Nathan Lo | The University of Sydney (original) (raw)

Papers by Nathan Lo

Following the acceptance of plate tectonics theory in the latter half of the 20th century, vicari... more Following the acceptance of plate tectonics theory in the latter half of the 20th century, vicariance became the dominant explanation for the distributions of many plant and animal groups. In recent years, however, molecular-clock analyses have challenged a number of well-accepted hypotheses of vicariance. As a widespread group of insects with a fossil record dating back 300 My, cockroaches provide an ideal model for testing hypotheses of vicariance through plate tectonics versus transoceanic dispersal. However, their evolutionary history remains poorly understood, in part due to unresolved relationships among the nine recognized families. Here, we present a phylogenetic estimate of all extant cockroach families, as well as a timescale for their evolution, based on the complete mitochondrial genomes of 119 cockroach species. Divergence dating analyses indicated that the last common ancestor of all extant cockroaches appeared 235Ma,235 Ma, 235Ma,95 My prior to the appearance of fossils that can be assigned to extant families, and before the breakup of Pangaea began. We reconstructed the geographic ranges of ancestral cockroaches and found tentative support for vicariance through plate tectonics within and between several major lineages. We also found evidence of transoceanic dispersal in lineages found across the Australian, Indo-Malayan, African, and Madagascan regions. Our analyses provide evidence that both vicariance and dispersal have played important roles in shaping the distribution and diversity of these insects.

Rhinoceros (rhinos) have suffered a dramatic increase in poaching over the past decade due to the... more Rhinoceros (rhinos) have suffered a dramatic increase in poaching over the past decade due to the growing demand for rhino horn products in Asia. One way to reverse this trend is to enhance enforcement and intelligence gathering tools used for species identification of horns, in particular making them fast, inexpensive and accurate. Traditionally, species identification tests are based on DNA sequence data, which, depending on laboratory resources, can be either time or cost prohibitive. This study presents a rapid rhino species identification test, utilizing species-specific primers within the cytochrome b gene multiplexed in a single reaction, with a presumptive species identification based on the length of the resultant ampli-con. This multiplex PCR assay can provide a presumptive species identification result in less than 24 hours. Sequence-based definitive testing can be conducted if/when required (e.g. court purposes). This work also presents an actual casework scenario in which the pre-sumptive test was successfully utlitised, in concert with sequence-based definitive testing. The test was carried out on seized suspected rhino horns tested at the Institute of Ecology and Biological Resources, the CITES mandated laboratory in Vietnam, a country that is known to be a major source of demand for rhino horns. This test represents the basis for which future 'rapid species identification tests' can be trialed.

Aim: The ways in which abiotic factors contribute to parallel evolution-the evolution of similar,... more Aim: The ways in which abiotic factors contribute to parallel evolution-the evolution of similar, derived phenotypes in independent, closely related lineages-remain understudied. Australian cockroaches of the subfamilies Panesthiinae ("wood feed-ers") and Geoscapheinae ("soil burrowers") are two closely related groups that provide a striking example of parallel evolution of burrowing behaviour. The ancestral wood-feeding panesthiines migrated from Asia ~20 million years ago before soil burrowing was independently acquired multiple times in the derived geoscapheines. Here, we investigate whether specific abiotic factors were associated with the parallel evolution of soil burrowing behaviour, and whether divergence events of geosca-pheines from panesthiine ancestors are consistent with niche conservatism or divergence. Location: The Australian mainland, including the areas in which selected Australian Panesthiinae and Geoscapheinae cockroach species are distributed. Methods: We generated environmental niche models for members of the Australian Geoscapheinae and Panesthiinae using presence-only data and abiotic variables related to temperature, precipitation, and soil composition from BioClim and the Australian Soil Resource Information System. We used an existing phylogenetic framework to compare environmental niche models and tested for niche conservatism versus divergence. Morphological convergence was assessed by a regression analysis and principal components analysis of leg segment and body dimensions in soil burrowers and wood feeders. Results: We found no relationship between niche similarity and time since divergence , and only limited evidence for phylogenetic signal with respect to the environmental variables examined. We found that soil burrowing behaviour is consistently correlated with thirteen abiotic factors associated with aridity, including a wider range of temperatures and lower precipitation levels. Evidence for convergence in leg morphology and body dimensions across soil burrowers was found. Main conclusions: Our results are consistent with soil burrowing behaviour evolving in response to ancient aridification events following the arrival of the Panesthiinae in Australia. Our results suggest a scenario of niche divergence between soil bur-rowers and each of their wood feeding sister taxa. There is evidence for morphological convergence on a "shovel-like" protibiotarsus in the Geoscapheinae that would aid in burrowing into soil.

