bluuebunny/crossref_metadata_2025 huggingface dataset:
The power of odour cues in shaping fine-scale search patterns of foraging mammalian herbivores
Authors: Orlando Cristian Gabriel , Tews Ashley , Banks Peter , McArthur Clare
Published: July 2020

Foraging by mammalian herbivores has profound impacts on natural and modified landscapes, yet we know little about how they find food, limiting our ability to predict and manage their influence. Mathematical models show that foragers exploiting odour cues outperform a random walk strategy. However, discovering how free-ranging foragers exploit odours in real, complex landscapes has proven elusive because of technological constraints. We took a novel approach, using a sophisticated purpose-built thermal camera system to record fine-scale foraging by a generalist mammalian herbivore, the swamp wallaby (
Wallabia bicolor
). We tested the hypothesis that odour cues shape forager movement and behaviour in vegetation patches. To do this, we compared wallaby foraging in two odour landscapes:
Control
(natural vegetation with food and non-food plants interspersed) and
+Apple
(the same natural vegetation plus a single, highly palatable food source with novel odour (apple)). The
+Apple
treatment led to strongly directed foraging by wallabies: earlier visits to vegetation patches, straighter movement paths, more hopping and fewer stops than in the
Control
treatment. Our results provide clear empirical evidence that odour cues are harnessed for efficient, directed search even at this fine scale. We conclude that random walk models miss a key feature shaping foraging within patches.
Exploring the attachment of the Mediterranean medicinal leech (Hirudo verbana) to porous substrates
Authors: Kampowski Tim , Thiemann Lara-Louise , Kürner Lukas , Speck Thomas , Poppinga Simon
Published: July 2020

Haematophagous ectoparasites must ensure a reliable hold to their host during blood meals and, therefore, have evolved a broad spectrum of versatile and effective attachment mechanisms. The Mediterranean medicinal leech (Hirudo verbana), for example, uses suction on both smooth and textured air-tight substrates. However, preliminary studies showed thatH. verbanais also capable of attaching itself to air-permeable substrates, where suction does not work. Using high-speed videography and mechanical tests, we comparatively investigated the attachment ofH. verbanaon both smooth and textured air-tight as well as on porous artificial substrates, also considering the influence of mucus on sucker surfaces. In general, the leech-specific locomotion cycle did not differ between the tested surfaces, and the leeches were able to reliably attach to both air-tight and porous substrates. From our results, we conclude that suction is presumably the primary attachment mechanism ofH. verbana. However, secondary mechanisms such as mechanical interlocking with surface asperities and pores or capillary forces occurring at the interface between the mucus-covered suckers and the substratum are also employed. In any case, the rich repertoire of applicable attachment principles renders the organs ofH. verbanafunctionally highly resilient.
The use of wound dressings as a means to alter the composition of biofilms in a chronic wound model
Authors: Nedelea Gabriela , Maddocks Sarah
Published: July 2020

It is estimated that £5 billion are invested yearly into chronic wound management by the NHS. Whilst the demand for treatment rises every year, it has become harder to treat wounds given the burden of antimicrobial resistance. Chronic wounds can easily become harbouring grounds for polymicrobial biofilms in which species interact in specific ways.

This study assessed the interactions between two commonly co-isolated chronic wound pathogens: Pseudomonas aeruginosa (ATCC 9027) and Staphylococcus aureus (EMRSA 15), whose biofilm relation initiates a Gram-negative shift. During this phenomenon, P. aeruginosa takes over the majority of the bacterial community, at the detriment of S. aureus. The Gram-negative shift marks the turning point from an acute to a chronic wound. The pH of a chronic wound is typically alkaline, and it was hypothesised that topical dressings with an acidic pH could disrupt the onset of the Gram negative shift, and therefore chronicity. Six different topical dressings with low pH were used in achronic wound model to assess their ability to reverse or delay the Gram-negative shift. It was found that they did not have an impact on the onset of the Gram-negative shift, despite their low pH values. However, the lower the pH of the dressings, the more frequently small colony variant (SCV) bacteria were observed in the biofilm. SCVs are known for causing persistent or chronic infections. It was therefore concluded that low pH dressings alone may not be favourable for managing chronic wound infection.
The SKI complex is a broad-spectrum antiviral drug target
Authors: Weston Stuart , Keller Chloe , MacKerell Alexander , Frieman Matthew
Published: July 2020

