Aspergillus fumigatus is the most common and deadly pulmonary fungal infection worldwide. In the lung, the fungus usually forms a dense colony of filaments embedded in a polymeric extracellular ...matrix. To identify candidate genes involved in this biofilm (BF) growth, we used RNA-Seq to compare the transcriptomes of BF and liquid plankton (PL) growth. Sequencing and mapping of tens of millions sequence reads against the A. fumigatus transcriptome identified 3,728 differentially regulated genes in the two conditions. Although many of these genes, including the ones coding for transcription factors, stress response, the ribosome, and the translation machinery, likely reflect the different growth demands in the two conditions, our experiment also identified hundreds of candidate genes for the observed differences in morphology and pathobiology between BF and PL. We found an overrepresentation of upregulated genes in transport, secondary metabolism, and cell wall and surface functions. Furthermore, upregulated genes showed significant spatial structure across the A. fumigatus genome; they were more likely to occur in subtelomeric regions and colocalized in 27 genomic neighborhoods, many of which overlapped with known or candidate secondary metabolism gene clusters. We also identified 1,164 genes that were downregulated. This gene set was not spatially structured across the genome and was overrepresented in genes participating in primary metabolic functions, including carbon and amino acid metabolism. These results add valuable insight into the genetics of biofilm formation in A. fumigatus and other filamentous fungi and identify many relevant, in the context of biofilm biology, candidate genes for downstream functional experiments.
Next-generation sequencing has opened the door to genomic analysis of nonmodel organisms. Technologies generating long-sequence reads (200-400 bp) are increasingly used in evolutionary studies of ...nonmodel organisms, but the short-sequence reads (30-50 bp) that can be produced at lower cost are thought to be of limited utility for de novo sequencing applications. Here, we tested this assumption by short-read sequencing the transcriptomes of the tropical disease vectors Aedes aegypti and Anopheles gambiae, for which complete genome sequences are available. Comparison of our results to the reference genomes allowed us to accurately evaluate the quantity, quality, and functional and evolutionary information content of our "test" data. We produced more than 0.7 billion nucleotides of sequenced data per species that assembled into more than 21,000 test contigs larger than 100 bp per species and covered ∼27% of the Aedes reference transcriptome. Remarkably, the substitution error rate in the test contigs was ∼0.25% per site, with very few indels or assembly errors. Test contigs of both species were enriched for genes involved in energy production and protein synthesis and underrepresented in genes involved in transcription and differentiation. Ortholog prediction using the test contigs was accurate across hundreds of millions of years of evolution. Our results demonstrate the considerable utility of short-read transcriptome sequencing for genomic studies of nonmodel organisms and suggest an approach for assessing the information content of next-generation data for evolutionary studies.
Tuberculosis in the UK is more prevalent in people with social risk factors- e.g. previous incarceration, homelessness - and in migrants from TB endemic countries. The management of TB infection is ...part of TB elimination strategies, but is challenging to provide to socially excluded groups and the evidence base for effective interventions is small.
We evaluated a TB infection screening and treatment programme provided by a peer-led service (Find&Treat) working in inclusion health settings (e.g. homeless hostels) in London. IGRA (interferon-gamma release assay) testing and TB infection treatment were offered to eligible adults using a community-based model. The primary outcome was successful progression through the cascade of care. We also evaluated socio-demographic characteristics associated with a positive IGRA.
42/312 (13.5%) participants had a positive IGRA and no one had evidence of active TB. 35/42 completed a medical evaluation; 22 started treatment, and 17 completed treatment. Having a positive IGRA was associated with previous incarceration and being born outside of the UK.
Provision of TB infection diagnosis and management to this socially excluded population has several challenges including maintaining people in care and drug-drug interactions. Peer-support workers provided this service safely and effectively with appropriate support. Further work to generate data to inform risks and benefits of treatment for TB infection in this group is needed to facilitate joint decision making.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Introduction
COVID-19 was declared a pandemic in March 2020. With the sudden surge in demand for personal protective equipment (PPE), significant concerns regarding the ongoing availability emerged. ...One solution proposed is re-sterilisation of respirator masks and this has been commenced in some parts of the world. On review of the literature, very little is documented regarding the knowledge of masks and the attitudes of healthcare workers towards using re-sterilised masks.
