Biology subjects

Douglas, R. G.

Publications and source records attributed to Douglas, R. G..

7 recordsLinked to original sources

bioRxiv2026

Actin-related protein Alp1 governs malaria parasite motility and transmission

Motility of the malaria-causing parasite Plasmodium is essential for transmission to and from mosquitoes, with the turnover of actin filaments being a central feature of productive cell movement. Actin-related proteins (Arps) are known to play critical roles in motility, trafficking and chromatin remodelling. Here, we show that the actin-like protein 1 (Alp1), an apicomplexan Arp, is essential for Plasmodium ookinete motility and establishing of infection in mosquitoes. We identified an insertion region in subdomain 4 that contributes to Alp1 function in ookinetes and show novel actin filament structures in ookinetes. A combination of gene deletion and actin filament recognizing chromobody expression revealed a role of Alp1 in promoting actin filament turnover in ookinetes. We have thus identified a novel Arp that has evolved a specialist function to regulate actin dynamics, govern malaria parasite motility and facilitate malaria transmission.

cell biology↗

bioRxiv2026

Plasmodium actin-like proteins 3 and 5a are essential for subsequent steps of mosquito infection

Actin superfamily members are critical for the biology of eukaryotes and archaea. Actin-related proteins (Arps) are a subgroup within the actin superfamily and play essential roles in trafficking, replication and motility. The genome of the malaria parasite Plasmodium contains a set of Arps unique to apicomplexans, termed actin-like proteins (Alps). However, the importance and specific roles of many of these Alps in Plasmodium progression are not yet understood. Here, we determined the functional contribution of Plasmodium berghei Alp3 and Alp5a (recently relabelled as Arp3) by generation of knock-out (KO) lines and their subsequent characterisation across different life cycle stages. Deletion of either Alp did not affect blood stage growth, gametogenesis and ookinete gliding motility. However, deletion of Alp5a lead to smaller and fewer oocysts as well as severely impaired sporozoite formation. The Alp3KO line had highly reduced oocyst loads compared to wild-type parasites. This striking decrease was due to impaired ookinete penetration of the mosquito midgut epithelium. Our study shows that both Alp3 and Alp5a are indispensable for Plasmodium transmission at different steps of initial mosquito infection, provides insights into the role of specific unique members of the actin superfamily during parasite progression and the requirements for efficient midgut penetration.

molecular biology↗

bioRxiv2024

Molecular architecture of glideosome and nuclear F-actin in Plasmodium falciparum

Actin-based motility is required for the transmission of malaria sporozoites. While this has been shown biochemically, filamentous actin has remained elusive and has to date never been directly visualised inside the parasite. Using focused ion beam milling and electron cryo-tomography, we studied dynamic actin filaments in unperturbed Plasmodium falciparum cells for the first time. This allowed us to dissect the assembly, path and fate of actin filaments during parasite gliding and determine a complete 3D model of F-actin within sporozoites. We show that within the cell, actin assembles into micrometre long filaments, much longer than observed in in vitro studies. After their assembly at the parasites apical end, actin filaments continue to grow as they are transported down the cell as part of the glideosome machinery, and are disassembled at the basal end in a rate-limiting step. Large pores in the IMC, constrained to the basal end, may facilitate actin exchange between the pellicular space and the cytosol for its recycling and maintenance of directional actin flow for efficient gliding. The data also reveal striking and extensive actin bundles in the nucleus. Implications of these structures for motility and transmission are discussed.

microbiology↗

bioRxiv2020

Antibiotic concentrations in the sinonasal secretions and tissue in CRS patients after oral therapy: a randomized trial

Background Despite the widespread prescription of antibiotics for patients with chronic rhinosinusitis (CRS), the extent to which drug distribution to the sinonasal mucosa influences their efficacy remains largely undefined.Methods Thirty subjects undergoing functional endoscopic sinus surgery (FESS) for bilateral CRS were randomized to one of three groups: 1) doxycycline (100 mg daily for seven days) 2) roxithromycin (300 mg daily for seven days) and 3) control (no antibiotics given). Drug levels were measured using liquid chromatography-tandem mass spectrometry in sinonasal secretions, sinonasal tissues and serum at steady state. Nasal endoscopy (Modified Lund-Kennedy) and Gastrointestinal Symptom Rating Scale (GSRS) scores were recorded.Results Antibiotic concentrations in the nasal secretions were significantly lower compared to those in the serum and tissue (mean mucus/serum ratio at steady state = 0.16 and 0.37 for doxycycline and roxithromycin respectively; p<0.01). A short course of antibiotic intake did not correlate with any difference in clinical outcomes except where slightly higher GSRS scores were reported in the roxithromycin group (p=0.04).Conclusions Although the efficacy of doxycycline and roxithromycin in sinonasal mucus in vivo cannot be predicted solely from reported minimum inhibitory concentrations, given the added complication of bacterial biofilm antimicrobial tolerance, these results suggest that low mucosal penetration of antibiotics may be one of the factors contributing to the limited efficacy of these agents in the treatment of CRS.View Full Text