Starting from a yeast suppressor screening platform, we have identified the SKI complex as a potential broad-spectrum antiviral target. We found that the NS1 protein of influenza A virus (IAV) and the ORF4a protein of Middle East respiratory syndrome coronavirus (MERS-CoV), which both function to bind double-strand RNA and inhibit cellular interferon responses, cause a slow growth phenotype when expressed in yeast. Knockout of the components of the yeast SKI complex caused a loss of this slow growth phenotype, suggesting a functional link between the viral proteins and the SKI complex. The SKI complex is a helicase that unwinds double-strand RNA and sends it to the RNA exosome for degradation. We next investigated whether the highly conserved human SKI complex was important for replication of IAV and MERS-CoV. RNAi based experiments showed that both viruses were inhibited when the SKI complex was removed, suggesting the complex has a proviral role in replication. Through in silico modelling using the published crystal structure of the SKI complex, we looked for potential binding pockets for chemical compounds. We screened a selection of these compounds for antiviral activity and have found four different chemicals capable of inhibiting IAV infection. Our most studied of these also inhibits not only MERS-CoV, but also Ebolavirus Makona. Our data suggests the SKI complex may be a target for broad-spectrum antiviral therapy and we have multiple chemical structures from which to work to develop therapeutic approaches.
Digital Outreach: Promoting a global understanding of antimicrobial resistance (AMR)
Authors: Shaw Michael
Published: July 2020

Technology allows educators to reach a global audience without amassing a large carbon footprint. The Skype in the Classroom programme from Microsoft enables me to spark curiosity and engage a wide range of students in dialogue to explore global issues, in particular, the threat of antimicrobial resistance, while supporting me to develop my skills as an educator. Through a multi-disciplinary approach, I employ Skype in the Classroom as a future-proof, powerful tool in our arsenal against the evolving microbial threat. We are not only responsible for teaching the next generation of scientists, but also the next generation of thought-leaders and professionals, who will work collaboratively towards our common goals.

Antimicrobial resistance is one of the grand challenges of the 21st century, and global connectivity is one of the best solutions to promoting better understanding of this threat. The Skype in the Classroom programme has allowed me to teach sessions to students in Norway, India, Brazil, Miami, Florida and Puerto Rico, with students ranging from 10 to 18 years old. I use skype and Teams to deliver online lessons in an accessible way, starting with the scope of the problem, covering the technical detail of how resistance emerges and is spread, and looking at some of the newest approaches to tackling AMR. Using a ‘challenge-based-learning’ approach, we discuss how different professionals and roles within society can contribute to the issue. This makes the class applicable to students with a wide range of backgrounds, including those without a keen interest in science.
Bacterial profile of meconium of neonates born at National Referral Hospital Cipto Mangunkusumo, Jakarta, Indonesia
Authors: Malik Amarila , Ong Grasella , Amanah Prasetyaningsih Firsty , Juni Kristien , Amandito Radhian , Rohsiswatmo Rinawati
Published: July 2020

