Uncovering the magnitude of African pangolin poaching with extensive nanopore DNA genotyping of seized scales.

Trade in pangolins is illegal, and yet tons of their scales and products are seized at various ports. These large seizures are challenging to process and comprehensively genotype for upstream provenance tracing and species identification for prosecution. We implemented a scalable DNA barcoding pipeline in which rapid DNA extraction and MinION sequencing were used to genotype a substantial proportion of pangolin scales subsampled from 2 record shipments seized in Singapore in 2019 (37.5 t). We used reference sequences to match the scales to phylogeographical regions of origin. In total, we identified 2346 cytochrome b (cytb) barcodes of white-bellied (Phataginus tricuspis) (from 1091 scales), black-bellied (Phataginus tetradactyla) (227 scales), and giant (Smutsia gigantea) (1028 scales) pangolins. Haplotype diversity was higher for P. tricuspis scales (121 haplotypes, 66 novel) than that for P. tetradactyla (22 haplotypes, 15 novel) and S. gigantea (25 haplotypes, 21 novel) scales. Of the novel haplotypes, 74.2% were likely from western and west-central Africa, suggesting potential resurgence of poaching and newly exploited populations in these regions. Our results illustrate the utility of extensively subsampling large seizures and outline an efficient molecular approach for rapid genetic screening that should be accessible to most forensic laboratories and enforcement agencies.

Revelación de la magnitud de la caza furtiva del pangolín africano mediante el genotipo extenso de nanoporos de ADN de escamas incautadas Resumen Aunque el mercado de pangolines es ilegal, se incautan toneladas de sus escamas y productos derivados en varios puertos comerciales. Es un reto procesar estas magnas incautaciones y obtener el genotipo completo para usarlo en la trazabilidad logística ascendente e identificación de la especie y así imponer sanciones. Implementamos una canalización escalable del código de barras de ADN en el cual usamos la extracción rápida de ADN y la secuenciación MinION para obtener el genotipo de una proporción sustancial de las escamas de pangolín submuestreadas en dos cargamentos incautados en 2019 en Singapur (37.5 t). Usamos secuencias referenciales para emparejar las escamas con las regiones filogeográficas de origen. Identificamos en total 2,346 códigos de citocromo b (cytb) del pangolín de vientre blanco (Phataginus tricuspis) (de 1,091 escamas), de vientre negro (P. tetradactyla) (227 escamas) y del pangolín gigante (Smutsia gigantea) (1,028 escamas). La diversidad de haplotipos fue mayor en las escamas de P. tricuspis (121 haplotipos, 66 nuevos) que en las de P. tetradactyla (22 haplotipos, 15 nuevos) y S. gigantea (25 haplotipos, 21 nuevos). De los haplotipos nuevos, el 74.2% probablemente provenía del occidente y centro­occidente de África, lo que sugiere un resurgimiento potencial de la caza furtiva y poblaciones recién explotadas en estas regiones. Nuestros resultados demuestran la utilidad de submuestrear extensivamente las grandes incautaciones y esboza una estrategia molecular eficiente para un análisis genético rápido que debería ser accesible para la mayoría de los laboratorios forenses y las autoridades de aplicación.

Isopod mouthpart traits respond to a tropical forest recovery gradient.

Functional trait ecology has the potential to provide generalizable and mechanistic predictions of ecosystem function from data of species distributions and traits. The traits that are selected should both respond to environmental factors and influence ecosystem functioning. Invertebrate mouthpart traits fulfill these criteria, but are seldom collected, lack standardized measurement protocols, and have infrequently been investigated in response to environmental factors. We surveyed isopod species that consume plant detritus, and tree communities in 58 plots across primary and secondary forests in Singapore. We measured body dimensions (body size traits), pereopod and antennae lengths (locomotory traits), dimensions of mandible structures (morphological mouthpart traits), and mechanical advantages generated by mandible shape (mechanical mouthpart traits) for six isopod species found in these plots and investigated if these traits respond to changes in tree community composition, tree diversity, and forest structure. Morphological mouthpart traits responded to a tree compositional gradient reflecting forest recovery degree. Mouthpart features associated with greater consumption of litter (broader but less serrated/rugose lacinia mobilis [an important cutting and chewing structure on the mandible]) were most prevalent in abandoned plantation and young secondary forests containing disturbance-associated tree species. Feeding strategies associated with fungi grazing (narrower and more serrated/rugose lacinia mobilis) were most prevalent in late secondary forests containing later successional tree species. Since morphological mouthpart traits likely also predict consumption and excretion rates of isopods, these traits advance our understanding of environment-trait-ecosystem functioning relationships across contrasting tropical forest plots that vary in composition, disturbance history, and post-disturbance recovery.

