John-James Wilson

Compositional heterogeneity in true bug mitochondrial phylogenomics

Mitochondrial phylogenomics is often controversial, in particular for inferring deep relationships. The recent rapid increase of mitochondrial genome data provides opportunities for better phylogenetic estimates and assessment of potential biases resulting from heterogeneity in nucleotide composition and mutation rates. Here, we gathered 76 mitochondrial genome sequences for Heteroptera representing all seven infraorders, including 17 newly sequenced mitochondrial genomes. We found strong heterogeneity in base composition and contrasting evolutionary rates among heteropteran mitochondrial genomes, which affected analyses with various datasets and partitioning schemes under site-homogeneous models and produced false groupings of unrelated taxa exhibiting similar base composition and accelerated evolutionary rates. Bayesian analyses using a site-heterogeneous mixture CAT+GTR model showed high congruence of topologies with the currently accepted phylogeny of Heteroptera. The results confirm the monophyly of the six infraorders within Heteroptera, except for Cimicomorpha which was recovered as two paraphyletic clades. The monophyly of Terheteroptera (Cimicomorpha and Pentatomomorpha) and Panheteroptera (Nepomorpha, Leptopodomorpha and Terheteroptera) was recovered demonstrating a significant improvement over previous studies using mitochondrial genome data. Our study shows the power of the site-heterogeneous mixture models for resolving phylogenetic relationships with Heteroptera and provides one more case showing that model adequacy is critical for accurate tree reconstruction in mitochondrial phylogenomics.

DNA barcodes for dragonflies and damselflies (Odonata) of Mindanao, Philippines

Abstract Reliable species identification provides a sounder basis for use of species in the order Odonata as biological indicators and for their conservation, an urgent concern as many species are threatened with imminent extinction. We generated 134 COI barcodes from 36 morphologically identified species of Odonata collected from Mindanao Island, representing 10 families and 19 genera. Intraspecific sequence divergences ranged from 0 to 6.7% with four species showing more than 2%, while interspecific sequence divergences ranged from 0.5 to 23.3% with seven species showing less than 2%. Consequently, no distinct gap was observed between intraspecific and interspecific DNA barcode divergences. The numerous islands of the Philippine archipelago may have facilitated rapid speciation in the Odonata and resulted in low interspecific sequence divergences among closely related groups of species. This study contributes DNA barcodes for 36 morphologically identified species of Odonata reported from Mindanao including 31 species with no previous DNA barcode records.

Impact of urbanisation and agriculture on the diet of fruit bats

The expansion of cities and agricultural plantations have unpredictable impacts on biodiversity and ecosystem services. Yet some species are capable of tolerating anthropogenic impacts and continue to provide ecological services in highly disturbed landscapes. The objective of this study was to use DNA barcoding to identify digested plant materials and seeds in the faeces of frugivorous bats (Cynopterus brachyotis) and investigate whether (1) C. brachyotis in urban and agricultural areas exploit cultivated and exotic plants as a novel food resource and as a consequence, potentially facilitate the invasion of cultivated and exotic plants, or whether (2) C. brachyotis exploit native plants and as a consequence, potentially promote forest regeneration. A native species, Ficus fistulosa, was the most frequently detected plant and the seeds were found in bat faeces from all sampling sites suggesting the potential of fruit bats in dispersing seeds. However, we also detected several exotic plants in the faeces of C. brachyotis which suggests that the fruit bats exploit novel food resources at all sites. We recorded a diverse diet of C. brachyotis at an oil palm plantation which indicated that the fruit bats are not predominantly feeding on oil palm fruits. By using DNA barcoding, we detected plants that have not been reported in previous studies of the diet of C. brachyotis, although we could not identify which part of the plant was being consumed by the fruit bats. Given the varied diet of C. brachyotis, the potential of this bat to adapt to changing landscapes is high and they are likely dispersing seeds of native pioneer plants (Ficus).

Temporal changes in arthropod activity in tropical anthropogenic forests.

Arthropod communities in the tropics are increasingly impacted by rapid changes in land use. Because species showing distinct seasonal patterns of activity are thought to be at higher risk of climate-related extirpation, global warming is generally considered a lower threat to arthropod biodiversity in the tropics than in temperate regions. To examine changes associated with land use and weather variables in tropical arthropod communities, we deployed Malaise traps at three major anthropogenic forests (secondary reserve forest, oil palm forest, and urban ornamental forest (UOF)) in Peninsular Malaysia and collected arthropods continuously for 12 months. We used metabarcoding protocols to characterize the diversity within weekly samples. We found that changes in the composition of arthropod communities were significantly associated with maximum temperature in all the three forests, but shifts were reversed in the UOF compared with the other forests. This suggests arthropods in forests in Peninsular Malaysia face a double threat: community shifts and biodiversity loss due to exploitation and disturbance of forests which consequently put species at further risk related to global warming. We highlight the positive feedback mechanism of land use and temperature, which pose threats to the arthropod communities and further implicates ecosystem functioning and human well-being. Consequently, conservation and mitigation plans are urgently needed.

