Edinburgh Genomics

Belonocnema Mayr (Cynipidae, Cynipini) is an oak gall wasp genus currently known by three described species distributed in the southern and southeastern U.S. and northern Mexico, all associated with live oaks of the Quercus section Virentes. Here, we conducted an integrative systematic study of a new species of Belonocnema , B. mesoamericana Zhang, Zaldívar-Riverón & Fernández-Flores, sp. nov ., reared from galls of the live oak Quercus oleoides Schltdl. in the state of Veracruz, Mexico. We morphologically described the sexual and asexual generations of the new species and their galls, molecularly characterized it based on the DNA barcoding locus, reconstructed its phylogenetic affinities using multilocus data from Ultraconserved Elements, and identified its associated hymenopteran community. The new species is morphologically similar to its sister species, B. kinseyi Weld, though it can be distinguished by color and sculpture features, as well as by the sexual generation forming velvety rootlet galls. Since Q. oleoides is widely distributed across Mesoamerica, we expect the new species to have a broader distribution. A considerably high species richness and phylogenetic diversity of the hymenopteran community associated with the galls were recovered, with 43 morphospecies belonging to 15 genera and 13 families, of which 41 were found in the asexual generation and only three in the sexual generation. This study increases the systematic knowledge of a poorly known Cynipini genus and highlights the extraordinary, yet neglected, insect diversity associated with galls, especially in the Neotropics.

ABSTRACT Pathogens frequently produce proteins to evade or inhibit host immune responses. One such protein is lymphostatin from attaching and effacing Escherichia coli (also known as lymphocyte inhibitory factor A; LifA), which influences intestinal colonization and inhibits mitogen- and antigen-activated proliferation of T lymphocytes and pro-inflammatory cytokine synthesis. Here, we report the cloning, purification and characterization of a LifA homologue from Chlamydia pecorum. The predicted 382 KDa protein (CPE2_0552) exhibited 36% identity and 55% similarity over 3171 amino acids to lymphostatin from enteropathogenic E. coli strain E2348/69. CPE2_0552 shares glycosyltransferase and cysteine protease motifs required for lymphostatin activity, including similarity in the tertiary structure of these domains predicted by AlphaFold 3. Purified CPE2_0552 did not share the L-shaped globular structure of lymphostatin when analyzed by transmission electron microscopy. CPE2_0552 inhibited concanavalin A-stimulated proliferation of bovine T cells in a concentration-dependent manner, with an inhibitory dose 50 (ID50) of 812 pg/mL. This was 38-fold higher than the ID50 of E. coli E2348/69 lymphostatin tested in parallel on T cells from the same donors (21 pg/mL), but was similar to another LifA homologue from E. coli O157:H7 (ToxB). Moreover, CPE2_0552 inhibited the secretion of interferon gamma (IFNγ), a key cytokine that influences the outcome of Chlamydia infections. At the concentrations at which CPE2_0552 inhibited T lymphocyte proliferation and IFNγ secretion, negligible cytotoxicity was observed after 72 h of stimulation. Our study indicates that E. coli lymphostatin belongs to a wider family of lymphocyte-inhibitory molecules that exist in distantly related bacterial pathogens.

Background: Orofacial traits are influenced by a large number of loci; requiring large population-based studies for their detection. Public willingness to participate in and share three-dimensional (3D) image data from orofacial genetic research is essential to ensure feasible implementation of these studies. This preliminary study aims to explore the Scottish public’s willingness to participate in and share 3D image data from orofacial genetic research, providing baseline data for future research. Methods: The online survey was administered from the 14th June–14th July 2017. Multinomial logistic regression was used to test for association with willingness to participate in and share image data from orofacial genetic research, with baseline demographics and prior knowledge of orofacial research as covariates. Results: A total of 82 members of the Scottish public were recruited. The majority of respondents (83%) would participate in a research project that involves the collection of facial images. Of those who would participate, most (93%) would share their data outside of the UK. Fewer (3%) would only share their data within the UK. Education level and prior knowledge of the oral-systemic health relationship were associated with respondent’s willing ness to participate. Education level was also associated with respondent’s willingness to share image data. Conclusions: This preliminary study indicates a high level of willingness among the Scottish public to participate in and share data from orofacial genetic research. The findings suggest that education level and prior knowledge of oral-systemic health influence participation, highlighting the need for targeted public engagement in orofacial genetic research. These results lay the groundwork for larger, more diverse studies to further explore these associations.