Microbial colonization of a neonate’s gastrointestinal tract has significant perinatal and lifetime health consequences, with some clinical outcomes that have been linked to differences in the diversity and composition of gut microbiota. The effort to engineer intestinal ecosystem has led us to preserve the cultivable commensal microbiota. Here we investigated the association of cultivable bacterial diversity of neonates meconium from Indonesian National Referral Hospital Cipto Mangunkusumo (NRHCM) with mode of delivery and feeding patterns, as well as hyperbilirubinemia. We performed a cross-sectional study of meconium and clinical data collected from 14 Indonesian neonates born at NRHCM. Culture-dependent identification of bacterial isolates was conducted by performing simultaneous microbiological and molecular 16S rDNA PCR-Sanger sequencing methods. Phylogenetic tree and principal components analysis were employed to determine the bacterial profile and their association with clinical characteristics and outcomes. Cultivable bacterial profile indicates the predominance of Firmicutes (84,41%), with an abundant population of Staphylococcus (53.24%) with top three most significant population present are, i.e. S. hominis (12.99%), S. epidermidis (11.68%), and S. haemolyticus (10.39%). Bacterial diversity was associated with mode of delivery which showed that vaginal route populated by lower diversity of cultivable bacteria but by fewer opportunistics one than that of cesarean, with Staphylococcus hominis dominates the population, whereas with feeding patterns showed that the exclusive breast-fed was most populated by Staphylococcus, whereas non-exclusive one shared the same proportion of Staphylococcus and Bacterioides. While, non-hyperbilirubinemia group showed more abundant and diverse Staphylococcus than that of the opposite group.
Comparison of proteins expressed during urinary tract infection in young females
Authors: Alsubhi Manal , Alsharif Ghadah
Published: July 2020

Introduction: Gram-negative bacteria are a major cause of urinary tract infections (UTIs) and particularly Klebsiella pneumoniae (K. pneumoniae), which is a causative agent of 60-70% of community-acquired infections, about 30% of nosocomial UTIs and 20% of recurrent infections.

Materials and methods: Nine urine samples were collected from patients from various clinical departments in King Abdulaziz University Hospital from 2/3/2019 to 2/4/2019. The microbial contents in the urine samples was analysed by urine culture and VITEK analyses. Here, we compared K. pneumoniae proteins profiles to find possible proteins which could shed a light on host-pathogen interactions. The colonies were suspended in a lysing buffer, which then were sonicated, and the proteins contents were separated using 1D SDS-PAGE, analyzed using liquid chromatography-mass spectrometry LC/MS. Proteins showing different expressions in samples were identified by TripleTOF 5600 mass spectrometer.

Results: All of the Klebsiella pneumoniae isolates were ESBL+ and KPC+ as shown on ChromAgar plates. There was no available data on resistance, ESBL or KPC from VITEK2. Hence, ESBL+ or KPC+ data were only obtained from ChromAgar. Additionally, proteomics analysis revealed the following, the total number of different proteins that are expressed from all of the isolates is 2958 proteins. Where in sample U-102, 328 different proteins expressed, 300 different proteins expressed from isolate U-871, 350 different proteins expressed from isolate U-713, in isolate U-755, 378 different proteins were expressed, 207 different proteins expressed from isolate U-754, 305 different proteins expressed from sample U-134, 290 different proteins expressed from isolate U-968, 600 different proteins expressed from isolate U-104, and 200 different proteins expressed from isolate U-659.

Conclusion: The different proteins between the UTI patients indicated specific host pathogen interaction, each isolate expressed different proteins than the other isolate could be reasoned by host pathogen interaction. Host pathogen interaction are influenced by numbers of factor: age, gender, immunity, underling health conditions, antibiotic treatment, acute UTI or recurrent infection all of these factor could have an impact on the type of proteins that are expressed during infection. Even though the isolates are Klebsiella pneumoniae from young females with UTI, they expressed different proteins, these proteins could explain evolutionary development of pathogens and survival in urinary niche, as pathogens need to express certain type of proteins to enable them to live and survival in urinary niche.
The investigation of microbial induced calcium carbonate precipitation for soil improvement
Authors: Haystead Jamie , Dade-Robertson Martyn , Arnardottir Thora , Christgen Beate , Zhang Meng
Published: July 2020

Weak and unstable soils can limit the building of new infrastructure. Current soil strengthening techniques such as chemical grouting have detrimental effects on the environment from greenhouse gas production, soil pH modification and groundwater contamination. Microbial-induced calcium carbonate precipitation (MICCP) is a technique that utilises the ability of bacteria to precipitate calcium carbonate, which can be used for a variety of applications including binding adjacent soil particles and filling the pore spaces of soils to increase their mechanical properties. A commonly used bacterium is Sporosarcina pasteurii. A range of factors influences MICCP which presents challenges with process optimisation. Some studies have made use of computational models to predict biocementation at a larger scale, however aspects of models are based on assumption of conditions instead of experimental data.