Background: Sexual reproduction is the norm in almost all animal species, and in many advanced an... more Background: Sexual reproduction is the norm in almost all animal species, and in many advanced animal societies, both males and females participate in social activities. To date, the complete loss of males from advanced social animal lineages has been reported only in ants and honey bees (Hymenoptera), whose workers are always female and whose males display no helping behaviors even in normal sexual species. Asexuality has not previously been observed in colonies of another major group of social insects, the termites, where the ubiquitous presence of both male and female workers and soldiers indicate that males play a critical role beyond that of reproduction. Results: Here, we report asexual societies in a lineage of the termite Glyptotermes nakajimai. We investigated the composition of mature colonies from ten distinct populations in Japan, finding six asexual populations characterized by a lack of any males in the reproductive, soldier, and worker castes of their colonies, an absence of sperm in the spermathecae of their queens, and the development of unfertilized eggs at a level comparable to that for the development of fertilized eggs in sexual populations of this species. Phylogenetic analyses indicated a single evolutionary origin of the asexual populations, with divergence from sampled sexual populations occurring about 14 million years ago. Asexual colonies differ from sexual colonies in having a more uniform head size in their all-female soldier caste, and fewer soldiers in proportion to other individuals, suggesting increased defensive efficiencies arising from uniform soldier morphology. Such efficiencies may have contributed to the persistence and spread of the asexual lineage. Cooperative colony foundation by multiple queens, the single-site nesting life history common to both the asexual and sexual lineages, and the occasional development of eggs without fertilization even in the sexual lineage are traits likely to have been present in the ancestors of the asexual lineage that may have facilitated the transition to asexuality.

Almost all examined cockroaches harbor an obligate intracellular endosymbiont, Blattabacterium cu... more Almost all examined cockroaches harbor an obligate intracellular endosymbiont, Blattabacterium cuenoti. On the basis of genome content, Blattabacterium has been inferred to recycle nitrogen wastes and provide amino acids and cofactors for its hosts. Most Blattabacterium strains sequenced to date harbor a genome of 630 kbp, with the exception of the termite Mastotermes darwiniensis (590 kbp) and Cryptocercus punctulatus (614 kbp), a representative of the sister group of termites. Such genome reduction may have led to the ultimate loss of Blattabacterium in all termites other than Mastotermes. In this study, we sequenced 11 new Blattabacterium genomes from three species of Cryptocercus in order to shed light on the genomic evolution of Blattabacterium in termites and Cryptocercus. All genomes of Cryptocercus-derived Blattabacterium genomes were reduced (614 kbp), except for that associated with Cryptocercus kyebangensis, which comprised 637 kbp. Phylogenetic analysis of these genomes and their content indicates that Blattabacterium experienced parallel genome reduction in Mastotermes and Cryptocercus, possibly due to similar selective forces. We found evidence of ongoing genome reduction in Blattabacterium from three lineages of the C. punctulatus species complex, which independently lost one cysteine biosynthetic gene. We also sequenced the genome of the Blattabacterium associated with Salganea taiwanensis, a subsocial xylophagous cockroach that does not vertically transmit gut symbionts via proctodeal trophallaxis. This genome was 632 kbp, typical of that of nonsubsocial cockroaches. Overall, our results show that genome reduction occurred on multiple occasions in Blattabacterium, and is still ongoing, possibly because of new associations with gut symbionts in some lineages.