Feeding behavior of Mimomyia (Etorleptiomyia) luzonensis (Ludlow, 1905) (Diptera, Culicidae) in Peninsular Malaysia

Mosquitoes are vectors of various human diseases in the tropics including yellow fever, dengue, malaria and West Nile virus. Mosquitoes can act as vectors between wildlife and humans, which is particularly important for diseases where wild animals serve as reservoirs of parasites in the absence of human infections. Research has mainly focused on the medical impacts of Anopheles, Aedes, Mansonia and Culex, however, very little attention has been directed towards other mosquito genera, especially those which act as vectors of diseases of wildlife. We have observed adults of Mimomyia (Etorleptiomyia) luzonensis (Ludlow, 1905) feeding on a toad, Ingerophrynus parvus, near an oil palm plantation settlement in Setia Alam, Selangor state, Peninsular Malaysia. Mimomyia is known to feed on reptiles and amphibians, and is a documented vector of several arboviruses, including West Nile virus. The observation of Mimomyia feeding on a common toad near a human settlement highlights a need to understand the relationships between mosquitoes, toads and humans from an ecological perspective. We report on-site observations of the feeding habit of Mimomyia; the first records from Malaysia.

A two-step DNA barcoding approach for delimiting moth species: moths of Dongling Mountain (Beijing, China) as a case study

DNA barcoding, based on a fragment of cytochrome c oxidase I (COI) mtDNA, is as an effective molecular tool for identification, discovery, and biodiversity assessment for most animals. However, multiple gene markers coupled with more sophisticated analytical approaches may be necessary to clarify species boundaries in cases of cryptic diversity or morphological plasticity. Using 339 moths collected from mountains surrounding Beijing, China, we tested a pipeline consisting of two steps: (1) rapid morphospecies sorting and screening of the investigated fauna with standard COI barcoding approaches; (2) additional analyses with multiple molecular markers for those specimens whose morphospecies and COI barcode grouping were incongruent. In step 1, 124 morphospecies were delimited into 116 barcode units, with 90% of the conflicts being associated with specimens identified to the genus Hypena. In step 2, 55 individuals representing all 12 Hypena morphospecies were analysed using COI, COII, 28S, EF-1a, Wgl sequences or their combinations with the BPP (Bayesian Phylogenetics and Phylogeography) multigene species delimitation method. The multigene analyses supported the delimitation of 5 species, consistent with the COI analysis. We conclude that a two-step barcoding analysis pipeline is able to rapidly characterize insect biodiversity and help to elucidate species boundaries for taxonomic complexes without jeopardizing overall project efficiency by substantially increasing analytical costs.

Pollination implications of the diverse diet of tropical nectar-feeding bats roosting in an urban cave

Background Intense landscaping often alters the plant composition in urban areas. Knowing which plant species that pollinators are visiting in urban areas is necessary for understanding how landscaping impacts biodiversity and associated ecosystem services. The cave nectar bat, Eonycteris spelaea, is an important pollinator for many plants and is often recorded in human-dominated habitats. Previous studies of the diet of E. spelaea relied on morphological identification of pollen grains found in faeces and on the body of bats and by necessity disregarded other forms of digested plant material present in the faeces (i.e., plant juice and remnants). The main objective of this study was to examine the diet of the nectarivorous bat, E. spelaea, roosting in an urban cave at Batu Caves, Peninsular Malaysia by identifying the plant material present in the faeces of bats using DNA metabarcoding. Methods Faeces were collected under the roost of E. spelaea once a week from December 2015 to March 2016. Plant DNA was extracted from the faeces, Polymerase chain reaction (PCR) amplified at ITS2 and rbcL regions and mass sequenced. The resultant plant operational taxonomic units were searched against NCBI GenBank for identification. Results A total of 55 species of plants were detected from faeces of E. spelaea including Artocarpus heterophyllus, Duabanga grandiflora and Musa spp. which are likely to be important food resources for the cave nectar bat. Discussion Many native plant species that had not been reported in previous dietary studies of E. spelaea were detected in this study including Bauhinia strychnoidea and Urophyllum leucophlaeum, suggesting that E. spelaea remains a crucial pollinator for these plants even in highly disturbed habitats. The detection of many introduced plant species in the bat faeces indicates that E. spelaea are exploiting them, particularly Xanthostemon chrysanthus, as food resources in urban area. Commercial food crops were detected from all of the faecal samples, suggesting that E. spelaea feed predominantly on the crops particularly jackfruit and banana and play a significant role in pollination of economically important plants. Ferns and figs were also detected in the faeces of E. spelaea suggesting future research avenues to determine whether the ‘specialised nectarivorous’ E. spelaea feed opportunistically on other parts of plants.