An aim of this project is to investigate urease activity in S. pasteurii by comparing different growth media, growth stages, pH and temperatures. Ureolysis kinetics of S. pasteurii will be investigated at different urea and calcium chloride concentrations in liquid media. Finally, the biocementation of S. pasteurii in sand syringe setups will also be investigated to compare the effects of changing influencing factors such as growth stage and cell concentration of S. pasteurii, sand particle size, cementation media concentration, duration between cementation media applications and overall number of cementation treatments. Experimental work will be particularly focused towards gaps in the experimental data used in computational models, to help improve these models and bring MICCP biocementation closer to commercial use.
Genotypic and phenotypic characterisation of the Klebsiella oxytoca complex
Authors: McCuaig Frazer , Winter Jody , Thomas Jonathan , McVicker Gareth , Hoyles Lesley
Published: July 2020

Klebsiella spp. are associated with 3 to 7% of nosocomial infections and can be responsible for a range of conditions including pneumonia, bloodstream infections, meningitis, and necrotizing enterocolitis in infants. The role of Klebsiella pneumoniae in causing disease is well-characterised but, to date, the closely related species Klebsiella oxytoca has not received the same attention, despite often encoding extended-spectrum beta-lactamases and carbapenemases in clinical settings. K. oxytoca is the causative agent of Clostridiodes difficile-negative antibiotic-associated haemorrhagic colitis, a rare condition seen in some individuals receiving antibiotics. Whole-genome sequence analyses have shown K. oxytoca to be a complex comprising at least six species (K. oxytoca, K. michiganensis, K. grimontii, K. huaxiensis, ‘K. pasteurii’, ‘K. spallanzanii’). Our study aims to better characterise the K. oxytoca complex using a polyphasic approach. Preliminary investigations into the genomes of three K. michiganensis clinical isolates revealed the presence of a plasmid-borne ccdABlocus. ccdAB is a toxin-antitoxin (TA) system known to maintain plasmids in other pathogenic enterobacteria. We aim to functionally validate this TA system by cloning and conducting toxicity assays on the CcdB toxin, and cloning and assessing the ability of CcdA to function as an antidote. We also aim to sequence and generate Illumina/Oxford Nanopore hybrid genome assemblies of a larger collection of K. oxytoca complex clinical isolates and investigate their plasmids and TA systems in the same manner.
The role of air pollution and bacteria in COPD
Authors: Faye Purser Lillie , Purves Jo , Corscadden Louise , Selley Liza , S. Monks Paul , M. Ketley Julian , W. Andrew Peter , Barer Michael , Brightling Christopher , A. Morrissey Julie
Published: July 2020

Air pollution is the single largest environmental health risk worldwide. Particulate matter (PM) air pollution is released as a result of fossil fuel combustion and vehicle motion, breaking and tyre wear. It has been shown that exposure to PM can cause increased levels of respiratory disease, including the exacerbation of COPD, which is frequently associated with bacterial infection. Despite this, the effects of air pollution exposure on COPD associated respiratory bacteria, includingHaemophilus influenzae, Moraxella catarrhalis and Streptococcus pneumoniae are largely unknown. Our recent publication was the first to document that as well as damaging the host, PM has a direct impact on bacteria that can cause respiratory infections. We showed that exposure to black carbon (BC), an important component of PM, results in alterations in biofilm structure in both Streptococcus pneumoniae and Staphylococcus aureus, and increases dissemination of colonising S. pneumoniaein in vivo models.