We investigated the phylogenetic diversity, localisation and metabolism of an uncultured bacteria... more We investigated the phylogenetic diversity, localisation and metabolism of an uncultured bacterial clade, Termite Group 2 (TG2), or ZB3, in the termite gut, which belongs to the candidate phylum 'Margulisbacteria'. We performed 16S rRNA amplicon sequencing analysis and detected TG2/ZB3 sequences in 40 out of 72 termite and cockroach species, which exclusively constituted a monophyletic cluster in the TG2/ZB3 clade. Fluorescence in situ hybridisation analysis in lower termites revealed that these bacteria are specifically attached to ectosymbiotic spirochetes of oxymonad gut protists. Draft genomes of four TG2/ZB3 phylotypes from a small number of bacterial cells were reconstructed, and functional genome analysis suggested that these bacteria hydrolyse and ferment cellulose/cellobiose to H 2 , CO 2 , acetate and ethanol. We also assembled a draft genome for a partner Treponema spirochete and found that it encoded genes for reductive acetogenesis from H 2 and CO 2. We hypothesise that the TG2/ZB3 bacteria we report here are commensal or mutualistic symbionts of the spirochetes, exploiting the spirochetes as H 2 sinks. For these bacteria, we propose a novel genus, 'Candidatus Termititenax', which represents a hitherto uncharacterised class-level clade in 'Margulisbacteria'. Our findings add another layer, i.e., cellular association between bacteria, to the multi-layered symbiotic system in the termite gut.

The mantis shrimp superfamily Squilloidea, with over 185 described species, is the largest superf... more The mantis shrimp superfamily Squilloidea, with over 185 described species, is the largest superfamily in the crustacean order Stomatopoda. To date, phylogenetic relationships within this superfamily have been comprehensively analysed using morphological data, with six major generic groupings being recovered. Here, we infer the phylogeny of Squilloidea using a combined dataset comprising 75 somatic morphological characters and four molecular markers. 5 Nodal support is low when the morphological and molecular datasets are analysed separately but improves substantially when combined in a total-evidence phylogenetic analysis. We obtain a well resolved and strongly supported phylogeny that is largely congruent with previous estimates except that the Anchisquilloides-group, rather than the Meiosquilla-group, is the earliest-branching lineage in Squilloidea. The splits among the Anchisquilloides-and Meiosquilla-groups are followed by those of the Clorida-, Harpiosquilla-, Squilla-and Oratosquilla-groups. Most of the generic groups are 10 recovered as monophyletic, with the exception of the Squilla-and Oratosquilla-groups. However, many genera within the Oratosquilla-group are not recovered as monophyletic. Further exploration with more extensive molecular sampling will be needed to resolve relationships within the Oratosquilla-group and to investigate the adaptive radiation of squilloids. Overall, our results demonstrate the merit of combining morphological and molecular datasets for resolving phylogenetic relationships.

Biological invasions represent a major threat to agriculture and forestry across the globe. Thaum... more Biological invasions represent a major threat to agriculture and forestry across the globe. Thaumastocoris peregrinus is a small sap-sucking heteropteran bug that has recently invaded a number of eucalypt plantations worldwide from its native range in Australia. To date, no studies have examined the range of this insect within Australia, and its population genetics remain poorly understood. We sampled T. peregrinus from 16 populations from South East Queensland, across New South Wales (NSW) and Victoria to southeastern South Australia, and generated microsatellite and mtDNA data for ~ 200 individuals. Population genetic analyses consistently revealed moderate levels of genetic isolation by distance among populations across the range. Nonetheless, T. peregrinus has undergone dispersal across large distances, as revealed by the presence of identical mitochondrial haplotypes in both South East Queensland and South Australia. Two populations within the Sydney area (NSW) were divergent from other populations based on STRU CTU RE and factorial correspondence analysis. They also had relatively low allelic richness and haplotype diversity indices. These results suggest they are the result of a relatively recent invasion event, consistent with their discovery in 2001. Pairwise genetic distance analyses suggest that the source of the invasion may have been central NSW. Our study provides an important framework for understanding the biology of this pest in its native environment, and may have implications for determining how it has invaded multiple areas worldwide. Keywords Bronzing bug · Hemiptera · Invasive insect pest · Microsatellites · Mitochondrial COI · Plantation forestry Key Message • Thaumastocoris peregrinus is an emerging pest of euca-lypt plantations worldwide; however, no studies have examined the population genetics of this species in its native range of Australia. • We sequenced both mitochondrial and microsatellite DNA from 16 T. peregrinus populations in order to investigate the level of genetic structure of this species across its native range, and test the hypothesis that individuals in Sydney represent a recent invasion. • We provide evidence that the insect in Sydney is distinct from other populations, and that it appears to be a recent invasion, whose source may be central NSW.