Tracking the southern river terrapin (Batagur affinis) through environmental DNA: prospects and challenges

Abstract Environmental DNA detection has emerged as a powerful tool to monitor aquatic species without the need for capture or visual identification and is particularly useful for rare or elusive species. Our objective was to develop an eDNA approach for detecting the southern river terrapin (Batagur affinis) in Malaysia. We designed species-specific primers for a fragment of B. affinis mtDNA and evaluated their effectiveness in silico, in vitro and in situ. The primers amplified 110 bp of the cytochrome b mtDNA sequence of B. affinis from aquarium water samples housing nine juvenile B. affinis. We also successfully detected B. affinis eDNA from river samples taken from a site where turtles were known to be in the vicinity. Prospects and challenges of using an eDNA approach to help determine the distribution of B. affinis, essential information for an effective conservation plan, are discussed.

High-throughput terrestrial biodiversity assessments: mitochondrial metabarcoding, metagenomics or metatranscriptomics?

Abstract Consensus on the optimal high-throughput sequencing (HTS) approach to examine biodiversity in mixed terrestrial arthropod samples has not been reached. Metatranscriptomics could increase the proportion of taxonomically informative mitochondrial reads in HTS outputs but has not been investigated for terrestrial arthropod samples. We compared the efficiency of 16S rRNA metabarcoding, metagenomics and metatranscriptomics for detecting species in a mixed terrestrial arthropod sample (pooled DNA/RNA from 38 taxa). 16S rRNA metabarcoding and nuclear rRNA-depleted metatranscriptomics had the highest detection rate with 97% of input species detected. Based on cytochrome c oxidase I, metagenomics had the highest detection rate with 82% of input species detected, but metatranscriptomics produced a larger proportion of reads matching (Sanger) reference sequences. Metatranscriptomics with nuclear rRNA depletion may offer advantages over metabarcoding through reducing the number of spurious operational taxonomic units while retaining high detection rates, and offers natural enrichment of mitochondrial sequences which may enable increased species detection rates compared with metagenomics.

Hookworm infections among migrant workers in Malaysia: molecular identification of Necator americanus and Ancylostoma duodenale

Ongoing urbanisation of the working population as well as cross-border migration of workers particularly into large cities has contributed to the development and growth of urban slums. These deprived areas are conducive for the transmission of intestinal pathogens including hookworm. The aim of this study was to determine both the prevalence and species identity of hookworm infections among the migrant worker community in Malaysia. A total of 388 faecal samples were collected from migrant workers between September 2014 and August 2015, representing workers from five employment sectors: construction, manufacturing, agriculture and plantations, food services and domestic services. Faecal samples were examined by microscopy and positive samples were subjected to molecular analysis. A total of 51 samples (13.1%) were positive by microscopy for hookworm infections. A two-step PCR based method amplifying a fragment of the 28S rRNA-ITS2 region was used to identify infections by Necator americanus and Ancylostoma spp. PCR products positive for Ancylostoma spp. were sequenced bidirectionally, and sequences analysed through BLAST and phylogenetic analysis. Samples containing Ancylostoma duodenale were further characterized by amplification and sequencing a fragment of cytochrome c oxidase subunit 1 (cox1) gene. PCR amplicons were successfully obtained from 42 (82.4%) of 51 samples, with 81.0% (34 of 42) identified as Necator americanus, 16.7% (7 of 42) as Ancylostoma spp. and 2.4% (1 of 42) as mixed infections of both species. All eight Ancylostoma spp. were confirmed to be Ancylostoma duodenale and this is the first time A. duodenale was reported in Malaysia. Samples containing A. duodenale from Nepalese and Indonesian workers shared high-similarity and were distinct compared to sequences from other countries. This study highlights the prevalence of hookworm infections among migrant workers living in Malaysia. Our findings underscore the necessity of screening migrant workers for hookworm infections, particularly those working in food-related services and industries.