Following on from this work, we aim to determine how BC impacts the growth, behaviour and virulence of bacteria associated with the COPD exacerbation, including non-typeable Haemophilus influenzaeand Moraxella catarrhalis. Current data show that BC exposure is decreasing the biofilm forming ability of NTHistrains 162 and 375. M. catarrhalis strain M61 biofilm formation is also decreased in the presence of BC, while its growth rate is increased. In addition, pre-exposing NTHi375 cells to BC, prior to infection of A549 cells, increases their ability to adhere to human epithelial cells. This suggests that the frequency of bacterial infection induced COPD exacerbation may be altered in patients from highly polluted areas.
Intercorrelation mechanism of Hypoxia and Cordycepin Biosynthesis in Zombie fungus
Authors: Suparmin Ahmad , Kato Tatsuya , Takemoto Hiroyuki , Y. Park Enoch
Published: July 2020

Cordycepin is an anticancer metabolite produces by a zombie fungus species of Cordyceps militaris. They are capable to infect and hijack insect’s nervous neuron system. Hypoxic environment commonly must be faced by the pathogenic fungus during infection either this zombie fungus. They activate oxygen sensing mode, heme, siderophore, and sterol biosynthesis to overcome it. Underlined our previous study that liquid surfaced culture of C. militaris NBRC103752 produced a higher amount of cordycepin than submerged culture, suggesting that hypoxic conditions might induce it. However, when and how the mechanism of cordycepin production started in liquid surfaced culture is not understood, yet. In our present study, the combination of transcriptomics and gas chromatography-mass spectrometry were carried out during the production phases of cordycepin (5d, 12d, and 19d of incubation periods) and the mechanism of cordycepin production was figured out. The expression of genes in the fermentation pathway and the oxidative phosphorylation pathway were significantly upregulated and down regulated, respectively. Expression of four genes in the heme biosynthesis, including 5-aminolevulinic acid synthase (CCM_01504), delta-aminolevulinic acid dehydratase (CCM_00935), coproporphyrinogen III oxidase (CCM_07483) and cytochrome c oxidase15 (CCM_05057) were upregulated at the beginning of the exponential phase (12d). Further, the activation of Zn(2)-C6 transcription factor that regulates the iron acquisition and ergosterol biosynthesis significantly upregulated and a metabolite reporter adenosine was detected only at 12d. The results in the present study show the correlation between hypoxia and the accumulation of heme before cordycepin biosynthesis.
Molecular identification of ticks and tick borne pathogens in Nigerian Dogs: A public health threat
Authors: Apaa Ternenge , Dunham Stephen , Tarlinton Rachael
Published: July 2020

Global warming has changed the range of tick species and the tick borne diseases (TBDs) they carry; there is an increasing need to assess the risk and impact of TBDs. This study focused on TBDs of Dogs in Nigeria, where despite a large burden of ticks and TBDs there is little data on the prevalence of either for veterinarians and human health professionals to make informed decisions regarding treatment and control.

Most existing studies on cattle have indicated that Anaplasma, Babesia, Theileria, Ehrlichia, Hepatozoon and Candidatus Neoehrlichia species are present in Nigeria. However, dogs kept in much closer contact with their owners, may represent a higher risk of zoonotic TBDs transmission. This study used a combination of morphological and molecular identification of ticks and tick borne pathogens, and IDEXX 4Dx kits for pathogen antibody detection from canine blood, we examined 93 dogs to date.

