Directional Next-Generation RNA Sequencing and Examination of Premature Termination Codon Mutations in Endoglin/Hereditary Haemorrhagic Telangiectasia

Hereditary haemorrhagic telangiectasia (HHT) is a disease characterised by abnormal vascular structures, and most commonly caused by mutations in ENG, ACVRL1 or SMAD4 encoding endothelial cell-expressed proteins involved in TGF-β superfamily signalling. The majority of mutations reported on the HHT mutation database are predicted to lead to stop codons, either due to frameshifts or direct nonsense substitutions. The proportion is higher for ENG (67%) and SMAD4 (65%) than for ACVRL1 (42%), p < 0.0001. Here, by focussing on ENG, we report why conventional views of these mutations may need to be revised. Of the 111 stop codon-generating ENG mutations, on ExPASy translation, all except one were premature termination codons (PTCs), sited at least 50-55 bp upstream of the final exon-exon boundary of the main endoglin isoform, L-endoglin. This strongly suggests that the mutated RNA species will undergo nonsense-mediated decay. We provide new in vitro expression data to support dominant negative activity of stable truncated endoglin proteins but suggest these will not generate HHT: the single natural stop codon mutation in L-endoglin (sited within 50-55 nucleotides of the final exon-exon boundary) is unlikely to generate functional protein since it replaces the entire transmembrane domain, as would 8 further natural stop codon mutations, if the minor S-endoglin isoform were implicated in HHT pathogenesis. Finally, next-generation RNA sequencing data of 7 different RNA libraries from primary human endothelial cells demonstrate that multiple intronic regions of ENG are transcribed. The potential consequences of heterozygous deletions or duplications of such regions are discussed. These data support the haploinsufficiency model for HHT pathogenesis, explain why final exon mutations have not been detected to date in HHT, emphasise the potential need for functional examination of non-PTC-generating mutations, and lead to proposals for an alternate stratification system of mutational types for HHT genotype-phenotype correlations

Activity and social interactions in a wideranging specialist scavenger, the Tasmanian devil (Sarcophilus harrisii), revealed by animalborne video collars

Observing animals directly in the field provides the most accurate understanding of animal behaviour and resource selection. However, making prolonged observation of undisturbed animals is difficult or impossible for many species. To overcome this problem for the Tasmanian devil (Sarcophilus harrisii), a cryptic and nocturnal carnivore, we developed animal-borne video collars to investigate activity patterns, foraging behaviour and social interactions. We collected 173 hours of footage from 13 individual devils between 2013 and 2017. Devils were active mostly at night, and resting was the most common behaviour in all diel periods. Devils spent more time scavenging than hunting and exhibited opportunistic and flexible foraging behaviours. Scavenging occurred mostly in natural vegetation but also in anthropogenic vegetation and linear features (roads and fence lines). Scavenging frequency was inversely incremental with size e.g. small carcasses were scavenged most frequently. Agonistic interactions with conspecifics occurred most often when devils were traveling but also occurred over carcasses or dens. Interactions generally involved vocalisations and brief chases without physical contact. Our results highlight the importance of devils as a scavenger in the Tasmanian ecosystem, not just of large carcasses for which devils are well known but in cleaning up small items of carrion in the bush. Our results also show the complex nature of intraspecific interactions, revealing greater detail on the context in which interactions occur. In addition, this study demonstrates the benefits of using animal-borne imaging in quantifying behaviour of elusive, nocturnal carnivores not previously seen using conventional field methods

HIV knowledge, disclosure and sexual risk among pregnant women and their partners in rural South Africa

Partner involvement has been deemed fundamental for the prevention of mother-to-child transmission (PMTCT) of HIV, although it remains difficult to achieve. This study aimed to explore the attitudes and behaviours of pregnant women and their partners who participated in a behavioural risk reduction intervention in six community health centres in the Mpumalanga province of South Africa. Qualitative methods only were used in this study. Women and their partners took part in four gender-concordant groups that addressed HIV, PMTCT, disclosure of HIV status and safer sex practices. The results indicate that men value and understand the importance of being involved in women’s reproductive health, although some components of the PMTCT programme such as condom use were still met with some resistance. Participants demonstrated high levels of HIV- and sexually transmitted infection-related knowledge. Men lacked knowledge about PMTCT but were interested in acquiring information so that they could support their partners. All groups highlighted the emotional and physical benefits of disclosing one’s HIV status. The involvement of men in antenatal care has the potential to prevent women from becoming infected with HIV both during pregnancy and post-partum when they are more vulnerable to infection and have a high risk of transmission to the infant. There is a need for interventions that focus on both increasing male involvement and promoting condom use during pregnancy.Keywords: HIV, PMTCT, disclosure, partners, risk reduction intervention, Mpumalang