGU ] UU ( GACUGGGGCGGUAC ) ( AUCUGUC Megisthanus CCGAG ) GACAGU [ UUCAGGUGGGGAGU ] UU ( GA ? UGGGGCGGUAC ) ( AUCUGUC Trichodiplog CCGAG ) GACAGU [ UUCAGACAGAUAGU1 UU t GACUGGGGCGGUAC ) ( AUCUGUC Passalacarus CCGAG ) GACAGU [ GUCAGAUUGACAGU ] UU ( GACUGGGGCGGUAC ) ( AUCUGUC Euzercon CCGUG)GACAGC [GGCAGGUCGGCAGU] UC (GACUGGGGCGGUAC) (AACUGUC Neoeuzercon CCGUG)GACAGC[GGGAGGUCGGGAGU]UC(GACUGGGGCGGU9C) (AACUGUC P o ly a sp is CCGAG ) GACAGU [ GUCAGGUGGGAAGU ] UU ( GACUGGGGCGGUAC ) ( AUCUGUC O p litis CCGAG ) GACAGU [ UUCAGUUGGGGAGU ] UU ( GACUGGGGCGGUAC ) ( AUCUGUC Asternolaelap CCGAG ) GACAGU [ GUCAGGUGGGGAGU ] UU ( GACUGGGGCGGUAC ) ( AUCUGUC Epicroseius CCGAG ) GACAGU [ GUUAGGUGGGGAGU ] UC ( GACUGGGGCGGUAC ) ( AUCUGUC S eju s CCGAG ) GACÀCC [ GUCAGGUGGGGAGU ] UU ( GACUGGGGCGGUAC ) ( AUCUGUC Discozercon CCGAG ) GACAGU [ GUCAGGUGGGGAGU ] UU ( GACUGGGGCGGUAC ) ( AUCUGUC Heterozerconid CCGAG ) GACAGU [ GUCAGGUGGGGAGU ] UU ( GUCUGGGGCGGAAC ) ( AUCUGUC
160 8 1 ) [82] [ 83 ( 84 Astemoseius AAAUGGUAACGCAGGU) GU[CC] CAA[GGCUGGC U {CAGCGAGGACAGAAACC Cercoleipus AAAUGGUAACGCAGGU) GU[CC] AAA[GGCCAGC U ( CAGCGAGGACAGAAACC Micromegistus AAAUGGUAACGCAGGU)GU[CC]UAA[GGCCAGC U ( CAGCGAGGACAGAAACC Promegistus AAAGCGUAACGCAGGU) GU[CC] CAA[GACCGGC U (CAGCGAGGACAGAAACC Paramegistus AAAAGGUAACGUAGGU) GU[CC] UAA[GGCCAGC U (CAGCGAGGACAGAAACC Antennophorus AAAUGGUAACGCAGGU) GU[CC] UAA[GGUCGGC U ( CAGCGAGGACAGAAACC Klinkowstro AAAUGGUAACGCAGGU) GU[CC] UAA[GGCCGGC U ( CAGCGAGGACAGAAACC F e d rizzia AAAUGGUAACGCAGGU) GU[CC] UAA[GGCCGGC U (CAGCGAGGACAGAAACC Neofedrizzia AAAUGGUAACGCAGGU) GU[CC] UAA[GGCCGGC U (CAGCGAGGACAGAAACC Megisthanus AAAUGGUAACGCA?GU) GU[CC] UAA[GGCCGGC U ( CAGCGAGGA? AGAAACC Trichodiplog AAAUGGUAACGCAGGU) GU[CC] UAA[GGCCGGC U ( CAGCGAGGACAGAAACC Passalacarus AAAAGGUAACGCAGGU)GU[CC]UAA[GGCCGGC U (CAGCGAGGACAGAAACC Euzercon AAACGGUAACACAGUU) GU[CC] UAU[GGUCGGC U (CAGCGAGGACAGAAACC Neoeuzercon AAAAGGUAACACAGUU)GU[CC]UAA[GGUCGGC U ( CAGCGGGGACAGAAACC P o lya sp is AAAUGGUAACGCAGGU) GU[CC] UAA[GGCGAGC U ( CAGCG? 7GACAGAAACC O p litis AAAUGGUAACGCAGGU) GU[CC] UAA[GGCAAGC U (CAGCGAGGACAGAAACC Asternolaelap AAAUGGUAACGCAGGU)GU[CC]UAA[GGCGAGC ] U(CAGCGAGGACAGAAACC Epicroseius AAAUGGUAACGCAGGU) GU[CC] UAA[GGCAAGC ] U(CAGCGAGGACAGAAACC S eju s AAAUGGUAACGCAG ? U) GU[CC] UAA[GGUGAGC ] U (CAGCGAGGACAGAAACC Discozercon AAAUGGUAACGCAGGU)GU[CC] UAA[GGCGAGC ] U(CAGCGAGGACAGAAACC Heterozerconid AAAUGGUAACGCAGGU) GU[CC] UAA[GGCUAGC ] U(CAGCGAGGACAGAAACC
