Phylogenetics and Host Relationships of Plantbugs Inhabiting Native Cypress Pine Celia Symonds The University of Sydney and the Australian Museum Honours in Biological Science 2005 Supervisors Dieter Hochuli & Gerry Cassis

Introduction “Much of evolution is coevolution” (John Thompson (2004) The Geographic Mosaic Theory of Coevolution) However, the study of coevolution has been slow to progress In part, due to:  the complexity of species interactions, and  the lack of phylogenies for interacting organisms, to enable cospeciation analysis.

Coevolution and Cospeciation Coevolution reciprocal adaptation between two interacting organisms e.g. Ehrlich and Raven’s study of butterflies and their larval host plants (1964) Cospeciation the joint speciation of two or more lineages that are ecologically associated coevolution that occurs in macroevolutionary time e.g. classic example is between a host and its parasite

Describing Cospeciation Assessing congruence between phylogenies (relationship trees) of two interacting organisms Source: Page ed. (2003) Tangled Trees Gopher Phylogeny Louse Phylogeny

Cospeciation and other processes Very few cases, mostly animal parasites and animal hosts. Plant-insect associations are one of the most dominant interactions in natural systems. Few clear coevolutionary relationships established between plants and insects. Noise in the data to date…….. Source: Page ed. (2003) Tangled Trees

The aim of my project To investigate the potential for cospeciation to explain the relationship between a host specific group of undescribed plantbugs and native cypress pine.

Study System - Plantbugs (Miridae) sp. 22, ex. Callitris preissii, WA sp. 25, ex. Callitris rhomboidea, NSW

Native Cypress Pine Callitris & Actinostrobus Callitris bayleyi, NSW Callitris gracilis, VIC Callitris rhomboidea NSW Callitris sp. ‘Emerald Creek’, QLD Callitris intratropica, QLD

Approach Species discrimination – morphospecies Morphological analysis – comparative taxonomic study Molecular analysis – sequencing of 2 mitochondrial DNA loci, 16s & COI Phylogenetic analysis Separate and combined analysis of morphological and molecular data using maximum parsimony in PAUP* Cospeciation analysis Biogeographic associations?

Morphological Characters 1mm sp. 25

Morphological Characters 0.5mm Dorsal vestiture simple setae simple & scale like setae sp. 7 sp. 17

Pygophore ultimate abdominal segment of male plantbugs Diagnostic characters include: dorsal opening lobes tubercles left and right parameres Morphological Characters sp. 23 sp. 25 sp. 7 Dorsal Ventral

Parameres (male “claspers”) left right Morphological Characters 0.1mm sp. 4 sp. 10 sp. 13 sp. 20 sp. 26

Aedeagus (male genitalia) Morphological Characters sp. 23sp. 25sp. 7

Morphological analysis Findings: 66 characters, of which 62 are of the male genitalia 31 species from total of 42 localities and 13 Cypress pine species complex associations –up to 3 species from the same locality –29 species of plantbugs found on only one Cypress pine species –multiple species of plantbugs from a single Callitris species (up to 10)

Morphology strict consensus tree ] Three main species groups: One strongly supported Higher level: Strong support for species groups Basally: Unresolved Synapomorphy support for species groupings outgroup Bootstrap support Bootstrap support >80

Molecular analysis Findings: 16s - ~500bp sequence for 17 species COI - ~500bp sequence for 10 species Limitations: Incomplete data set Generally only one sample per species

Molecular – 16s strict consensus (MP) Similar patterns: strong support for species groups observed in morphological data basally unresolved some conflict with morphology in placing of particular species Likelihood analysis supporting parsimony results sp25 sp24 sp15 sp5 sp12 sp16 sp29 sp8 sp1 sp3 sp10 sp11 sp20 sp21 sp22 sp9 O. clermontiella sp27 Austromiris sp. Bootstrap support Bootstrap support >80

Molecular – COI strict consensus Similar patterns: support for species groups observed in morphology and 16s basally unresolved some differing placement of species to 16s tree artefact of small sample Bootstrap support Bootstrap support >80 sp3 sp10 sp29 sp25 sp26 sp28 sp1 sp12 sp16 sp9 O. clermontiella O. marginalis Austromiris sp.

Cospeciation analysis Method: Tree reconciliation analysis approach using the ‘Jungles’ method in TreeMap 2.0 (Charleston and Page 2002): –Mapping of parasites (plantbugs) onto the host (plant) tree generating all potentially optimal solutions –Assessed for significance against a cospeciation distribution pattern from random host and parasite trees –Detects: cospeciation host switching duplication (intrahost speciation) extinctions (sorting events) Separate analysis of morphological and molecular trees undertaken, due to unresolved basal relationships in the plantbug phylogenies

Cospeciation analysis Tanglegram for morphological data Patterns found: high level of host switching no clear cut pattern of cospeciation most plantbugs species highly specific to one host plant species 2 bug species found to inhabit more than one host – host switching phylogenetically conserved ‘missing the boat’ or extinction events speciation in plantbugs occurring below the level of speciation in the plants host plant plantbug

Historical Biogeography Method: Three area statement analysis (TAS: Nelson & Ladiges, 1991, Syst. Zool. 40). Areas of endemism from Crisp et al. (1995; Syst. Biol. 44) and Cracraft (1991; Aust. Syst. Bot. 4). Findings: East / west coast divide - confers with Crisp et al. Novel area relationship - Eyre Peninsula + (Adelaide + Eastern Desert). From: Crisp et al. (1995)

Conclusions 1.This plantbug and cypress pine interaction is not a clear cospeciation relationship. 2.Biogeography may better explain the plantbug host relationships. Host specificity is not a sufficient criterion for predicting parallel cladogenesis. Insect-plant relationships are more complex than conventional host-parasite relationships and thus reciprocal evolution needs to be examined in the context of their: direct 1:1 relationships, their historical biogeography, and other selection pressures.

Acknowledgements My supervisors Dieter Hochuli and Gerry Cassis My colleagues Michael Wall and Nik Tatarnic Mike Charleston at the University of Sydney Gary Nelson at the University of Melbourne Illustrations by Hannah Finlay SEMs by Sue Lindsay