Termite guts harbor diverse yet-uncultured bacteria, including a non-photosynthetic cyanobacteria... more Termite guts harbor diverse yet-uncultured bacteria, including a non-photosynthetic cyanobacterial group, the class " Melainabacteria ". We herein reported the phylogenetic diversity of " Melainabacteria " in the guts of diverse termites and conducted a single-cell genome analysis of a melainabacterium obtained from the gut of the termite Termes propinquus. We performed amplicon sequencing of 16S rRNA genes from the guts of 60 termite and eight cockroach species, and detected melainabacterial sequences in 48 out of the 68 insect species, albeit with low abundances (0.02–1.90%). Most of the melain-abacterial sequences obtained were assigned to the order " Gastranaerophilales " and appeared to form clusters unique to termites and cockroaches. A single-cell genome of a melainabacterium, designated phylotype Tpq-Mel-01, was obtained using a fluorescence-activated cell sorter and whole genome amplification. The genome shared basic features with other melainabacterial genomes previously reconstructed from the metagenomes of human and koala feces. The bacterium had a small genome (~1.6 Mb) and possessed fermentative pathways possibly using sugars and chitobiose as carbon and energy sources, while the pathways for photosynthesis and carbon fixation were not found. The genome contained genes for flagellar components and chemotaxis; therefore, the bacterium is likely motile. A fluorescence in situ hybridization analysis showed that the cells of Tpq-Mel-01 and/or its close relatives are short rods with the dimensions of 1.1±0.2 μm by 0.5±0.1 μm; for these bacteria, we propose the novel species, " Candidatus Gastranaerophilus termiticola ". Our results provide fundamental information on " Melainabacteria " in the termite gut and expand our knowledge on this underrepresented, non-photosynthetic cyanobacterial group.

Cockroaches are among the most recognizable of all insects. In addition to their role as pests, t... more Cockroaches are among the most recognizable of all insects. In addition to their role as pests, they play a key ecological role as decomposers. Despite numerous studies of cockroach phylogeny in recent decades, relationships among most major lineages are yet to be resolved. Here we examine phylogenetic relationships among cockroaches based on five genes (mitochondrial 12S rRNA, 16S rRNA, COII; nuclear 28S rRNA and histone H3), and infer divergence times on the basis of 8 fossils. We included in our analyses sequences from 52 new species collected in China, representing 7 families. These were combined with data from a recent study that examined these same genes from 49 species, resulting in a significant increase in taxa analysed. Three major lineages, Corydioidea, Blaberoidea, and Blattoidea were recovered, the latter comprising Blattidae, Tryonicidae, Lamproblattidae, Anaplectidae, Cryptocercidae and Isoptera. The estimated age of the split between Mantodea and Blattodea ranged from 204.3 Ma to 289.1 Ma. Corydioidea was estimated to have diverged 209.7 Ma (180.5–244.3 Ma 95% confidence interval [CI]) from the remaining Blattodea. The clade Blattoidea diverged from their sister group, Blaberoidea, around 198.3 Ma (173.1–229.1 Ma). The addition of the extra taxa in this study has resulted in significantly higher levels of support for a number of previously recognized groupings.