84 ) [ 83' ] ( 86 Astem oseius UCGCGUAG ) AGCAAAGGGGCAAAU [ GCCAGCU ] UGA ( U-CUUGAUUUUCAGUAC Cercoleipus UCGGGUAG) AGCAAAAGGGCAAAC [GCUGGCU] UGA (U-CUUGGCUCUCAGUAC Micromegistus UCGCGUAG) AGCAAAAGGGCAAAU [GCUGGCU] UGA (U-CCUGAUUUUCAGUAC Promegistus UCGUGUAG) AGCAAAAGGGCAAAU [GCCGGUU] UGA (U-CUUGAUCUUCAGUAG Paramegistus UCGCGUAG) AGCAAAAGGGCAAAU [GCUGCCU] UGA (U-CUUGAUUUUCAGUAC Antennophorus UCGCGUAG) AGCAAAAGGGCAAAU [GCCGGCU] UGA (U-CUUGAUUUUCAGUAC Klinkowstro UCGCGUAG)AGCAAAAGGGCAAAU[GCCGGCU]UGA(U-CUUGAUUUUCAGUAC F e d rizzia UCGCGUAG ) AGCAAAAGGGCAAAU [GCCGGCU] UGA (U-CUUGAUUUUCAGUAC Neofedrizzia UCGCGUAG ) AGCAAAAGGGCAAAU [GCCGGCU] UGA (U-CUUGAUUUUCAGUAC Megisthanus UCGCGUGG ) AGCAAAAGGGCAAAU [ GCCGGCU ] UGA ( U-CUUGAUUUUCAGUAC Trichodiplog UCGCGUGG ) AGCAAAAGGGCAAAU [GCCGACU] UGA ( U-UUUGAUUCUCA6UAC Passalacarus UCGCGUGG) AGCAAAAGGGCAAAU [GCCGCCU] UGA (U-UUUGAUUCUCAGUAU Euzercon UCGCGUGG ) AGCAAAAGGGCUAAU [GCCGGCU] UGA (U-CUUGAUUUUCAGUAC Neoeuzercon UCGCGUAG ) AGCAAAAGGGCUAAU [GUCGGCU ] UGA ( U-CUUGAUUUUCAGU7C P o lya sp is UCGCG? AG ) AGCAAAAGGGCAAAU [ GC ? ?GCU ] UGA ( UACUUGAUUUUCAGUAC O p litis UCGCGUAG ) AGCAUAAGGGCAAAA [GCUUGCU] UGA ( U-CUUGAUUUUCAGUAC Asternolaelap UCGCGUAG ) AGCAAAAGGGCAAAA [GCUUGCU] UGA (U-CUUGAUUUUCAGUAC Epicroseius UCGCGUAG ) AGCAAAAGGGCAAAU [GCUUGCU] UGA (U-CUUGAUUUUCAGUAC S eju s U7GCGUAG) AGCAGAAGGGCAAAU [GCUUGCU] UGA (U-?UUGAUUUU??GUAC Discozercon UCGCGUAG) AGCAAAAGGGCAAAA [GCUUGCU] UGA (U-CUUGAUUUUCAGUAC Heterozerconid UCGCGUAG) AGCAAAAGGGCAAAA [GCUAGCU] UGA (U-CUUGAUUUUCAGUAC
161 86 ) ( 87 ) 8 2 ' ( 88 Astemoseius GAAUACAG-A) C (CGCGAAAGCG) G GG] CCCCUCGAUC (CUUUGGCU-UUUAC Cercoleipus GAGCACAG-A) C (CGAGAAAUCG) U GG] CCCCUCGAUC (CUUUUGCU-CUUAC Micromegistus GAAUACGG-A) C (CGCGAAAGCG) G GG]CCUAUCGAUC (CUUUUGUU-UUUAC Promegistus GGAUAAAG- A ) C (UGCGAAAGCA) G GG] CCUAUCGAUC (CUCUUUUU-ACGCÜ Parêunegistus GAAUACAG-A) C (CGCGAAAGCG) G GG] CCCCUCGAUC (CUUUUGCU-UUUAC Antennophorus GAAUACAG-A) C (CGCGAAAGCG) G GG] CCCCUCGAUC (CUUUUGCU-UUUAC Klinkowstro GAAUACAG-A) C (CCCGAAAGGG) G GG] CCCCUCGAUC (CUUUUGCU-UUUAC F e d rizzia GAAUACAG-A) C (CGCGAAAGCG) G GG] CCCCUCGAUC (CUUUUGCU-UUUAC Neofedrizzia