Methods
A comprehensive questionnaire was used to assess general knowledge and attitudes around facemasks and respirators.
Results
There were 190 respondents. There were significant gaps in knowledge and understanding of when particular face masks should be worn. One-third had significant concerns about ongoing availability. One-third had concerns about the quality of the masks as the pandemic continued. Only 10% of respondents underwent formal face-fitting. Eighty percent of respondents stated they would wear a re-sterilised mask. A further 15% would use a re-sterilised mask but required certain reassurances. Five percent of our respondents would not use a re-sterilised mask under any circumstances.
Discussion
Ensuring an adequate understanding of face masks is crucial among healthcare workers (HCWs) and this study highlights a need for further education. It also demonstrates a general acceptability among HCWs towards the use of re-sterilised face masks.
Aspergillus fumigatus is both an environmental saprobe and an opportunistic human fungal pathogen. Knowledge of genomic variation across A. fumigatus isolates is essential for understanding the ...evolution of pathogenicity, virulence, and resistance to antifungal drugs. Here, we investigated 206 A. fumigatus isolates (133 clinical and 73 environmental isolates), aiming to identify genes with variable presence across isolates and test whether this variation was related to the clinical or environmental origin of isolates. The PanOrtho genome of A. fumigatus consists of 13,085 ortholog groups, of which 7,773 (59.4%) are shared by all isolates (core groups) and 5,312 (40.6%) vary in their gene presence across isolates (accessory groups plus singletons). Despite differences in the distribution of orthologs across all isolates, no significant differences were observed among clinical versus environmental isolates when phylogeny was accounted for. Orthologs that differ in their distribution across isolates tend to occur at low frequency and/or be restricted to specific isolates; thus, the degree of genomic conservation between orthologs of A. fumigatus is high. These results suggest that differences in the distribution of orthologs within A. fumigatus cannot be associated with the clinical or environmental origin of isolates.
Aspergillus fumigatus is a cosmopolitan species of fungus responsible for thousands of cases of invasive disease annually. Clinical and environmental isolates of A. fumigatus exhibit extensive phenotypic differences, including differences related to virulence and antifungal drug resistance. A comprehensive survey of the genomic diversity present in A. fumigatus and its relationship to the clinical or environmental origin of isolates can contribute to the prediction of the mechanisms of evolution and infection of the species. Our results suggest that there is no significant variation in ortholog distribution between clinical and environmental isolates when accounting for evolutionary history. The work supports the hypothesis that environmental and clinical isolates of A. fumigatus do not differ in their gene contents.
Downregulation of the alveolar macrophage (AM) receptor with collagenous structure (MARCO) leads to susceptibility to postinfluenza bacterial pneumonia, a major cause of morbidity and mortality. We ...sought to determine whether immunomodulation of MARCO could improve host defense and resistance to secondary bacterial pneumonia. RNAseq analysis identified a striking increase in MARCO expression between
and
after influenza infection and indicated important roles for Akt and Nrf2 in MARCO recovery. In vitro, primary human AM-like monocyte-derived macrophages (AM-MDMs) and THP-1 macrophages were treated with IFNγ to model influenza effects. Activators of Nrf2 (sulforaphane) or Akt (SC79) caused increased MARCO expression and a MARCO-dependent improvement in phagocytosis in IFNγ-treated cells and improved survival in mice with postinfluenza pneumococcal pneumonia. Transcription factor analysis also indicated a role for transcription factor E-box (TFEB) in MARCO recovery. Overexpression of TFEB in THP-1 cells led to marked increases in MARCO. The ability of Akt activation to increase MARCO expression in IFNγ-treated AM-MDMs was abrogated in TFEB-knockdown cells, indicating Akt increases MARCO expression through TFEB. Increasing MARCO expression by targeting Nrf2 signaling or the Akt-TFEB-MARCO pathway are promising strategies to improve bacterial clearance and survival in postinfluenza bacterial pneumonia.