Rhipicephalus sanguineus was the most abundant tick found on Nigerian dogs. Babesia species were the most abundant tick borne pathogen, followed by Ehrlichia species. This study serves as the first report of Babesia microti like species, Theileria parva, Babesia bovis, Anaplasma phagocytophilium and Ehrlichia chaffeensis in Nigerian dogs. There was no evidence of Borrelia Burgdorferi sensu lato nor Dirofilaria species in Nigerian dogs. The presence of Babesia microti and Anaplasma phagocytophilium is a direct public threat. Further testing of additional 200 dog samples and multivariate modelling of demographic risk factors, will enable us develop treatment and control guidelines for TBDs for Nigerian Veterinarians and Dog owners.
Tropism and neutralisation studies on bat influenza H17N10
Authors: Giotis Efstathios , Carnell George , Young Erik , Ghanny Saleena , Soteropoulos Patricia , Pessi Antonello , Lacek Krzysztof , Scott Simon , Wang Linfa , Barclay Wendy , Skinner Michael , Temperton Nigel
Published: July 2020

The diversity of subtypes within Influenza A recently expanded with identification of H17N10 and H18N11 from bats. To study the tropism and zoonotic potential of these viruses, we successfully produced lentiviral pseudotypes bearing haemagglutinin H17 and neuraminidase N10. We investigated a range of cell lines from different species for their susceptibility to infection by these pseudotypes and show that a number of human haematopoietic cancer cell lines and the canine kidney MDCK II (but not MDCK I) cells are susceptible. Using microarrays and qRT-PCR we show that the dog leukocyte antigen DLA-DRA mRNA is over expressed in late passaged parental MDCK and commercial MDCK II cells, compared to early passaged parental MDCK and MDCK I cells, respectively. The human orthologue HLA-DRA encodes the alpha subunit of the MHC class II HLA-DR antigen-binding heterodimer. Small interfering RNA- or neutralizing antibody-targeting HLA-DRA, drastically reduced the susceptibility of Raji B cells to H17-PV. Conversely, over expression of HLA-DRA and its paralogue HLA-DRB1 on the surface of unsusceptible HEK293T/17 cells conferred susceptibility to H17-PV. The identification of HLA-DR as an H17N10 entry mediator will contribute to understanding the tropism of the virus and help to elucidate its zoonotic transmission. We also show that H17 pseudotypes can be efficiently neutralised by the broadly-neutralizing HA2 stalk monoclonal antibodies CR9114 and FI6. The lentiviral pseudotype system is a useful research tool, amenable for investigation of bat influenza tropism, restriction and pandemic preparedness, without safety issues of producing a replication-competent virus, to which the human population is naïve.
Characterisation of Burkholderia prophages and discovery of a novel inducible phage from B. vietnamiensis G4 that is widely distributed across the species
Authors: Weiser Rebecca , Ling Zhong , Otter Ashley , Mullins Alex , Mahenthiralingam Eshwar
Published: July 2020

Burkholderia species have environmental, industrial and medical significance, and are important opportunistic pathogens in individuals with cystic fibrosis (CF). Approximately 10% of Burkholderia genomes (6-9 Mb) are horizontally acquired material, representing a rich source of mobile genetic elements including prophages. There is limited research on Burkholderia bacteriophages, their contributions to genome evolution, virulence and antimicrobial resistance, or biotechnological and therapeutic applications. We investigated prophage carriage in Burkholderia and aimed to isolate and characterise inducible bacteriophages from B. vietnamiensis.

Burkholderia genomes were screened for prophages using PHASTER. Prophage genomes were compared using MASH and visualised with ProgressiveMauve. Phylogenomics was used to assess the distribution of prophages across B. vietnamiensis strains. Spontaneously induced phages were characterised to determine linkage between prophage regions and isolated phages, bacteriophage morphology and host range.

Prophage carriage across 456 Burkholderia strains (spanning 43 species) was high; 716 intact prophages were discovered and polylysogeny was common. In B. vietnamiensis alone, 115 prophages were identified from 81 strains, with evidence of shared prophage carriage between related and diverse strains. Three novel inducible phages were isolated from B. vietnamiensis strain G4 and their genomic origins localised putatively. One phage (vB_BvM-G4P1; family Myoviridae) had inhibitory activity against multiple strains of 5 B. cepacia complex species, including species prevalent in CF infections.