Abstract Long-read single molecule sequencing technologies continue to grow in popularity for genome assembly and provide an effective way to resolve large and complex genomic variants. However, uptake of these technologies for teaching and training is hampered by the complexity of high molecular weight DNA extraction protocols, the time required for library preparation and the costs for sequencing, as well as challenges with downstream data analyses. Here, we present a full long-read workflow optimised for teaching, that covers each stage from DNA extraction, to library preparation and sequencing, to data QC and genome assembly and characterisation, that can be completed in under two weeks. We use a specific case study of plant identification, where students identify an anonymous plant sample by sequencing and assembling the genome and comparing it to other samples and to reference databases. In testing, long-read genome skimming of nine wild-collected plant species extracted with a modified kit-based approach produced an average of 8Gb of Oxford Nanopore data, enabling the complete assembly of plastid genomes, and partial assembly of nuclear genomes. In the classroom, all students were able to complete the protocols, and to correctly identify their plant samples based on BOLD searches of barcoding loci extracted from the plastid genome, coupled with phylogenetic analyses of whole plastid genomes. We supply all the learning material and raw data allowing this to be adapted to a range of teaching settings.

NHS genetics centres in Scotland sought to investigate the Genomics England 100,000 Genomes Project diagnostic utility to evaluate genome sequencing for in rare, inherited conditions. Four regional services recruited 999 individuals from 394 families in 200 rare phenotype categories, with negative historic genetic testing. Genome sequencing was performed at Edinburgh Genomics, and phenotype and sequence data were transferred to Genomics England for variant calling, gene-based filtering and variant prioritisation. NHS Scotland genetics laboratories performed interpretation, validation and reporting. New diagnoses were made in 23% cases – 19% in genes implicated in disease at the time of variant prioritisation, and 4% from later review of additional genes. Diagnostic yield varied considerably between phenotype categories and was minimal in cases with prior exome testing. Genome sequencing with gene panel filtering and reporting achieved improved diagnostic yield over previous historic testing but not over now routine trio-exome sequence tests. Re-interpretation of genomic data with updated gene panels modestly improved diagnostic yield at minimal cost. However, to justify the additional costs of genome vs exome sequencing, efficient methods for analysis of structural variation will be required and / or cost of genome analysis and storage will need to decrease.

Abstract Multiple viruses cause a phenomenon termed superinfection exclusion whereby a currently infected cell is resistant to secondary infection by the same or a closely related virus. In alphaviruses, this process is thought to be mediated, at least in part, by the viral protease (nsP2) which is responsible for processing the non-structural polyproteins (P123 and P1234) into individual proteins (nsP1-nsP4), forming the viral replication complex. Taking a synthetic-biology approach, we mimicked this naturally occurring phenomenon by generating a superinfection exclusion-like state in Aedes aegypti mosquitoes, rendering them refractory to alphavirus infection. By artificially expressing Sindbis virus (SINV) and chikungunya virus (CHIKV) nsP2 in mosquito cells and transgenic mosquitoes, we demonstrated a reduction in both SINV and CHIKV viral replication rates in cells following viral infection as well as reduced infection prevalence, viral titres and transmission potential in mosquitoes.

Cape Primroses (Streptocarpus, Gesneriaceae) are an ideal study system for investigating the genetics underlying species diversity in angiosperms. Streptocarpus rexii has served as a model species for plant developmental research for over five decades due to its unusual extended meristem activity present in the leaves. In this study, we sequenced and assembled the complete nuclear, chloroplast, and mitochondrial genomes of S. rexii using Oxford Nanopore Technologies long read sequencing. Two flow cells of PromethION sequencing resulted in 32 billion reads and were sufficient to generate a draft assembly including the chloroplast, mitochondrial and nuclear genomes, spanning 776 Mbp. The final nuclear genome assembly contained 5,855 contigs, spanning 766 Mbp of the 929‐Mbp haploid genome with an N50 of 3.7 Mbp and an L50 of 57 contigs. Over 70% of the draft genome was identified as repeats. A genome repeat library of Gesneriaceae was generated and used for genome annotation, with a total of 45,045 genes annotated in the S. rexii genome. Ks plots of the paranomes suggested a recent whole genome duplication event, shared between S. rexii and Primulina huaijiensis. A new chloroplast and mitochondrial genome assembly method, based on contig coverage and identification, was developed, and successfully used to assemble both organellar genomes of S. rexii . This method was developed into a pipeline and proved widely applicable. The nuclear genome of S. rexii and other datasets generated and reported here will be invaluable resources for further research to aid in the identification of genes involved in morphological variation underpinning plant diversification.