Sesame
L. is a major oil-based seed crop that has been widely cultivated and consumed in Pakistan. Unfortunately, sesame is highly prone to
fungal growth in the field, and under inappropriate storage ...conditions can become contaminated with aflatoxins, the most potent carcinogen found in nature. Here, we have isolated a high number of
isolates from sesame seeds in fresh and stored conditions obtained from rainfed and irrigated zones of Punjab, Pakistan, and characterized them for aflatoxigenic potentials. Using morphological identification techniques, 260 isolates were grouped as potential
section Flavi, with 126 and 134 originating from the rainfed and irrigated zones, respectively. Out of 260 in total, 188 isolates were confirmed to produce aflatoxins. There were no significant differences in potential aflatoxigenic isolates with respect to the rainfed and irrigated zones. However, the number of potential aflatoxigenic isolates was significantly higher (
< 0.05) in stored samples than that of those from fresh sesame seeds in the rainfed and irrigated zone. Whole genome sequencing and comparative analyses of 12 select isolates have revealed that one of the
isolates, which produced very low aflatoxins (AFP10), has an elevated missense variant rate, numerous high impact mutations, and a 600 base pair deletion in the
gene. In summary, our study provides insights into aflatoxigenic potential and the associated genetic diversity of indigenous
section Flavi isolates and potential management strategies for reducing aflatoxin contamination levels in a major crop consumed in Punjab, Pakistan.
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A protocol is presented for the calculation of monitor units (MU) for photon and electron beams, delivered with and without beam modifiers, for constant source-surface distance (SSD) and source-axis ...distance (SAD) setups. This protocol was written by Task Group 71 of the Therapy Physics Committee of the American Association of Physicists in Medicine (AAPM) and has been formally approved by the AAPM for clinical use. The protocol defines the nomenclature for the dosimetric quantities used in these calculations, along with instructions for their determination and measurement. Calculations are made using the dose per MU under normalization conditions,
$D_0^\prime $
D
0
′
, that is determined for each user's photon and electron beams. For electron beams, the depth of normalization is taken to be the depth of maximum dose along the central axis for the same field incident on a water phantom at the same SSD, where
$D_0^\prime $
D
0
′
= 1 cGy/MU. For photon beams, this task group recommends that a normalization depth of 10 cm be selected, where an energy-dependent
$D_0^\prime $
D
0
′
≤ 1 cGy/MU is required. This recommendation differs from the more common approach of a normalization depth of d
m
, with
$D_0^\prime $
D
0
′
= 1 cGy/MU, although both systems are acceptable within the current protocol. For photon beams, the formalism includes the use of blocked fields, physical or dynamic wedges, and (static) multileaf collimation. No formalism is provided for intensity modulated radiation therapy calculations, although some general considerations and a review of current calculation techniques are included. For electron beams, the formalism provides for calculations at the standard and extended SSDs using either an effective SSD or an air-gap correction factor. Example tables and problems are included to illustrate the basic concepts within the presented formalism.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
The freshwater pearl mussel Margaritifera margaritifera is an ecologically important globally endangered species, yet little is known about its biodiversity and population genetics in North American ...populations. This study focused on M. margaritifera from six locations within two stream systems in central and eastern Massachusetts, USA, to understand the historical impact of damming and habitat fragmentation on local population structure and genetic diversity.
Approximately 300 informative single nucleotide polymorphisms were generated from 59 individuals across six geographical locations, using the restriction‐site‐associated DNA sequencing approach. Genotypes were also gleaned from publicly available RNA sequencing data of 23 French M. margaritifera samples.
Predictions of population structure using discriminant analysis of principal components and fastStructure both indicate the existence of a single genetic population in central and eastern Massachusetts, and a clear separation from French individuals. This observation is further supported by frequency‐based calculations of population genetics, which indicate near‐complete admixture and a high number of migrants between Massachusetts sites.
These results suggest that the effects of habitat fragmentation from damming in the past century have not radiated through the Massachusetts population of M. margaritifera, potentially because of the long lifespan and generation time of the species. However, the Massachusetts population has similar levels of genetic diversity to the endangered populations in Europe, and these factors should be considered by conservationists when creating conservation and management plans.
The findings of this study suggest that populations in North America are reasonably healthy and diverse and that, as one population, the study area represents one conservation unit. This indicates that dam removal, habitat restoration, and captive breeding programmes may be viable options to increase the small population numbers and retain a substantial portion of genetic diversity without sacrificing local adaptation.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
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