Prophages are numerous in Burkholderia genomes and contribute to strain diversity. There is huge potential for further investigation into the functional implications of prophage carriage and its impact on genome evolution, in addition to the isolation of novel bacteriophages.
Molecular detection of Mycoplasma amphoriforme and Ureaplasma spp. from patient samples previously investigated for Mycoplasma pneumoniae infection
Authors: Rehman Safi , Maddocks Sarah , Afshar Baharak , Day Jessica , Chalker Victoria , Beeton Michael
Published: July 2020

M. amphorifrome(MAM), is a novel pathogen associated with chronic respiratory tract infections in immunocompromised patients, but prevalence in immunocompetent patients is not established. Ureaplasma spp. are a known cause of hyperammonaemia in lung transplant patients. To date, their prevalence has been well documented in neonatal lungs but observations in adults is lacking.

161 samples were obtained from Public Health England, submitted previously for M. pnuemoniae (MPN) investigation, which were screened for MAM using quantitative PCR. MAM positive samples were then screened for macrolide resistance using 23s rRNA (domain V) PCR amplification and Sanger-sequencing. The 161 samples were also screened for Ureaplasma spp. using conventional PCR involving different primer sets for initial screening and subsequent differentiation between Ureaplasma spp. by targeting a variable region.

10 of 161 samples were found positive for MAM (6.2%), all of which were found to be genotypically macrolide susceptible and 9/10 positives were isolated from males. Three of the total positives were previously identified to be MPN positives. None of the samples taken during the summer months had any MAM DNA detected. Five of the 161 samples tested positive for U. parvum (3.1%), 3/5 of which were from isolated from males.

These data suggest that MAM maybe an emerging pathogen that can cause persistent respiratory infections. The lungs may represent a possible reservoir for U. parvum and source for donor-derived lung infections. Further investigation is required into the prevalence of these organisms.
Comparing the innate and humoral immune responses of different chicken lines to Infectious Bronchitis Virus (IBV) infection
Authors: Dowell Isaac , Steyn Angela , Keep Sarah , Bickerton Erica , Hammond John
Published: July 2020

Infectious Bronchitis Virus (IBV) is a gammacoronavirus that is prevalent in commercial chicken flocks, resulting in characteristic clinical signs including snicking, rales, decreased tracheal ciliary activity, reduced weight gain and reduced egg production. Preliminary results indicate that there is a different clinical response to IBV infection in different chicken lines. Therefore, we aim to determine whether there is a differential innate or humoral immune response to IBV between chicken lines.

A series of in vivo experiments were conducted comparing brown leghorns (Rhode Island Red, RIR (Roslin)) to white leghorns (from Valo and Ovagen). Trachea and bursa were collected from infected and control birds at four-, six- and fourteen-days post infection (dpi). There was a difference in snick rate and rales between the RIRs and the white leghorns (both lines). However, no difference was observed in ciliary activity. Viral load was determined by absolute quantification using qRT-PCR. The viral load in the trachea of RIRs was significantly lower (p<0.05) at 6 dpi compared to 4 dpi, unlike in Ovagen birds where there was no significant difference between the timepoints. Relative gene expression of IFN-α, IFN-β, IFN-γ, IL-6 and IL-1β in these tissues will be measuredby qRT-PCR. Serum was processed from whole blood collected at zero and ten dpi for use in IBV specific ELISAs which will measure antibody responses in the chicken lines. This project aims to explore immune responses against IBV as well as identifying the causes of variability in experimentation using chickens to investigate IBV infection.
Identification of interferon-stimulated genes with anti-HCMV activity
Authors: Lin Yao-Tang , Chiweshe Stephen , McCormick Dominique , Gaunt Eleanor , Simmonds Peter , J. Wilson Sam , Grey Finn
Published: July 2020