The marine metazoan fauna first diversified in the early Cambrian, but terrestrial environments w... more The marine metazoan fauna first diversified in the early Cambrian, but terrestrial environments were not colonized until at least 100 million years later. Among the groups of organisms that successfully colonized land is the crustacean order Isopoda. Of the 10,000 described isopod species, ~ 3,600 species from the suborder Oniscidea are terrestrial. Although it is widely thought that isopods colonized land only once, some studies have failed to confirm the monophyly of Oniscidea. To infer the evolutionary relationships among isopod lineages, we conducted phylogenetic analyses of nuclear 18S and 28S and mitochondrial COI genes using maximum-likelihood and Bayesian methods. We also analyzed a second data set comprising all of the mitochondrial protein-coding genes from a smaller sample of isopod taxa. Based on our analyses using a relaxed molecular clock, we dated the origin of terrestrial iso-pods at 289.5 million years ago (95% credibility interval 219.6– 358.9 million years ago). These predate the known fossil record of these taxa and coincide with the formation of the supercon-tinent Pangaea and with the diversification of vascular plants on land. Our results suggest that the terrestrial environment has been colonized more than once by isopods. The monophyly of the suborder Oniscidea was not supported in any of our analyses, conflicting with classical views based on morphology. This draws attention to the need for further work on this group of isopods.

—In Bayesian phylogenetic analyses of genetic data, prior probability distributions need to be sp... more —In Bayesian phylogenetic analyses of genetic data, prior probability distributions need to be specified for the model parameters, including the tree. When Bayesian methods are used for molecular dating, available tree priors include those designed for species-level data, such as the pure-birth and birth–death priors, and coalescent-based priors designed for population-level data. However, molecular dating methods are frequently applied to data sets that include multiple individuals across multiple species. Such data sets violate the assumptions of both the speciation and coalescent-based tree priors, making it unclear which should be chosen and whether this choice can affect the estimation of node times. To investigate this problem, we used a simulation approach to produce data sets with different proportions of within-and between-species sampling under the multispecies coalescent model. These data sets were then analyzed under pure-birth, birth–death, constant-size coalescent, and skyline coalescent tree priors. We also explored the ability of Bayesian model testing to select the best-performing priors. We confirmed the applicability of our results to empirical data sets from cetaceans, phocids, and coregonid whitefish. Estimates of node times were generally robust to the choice of tree prior, but some combinations of tree priors and sampling schemes led to large differences in the age estimates. In particular, the pure-birth tree prior frequently led to inaccurate estimates for data sets containing a mixture of inter-and intraspecific sampling, whereas the birth–death and skyline coalescent priors produced stable results across all scenarios. Model testing provided an adequate means of rejecting inappropriate tree priors. Our results suggest that tree priors do not strongly affect Bayesian molecular dating results in most cases, even when severely misspecified. However, the choice of tree prior can be significant for the accuracy of dating results in the case of data sets with mixed inter-and intraspecies sampling.

Aim The Australian Coptotermes (Family: Rhinotermitidae) are a small mono-phyletic group of termi... more Aim The Australian Coptotermes (Family: Rhinotermitidae) are a small mono-phyletic group of termites, some of which build mounds. In this study, we construct predicted distributions based on environmental data (niche models) for all Australian species of Coptotermes to test whether specific environmental factors have contributed to the evolution of mound-building behaviour and whether the degree of niche similarity and degree of phylogenetic similarity are correlated. Location The Australian mainland, including the known native ranges of all Australian species of Coptotermes. Methods We estimated the phylogenetic relationships between the species of Australian Coptotermes. We then generated and compared environmental niche models in a phylogenetic framework for all study species to test niche conservation. Our analyses were based on location data from our own sampling and from the Atlas of Living Australia, genetic data from a previous study of Australian Coptotermes, and environmental data from WorldClim and ASRIS. Results We found that no environmental variable differed consistently between mound-building and non-mound-building taxa and that the differences in niches between pairs of Australian species of Coptotermes are uncorre-lated with time since divergence. The environmental tolerances of the Australian Coptotermes termites are more restricted by rainfall than they are by soil or temperature. Main conclusions Our results show that mound-building behaviour has not necessarily evolved in response to similar abiotic conditions. Our results are consistent with ecological speciation leading to niche divergence since Coptoter-mes first arrived in Australia ~12.5 million years ago.