GAAUACAG-A) C (CGCGAAAGCG) G GG]CCCCUCGAUC (CUUUUGCU-UUUAC Megisthcuius GAAUACAGCA) C (CGCGAAAGCG) G GG]CCCCUCGAUC (CUUUUGUUCUUUAC Trichodiplog GAAUACAA-A) C (AGCGAAAGCU) G [GG] CCUCUCGAUC (CUUUUGGA-UUUAA Passalacarus GAAUACGA-A) C (CGCGAAAGCG) G [GG]CCUAUCGAUC (CUUUUGCC-UUUAC Euzercon GAAUACAG-A) C {CGCGAAAGCG) G [GG]CCCCUCGAUC (CUUUUGCU-UUUAC Neoeuzercon GAAUACAG-A) C (CGCGAAAGCG) G [GG]CCCCUCGAUC (CUUUUGCU-UUUAC P o lya sp is GAAUAAAG-A) C (CGCGAAAGCG) G [GG]CCCCU?GAUC (CUUUUGUU-?UUAC O p litis GAAUACAG-A) C (UGCGAAAGCA) G [GG] CCUUACGAUC (CUUUUGUU-UUUAC Asternolaelap GAAUACAG-A)C(CGCGAAAGCG)G [GG] CCCCUCGAUC (CUUUUGUU-UUUAC Epicroseius GAAUACAG-A) C (CGCGAAAGCG) G [GG] CCCCUCGAUC (CUUUUGCU-UUUAC S eju s GAAUACAG-A) C (CGCGAAAGCG) G [GG] CCCCUCGAUC (CUUUUGUU-UUUAC Discozercon GAAUACAG-A) C (CGCGAAAGCG) G [GG] CCCCUCGAUC (CUUUUGUU-UUUAC Heterozerconid GAAUACAG-A) C (CGUGAAAGCG) G [GG] CCCCUCGAUC (CUUUUGUU-UUUAC
88 ) [ 74' ] Astemoseius GAGUUUUAAGCUAGAG) GUGUCAGAA [AAGUUACCACAGGG] -AUAACUGGCUU Cercoleipus GAGCUUUAAGCAAGAG) GUGUCAGAC[AAGUUACCACAGGG]-AUAACUGGCUU Mi cromegistus GAGUUUUAAACAAGAG) GUACCAGAA [AAGUUACCACAGGG] -GUAACUGGCUU Promegistus GAGUUUACGCUAAGAG)GUGUCAGAA[UAGUUACUACAGGG]-GUAACUGGCUU Paramegistus GAGUUUUAAGCAAGAG) GUGUCAGAA [AAGUUACCACAGGG] -AUAACUGGCUU Antennophorus GAGUUUUAAGCAAGAG ) GUGUCAGAA [AAGUUACCACAGGG ] -AUAACUGGCUG Klinkowstro GAGUUUUAAGCAAGAG) GUGUC7GAA [UAGUUACCACAGGG] -AUAACUGGCUU F e d rizzia GAGUUUUAAGCAAGAG ) GUGUCAGAA [AAGUUACCACAGGG] -AUAACUGGCUU Neofedrizzia GAGUUUUAAGCAAGAG ) GUGUCAGAA [ GAGUUACCACAGGG ] -AUAACUGGCUU Megisthanus 7AGUUUUAAGCAAGAG) GUGUCA7AU[AAGUUACCACAGGG]UAUAACUGGCUU Trichodiplog GAGUUUUAAACAAGAG) GUGUCAGAA [ AAGUUACCAUAGGG} -AUAACUGGCUU Passalacarus GAGUUUUAAGCAAGAG) GUGUCAGAA [AAGUUACCAUAGGG] -AUAACUGGCUU Euzercon GAGUUUUAAGCAAGAG) GUGUCAGAA [UAGUUACCACAGGG] -AUAAUUGGCUU Neoeuzercon GAGUUUUAAGCAAGAG ) GUGUCAGAA [AAGUUACCACAGGG] -AUAACUG7CUU P o lya sp is G?GUUU? AAGCAAGAG ) GUGUCAGAA [AAGUUACCAUAGGG] -AUAACUGACUU O p litis GAGUUUUAAGCAAGAG) GUGUCAGAA [AAGUUACCAUAGGG] - AUAACUGGCUU Astemolaelap GAGUUUUAAGCAAGAG) GUGUCAGAA [AAGUUACCACAGGG] - AUAACUGGCUU Epicroseius GAGUUUUAAGCAAGAG) GUGUCAGAA [ AAGUUACUACAGGG] -AUAACUGGCUC S eju s GAGUUUUAAGCAAGAG) GUGUCAGAA [AAGUUACCACAGGG] -AUAACUGGCUU Discozercon GAGUUUUAAGCAAGAG) GUGUCAGAA [GAGUUACCACAGGG] - AUAACUGGCUU Heterozerconid GAGUUUUAAGCAAGAG) GUGUCAGAA [AAGUUACCACAGGG] -AUAACUGGCUU