pharmacology and toxicology↗

bioRxiv2019

Microbiotyping the sinonasal microbiome

This study offers a novel description of the sinonasal microbiome, through an unsupervised machine learning approach combining dimensionality reduction and clustering. We apply our method to the International Sinonasal Microbiome Study (ISMS) dataset of 410 sinus swab samples. We propose three main sinonasal microbiotypes or states: the first is Corynebacterium-dominated, the second is Staphylococcus-dominated, and the third dominated by the other core genera of the sinonasal microbiome (Streptococcus, Haemophilus, Moraxella, and Pseudomonas). The prevalence of the three microbiotypes studied did not differ between healthy and diseased sinuses, but differences in their distribution were evident based on geography. We also describe a potential reciprocal relationship between Corynebacterium species and Staphylococcus aureus, suggesting that a certain microbial equilibrium between various players is reached in the sinuses. We validate our approach by applying it to a separate 16S rRNA gene sequence dataset of 97 sinus swabs from a different patient cohort. Sinonasal microbiotyping may prove useful in reducing the complexity of describing sinonasal microbiota. It may drive future studies aimed at modeling microbial interactions in the sinuses and in doing so may facilitate the development of a tailored patient-specific approach to the treatment of sinus disease in the future.

microbiology↗

bioRxiv2019

The international sinonasal microbiome study (ISMS): a multi centre, international characterization of sinonasal bacterial ecology

The sinonasal microbiome remains poorly defined, with our current knowledge based on a few cohort studies whose findings are inconsistent. Furthermore, the variability of the sinus microbiome across geographical divides remains unexplored. We characterise the sinonasal microbiome and its geographical variations in both health and disease using 16S rRNA gene sequencing of 410 individuals from across the world. Although the sinus microbial ecology is highly variable between individuals, we identify a core microbiome comprised of Corynebacterium, Staphylococcus, Streptococcus, Haemophilus, and Moraxella species in both healthy and chronic rhinosinusitis (CRS) cohorts. Corynebacterium (mean relative abundance = 44.02%) and Staphylococcus (mean relative abundance = 27.34%) appear particularly dominant in the majority of patients sampled. There was a significant variation in microbial diversity between countries (p = 0.001). Amongst patients suffering from CRS with nasal polyps, a significant depletion of Corynebacterium (40.29% vs 50.43%; p = 0.02) and over-representation of Streptococcus (7.21% vs 2.73%; p = 0.032) was identified. The delineation of the sinonasal microbiome and standardised methodology described within our study will enable further characterisation and translational application of the sinus microbiota.

microbiology↗

bioRxiv2018

Comparison of subtyping approaches and the underlying drivers of microbial signatures for chronic rhinosinusitis

Chronic rhinosinusitis (CRS) is a heterogeneous condition characterised by persistent sinus inflammation and microbial dysbiosis. This study aimed to identify clinically relevant sub-groups of CRS patients based on distinct microbial signatures, with a comparison to the commonly used phenotypic subgrouping approach. The underlying drivers of these distinct microbial clusters were also investigated, together with associations with epithelial barrier integrity.\n\nSinus biopsies were collected from CRS patients (n=23), and disease controls (n=8). Expression of 42 tight junction genes was evaluated using quantitative PCR, together with microbiota analysis and immunohistochemistry for measuring mucosal integrity and inflammation.\n\nCRS patients clustered into two distinct microbial sub-groups using probabilistic modelling Dirichlet (DC) multinomial mixtures. DC1 exhibited significantly reduced bacterial diversity, increased dispersion, and was dominated by Pseudomonas, Haemophilus, and Achromobacter. DC2 had significantly elevated B-cells, incidence of nasal polyps, and higher numbers of Anaerococcus, Megasphaera, Prevotella, Atopobium, and Propionibacterium. In addition, each DC exhibited distinct tight junction gene and protein expression profiles compared with controls. Stratifying CRS patients based on clinical phenotypic subtypes (absence or presence of nasal polyps (CRSsNP or CRSwNP respectively) or with cystic fibrosis (CRSwCF)) did account for a larger proportion of the variation in the microbial dataset compared with DC groupings. However, no significant differences between CRSsNP and CRSwNP cohorts were observed for inflammatory markers, beta-dispersion and alpha diversity measures.\n\nIn conclusion, both approaches used for stratifying CRS patients had benefits and pitfalls, but DC clustering did provide greater resolution when studying tight junction impairment. Future studies in CRS should give careful consideration into the patient subtyping approach used.\n\nImportanceChronic rhinosinusitis (CRS) is a major human health problem that significantly reduces quality of life. While various microbes have been implicated, there is no clear understanding of the role they play in CRS pathogenesis. Another equally important observation made for CRS patients is that the epithelial barrier in the sino-nasal cavity is defective. Finding a robust approach to subtype CRS patients would be the first step towards unravelling the pathogenesis of this heterogeneous condition. Previous work has explored stratification based on clinical presentation of the disease (with or without polyps), inflammatory markers, pathology, or microbial composition. Comparing between the different stratification approaches used in these studies has not been possible due to different cohorts, analytical methods, or sample sites used. In this study, two approaches of subtyping CRS patients were compared and the underlying drivers of the heterogeneity in CRS were also explored.

microbiology↗