The crustacean order Stomatopoda comprises seven superfamilies of mantis shrimps, found in coasta... more The crustacean order Stomatopoda comprises seven superfamilies of mantis shrimps, found in coastal waters of the tropics and subtropics. These marine carnivores bear notable raptorial appendages for smashing or spearing prey. We investigated the evolutionary relationships among stomatopods using phylogenetic analyses of three mitochondrial and two nuclear markers. Our analyses recovered the superfamily Gonodactyloidea as polyphyletic, with Hemisquilla as the sister group to all other extant stomatopods. A relaxed molecular clock, calibrated by seven fossil-based age constraints, was used to date the origin and major diversification events of stomatopods. Our estimates suggest that crown-group stomatopods (Unipeltata) diverged from their closest crustacean relatives about 340 Ma (95% CRI [401–313 Ma]). We found that the specialized smashing appendage arose after the spearing appendage ∼126 Ma (95% CRI [174–87 Ma]). Ancestral state reconstructions revealed that the most recent common ancestor of extant stomatopods had eyes with six midband rows of hexagonal ommatidia. Hexagonal ommatidia are interpreted as plesiomorphic in stomatopods, and this is consistent with the malacostracan ground-plan. Our study provides insight into the evolutionary timescale and systematics of Stomatopoda, although further work is required to resolve with confidence the phylogenetic relationships among its superfamilies.

Termites differ from hymenopteran social insects in several important respects, perhaps most sign... more Termites differ from hymenopteran social insects in several important respects, perhaps most significantly in their sex and caste determination systems: while hyme-nopteran colonies are female dominated (and sex determination is haplo-diploid), termite colonies are usually split evenly between males and females (and sex determination is through sex chromosomes). Not all termite species have an equal sex ratio—in the termite genus Schedo-rhinotermes, almost all workers and soldiers are females. The mechanism maintaining this sex ratio skew is unknown, but a possible mechanism (known in other termites as a mechanism for producing reproductives) is parthenogene-sis. Under this scenario, soldiers and workers would be offspring of the queen only. In this study, we performed microsatellite analysis on 11 colonies of Schedorhinotermes intermedius (Isoptera: Rhinotermitidae) to investigate the parentage of workers, soldiers and alates within colonies, and to determine whether parthenogenesis is responsible for the production of female workers and soldiers. We also conducted a preliminary analysis of population and colony genetic structure. We found that females from a single colony had in some instances more than two alleles among them at a single microsatellite locus. This indicates that a single female cannot be producing these offspring, as she has a maximum of two different alleles at any locus, ruling out the possibility that the high proportion of females in this species comes about through parthenogenesis.

Woodroaches from the genus Cryptocercus Scudder, 1862 are known to display low levels of morpholo... more Woodroaches from the genus Cryptocercus Scudder, 1862 are known to display low levels of morphological divergence, yet significant genetic divergence and variability in chromosome number. Compared with Cryptocercus taxa from North America, the diversity of the genus in Asia has received relatively little attention. We performed morphological and karyotypic examinations of multiple taxa from several previously unsampled mountainous areas of central and southwestern China, and identified nine candidate species primarily on the basis of chromosome number. We then investigated diversity across all Asian Cryptocercus, through phylogenetic analyses of 135 COI sequences and 74 28S rRNA sequences from individuals of 28 localities, including species delimitation analysis in General Mixed Yule Coalescent (GMYC) and Automatic Barcode Gap Discovery (ABGD). Phylogenetic results indicated that individuals from the same locality constituted well supported clades. The congruence of GMYC and ABGD results were in almost perfect accord, with 28 candidate species described on the basis of karyotypes (including the nine identified in this study). We provide evidence that each valley population in the Hengduan Mountains contains a separate evolving lineage. We conclude that the principal cause of the rich Cryptocercus diversity in China has been the uplift of the Qinghai-Tibet Plateau.