Why do understorey Licuala palm fruits turn from red to white and then black when ripe?

Licuala ferruginea Becc., a tropical forest understorey palm, is observed to have fruits that appear red in colour when unripe, turning pink, then white, purple and finally black in colour as they ripen. We monitored 13 fruiting palms in rainforest fragments and recorded the consumption of fruits by animals via camera traps. We also documented the fruiting phenology of two palms in the nursery. In the rainforest fragments, a Cream-vented Bulbul (Pycnonotus simplex) was observed plucking a mature purple fruit from a L. ferruginea palm, before flying away with the fruit in its beak. This was the only bird that was observed feeding on the mature fruit. A range of mammals, dominated by edge species such as the Long-tailed Macaque and Wild Boar, were observed to consume L. ferruginea fruits indiscriminately across all five colour stages, thereby limiting the dispersal of the fruits. Forest bulbul gape sizes also matched the fruit size, suggesting that forest bulbuls are the likely dispersers of the palm in the original forest where edge species are not in high densities. We further posit that the initial phase of red fruits, with high contrasting red reflectance against a green foliage background, might be a form of early advertisement to birds. The fruit then turns pink and white, which have high green reflectance and is less contrasting, thereby reducing the conspicuity of the fruit. This allows the fruit to ripen with high fructose and glucose content, and turn purple and black, which are known visual cues for birds. This study provides indicative support for the dispersal syndrome hypothesis and highlights the potential effects of forest fragmentation on plant-frugivore interactions.

Detection of a novel Babesia sp. in Amblyomma javanense, an ectoparasite of Sunda pangolins.

BACKGROUND: Babesia is a protozoal, tick-borne parasite that can cause life-threatening disease in humans, wildlife and domestic animals worldwide. However, in Southeast Asia, little is known about the prevalence and diversity of Babesia species present in wildlife and the tick vectors responsible for its transmission. Recently, a novel Babesia species was reported in confiscated Sunda pangolins (Manis javanica) in Thailand. To investigate the presence of this parasite in Singapore, we conducted a molecular survey of Babesia spp. in free-roaming Sunda pangolins and their main ectoparasite, the Amblyomma javanense tick. METHODS: Ticks and tissue samples were opportunistically collected from live and dead Sunda pangolins and screened using a PCR assay targeting the 18S rRNA gene of Babesia spp. DNA barcoding of the cytochrome oxidase subunit I (COI) mitochondrial gene was used to confirm the species of ticks that were Babesia positive. RESULTS: A total of 296 ticks and 40 tissue samples were obtained from 21 Sunda pangolins throughout the 1-year study period. Babesia DNA was detected in five A. javanense ticks (minimum infection rate = 1.7%) and in nine different pangolins (52.9%) located across the country. Phylogenetic analysis revealed that the Babesia 18S sequences obtained from these samples grouped into a single monophyletic clade together with those derived from Sunda pangolins in Thailand and that this evolutionarily distinct species is basal to the Babesia sensu stricto clade, which encompasses a range of Babesia species that infect both domestic and wildlife vertebrate hosts. CONCLUSIONS: This is the first report documenting the detection of a Babesia species in A. javanense ticks, the main ectoparasite of Sunda pangolins. While our results showed that A. javanense can carry this novel Babesia sp., additional confirmatory studies are required to demonstrate vector competency. Further studies are also necessary to investigate the role of other transmission pathways given the low infection rate of ticks in relation to the high infection rate of Sunda pangolins. Although it appears that this novel Babesia sp. is of little to no pathogenicity to Sunda pangolins, its potential to cause disease in other animals or humans cannot be ruled out.

Description of the larva of Brachygonia oculata (Brauer, 1878) (Odonata: Libellulidae) with notes on rearing technique.

The larva of the genus Brachygonia Kirby, 1889 is described and illustrated for the first time based on the larvae of Brachygonia oculata (Brauer, 1878) discovered and successfully reared in Singapore. Compared to known larvae from genera in the rather heterogeneous subfamily Brachydiplacinae, B. oculata is most similar to species from the genus Brachydiplax Brauer, 1868 after comparison is made with the larva of Brachydiplax farinosa Krüger, 1902.