162 ( 89 ) Asternoseius GU(GGCG-GCCAAGAGUCCAUAUCGACGUCGCU)UUUUGAUCCUUCGAUG Cercoleipus GU(GGCA-GCAGAGC ?UCCAUAGCGGCGUUGCU) UUUUGAUCCUUCGAUG Mi cromegistus GU(GGCG-GCCCAGAGUUCCCAUCGACUUCGCU)UUUUGAUUCUUCGAUG Promegistus GU(GGCG- GCCAAGAGUCCACAUCGACGUCGCU) UUUUGAUCCUUCGAUG Paramegistus GU(GGCG-GCCAAGCGUUCAUAGCGACGUCGCU)UUUUGAUCCUUCGAUG Antennophorus GU(GGCG-GCCAAGCGUCCAUAGCGACGUCGCU) UUUUGAUCCUUCGAUG Klinkowstro GU(GGCG-GCCAAGCGUUCAUAGCGACGUCGCU)UUUUGAUCCUUCGAUG F e d r iz z ia GU(GGCG-GCCAAGCGUUCAUAGCGACGUCGCU)UUUUGAUCCUUCGAUG Neofedrizzia GU(GGCG-GCCAAGCGUUCAUAGCGACGUCGCU)UUUUGAUCCUUCGAUG Megistheunus GU(GGCG-GC? ?AUCGUUCAU7UCUACGUCGCU) UGUUUAUCCUUCGAUG Trichodiplog GU(GGUGUGCCAAGCGUUCAUAGCGACGUCACU)UUUUGAUCCUUCGAUG Passalacarus GU(GGUGUGCCAAGCGUUCAUAGCGACGUCACU)UUUUGAUCCUUUGAUG Euzercon GU(GGUC-GCCAAGCGUUCAUAGCGACGUGACU) UUUUGAUCCUUCGAUG Neoeuzercon GU(GGCC-GCCAAGCGUUC?UAGC?AC??GG?U)UUU3GAUCCUUCGAUG P o ly a sp is GU(GGC ? -GCCA3GCGUUCAUAGCGACGUUGCU) UUUUGACCCUUCGAUG O p litis GU(GGUG-GCCAAGCGUUCAUAGCGACGUCACU)UUUUGAUCCUUCGAUG Asternolaelap GU(GGCG-GCCAAGCGUUCAUAGCGACGUCGCU)UUUUGAUCCUUCGAUG Epicroseius GU(GGCG-GCCAAGCGUUCAUAGCGACGUCGCU)UUUUGAUCCUUCGAUG S ej us GU(GGCG-GCCAAGCGUUCAUAGCGACGUCGCU)UUUUGAUCCUUCGAUG Discozercon GU(GGCG-GCCAAGCGUUCAUAGCGACGUCGCU)UUUUGAUCCUUCGAUG Heterozerconid GU(GGCA-GCCAAGCGUUCAUAGCGACGUUGCU)UUUUGAUCCUUCGAUG
[ 90 ] ( 91 Astem oseius [UCGGCUCUUCCU] AU(CAUUGUG Cercoleipus [UCGGCUCUUCCU] AU(CAUUGUG Mi cromegistus [UCGGCUCUUCCU] AU(CAUUGUG Promegistus [UCGGCUCUUUCU] AU(CAUUGUG Paramegistus (UCGGCUCUUCCU] AU(CAUUGUG Antennophorus [UCGGCUCUUCCU] AU(CAUUGUG Klinkowstro [UCGGCUCUUCCU]AU(CAUUGUG F e d rizzia [UCGGCUCUUCCU]AU(CAUUGUG Neofedrizzia [UCGGCUCUUCCU] AU(CAUUGUG Megisthanus [UCGGCUCUUCCU]AU(CAUUGUG Trichodiplog [UCGGCUCUUCCU] AU(CAUUGUG Passalacarus [UCGGCUCUUCCU] AU(CAUUGUG Euzercon [UCGGCUCUUCCU]AU(CAUUGUG Neoeuzercon [UCGGCUCUUCCU]AU(CAUUGUG P o ly a sp is [UCGGCUCUUCCU]AU(CAUUGUG O p litis [UCGGCUCUUCCU]AU(CAUUGUG Astemolaelap [UCGGCUCUUCCU] AU(CAUUGUG Epicroseius [UCGGCUCUUCCU]A U(CAUUGUG S eju s [UCGGCUCUUCCU] AU(CAUUGUG Discozercon [UCGGCUCUUCCU] AU(CAUUGUG Heterozerconid [UCGGCUCUUCCU]AU(CAUUGUG
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