As a first line of defence, the interferon-mediated innate immune response is pivotal to protect cells against invading pathogens. At the heart of it are hundreds of interferon-stimulated genes (ISGs) upregulated substantially shortly after viral infection. Some of these ISGs act indirectly to interfere with virus life cycle via regulation of interferon signalling pathways whereas other ISG can act directly to inhibit viral replication via interaction with viral components. Although many of the ISGs have been identified decades ago, only a handful of them have been characterized about their antiviral activity. Here, we used an arrayed expression lentivirus library of more than 400 ISGs to identify the ISGs with anti-HCMV activity. We performed parallel screens using wild type fibroblast cells and IRF3 KO fibroblast cells generated by CRISPR/Cas9 editing to identify ISGs more likely to directly inhibit HCMV, as opposed to activation of IFN signalling. The IRF3-independent ISGs identified in the screen include those that signal through IRF3 independent pathways such as IRF7 or known to inhibit HCMV directly such as IDO, RIPK2 and AIM2 validating the screening approach. Interestingly, we also identified novel IRF3-independent anti-HCMV ISGs, indicating they may play a role in directly inhibiting the virus.
BK polyomavirus genotypes and progression of BKV disease in renal transplant recipients
Authors: Fallatah Nafraa
Published: July 2020

Background. BK virus (BKV) reactivation is a significant cause of BK Polyomavirus associated nephropathy (BKPyVN) resulting in acute graft rejection in 1-10% of post-renal transplant recipients. However, the association of BKV genotypes with development of BKPyVN is poorly understood. Here we aimed to determine the prevalence of BKV genotypes in post-renal transplant recipients, and its association with BKPyVN disease progression.

Method. Two methods were utilised to genotype BKV. A 800bp fragment of virus VP1 antigen region was amplified using nested (PCR) followed by sequencing. The genotypes were determined according to a previously developed algorithm based on analysing 100bp region of the VP1 gene. Furthermore, the logarithm results were validated with the constructed phylogenetic tree. The results were correlated with patient viral loads and development of BKPyVN.

Results. BK virus DNA was detected in 32 (69%) of 46 post-renal transplant recipients with BK viremia, while BKPyVN was only reported in two (4.3%)patients. 30 out of 32 samples were successfully genotyped (93.7%) with 23 (76.6%) belonging to the BKV Ib-2 subtype and seven (23.3%) belonging to the BKV Ib-1 subtype and with no cases representing genotype II, III and IV. All cases with confirmed BKPyVN matched to the BKV Ib-2 genotype. Additionally, no significance differences were observed between BKV genotypes in regards to viral loads, development of viremia, HLA mismatch, age or sex.

Conclusion. The results indicate no correlations between BKV genotypes and the development of BKPyVN. Furthermore,a high distribution of BKV genotype Ib-2 was found among BKV infected patients within this cohort.
The molecular interaction between ML336 and the VEEV non-structural proteins
Authors: M. Skidmore Abdrew , Chung Dong-Hoon
Published: July 2020

Venezuelan Equine Encephalitis Virus (VEEV) is a positive sense RNA virus in the family Togaviridae. VEEV circulates in the Americas, causing occasional large scale epidemics. Our group has previously discovered and described the anti-VEEV compound ML336. We found that ML336 inhibits viral RNA synthesis during infection. This RNA synthesis inhibition is highly specific for VEEV, and ML336 has no effect on the closely related chikungunya virus, or on cellular RNA synthesis. We also found that this activity was maintained in a cell-free viral RNA synthesis system, supporting our hypothesis that ML336 is a direct acting antiviral compound. We recently discovered that treatment with ML336 reduces the amount of double stranded RNA present in infected cells. This reduction supports that ML336 is interfering with the synthesis of viral RNA. This was measured qualitatively with microscopy, and quantitatively with flow cytometry. We have also reported that resistant viral mutants emerge when grown in the presence of inhibitory compounds and these mutations mapped to the N terminal domains of both nsP2 and nsP4. This region of nsP2 has recently been shown to be a helical region which serves as an accessory domain to the viral helicase. However, the region of nsP4 in question currently lacks known function. Based on this genetic data we hypothesized that ML336 and related compounds interact with these two domains to interfere with the activity of the replicase complex. We are currently examining this interaction using ectopically expressed protein.
Gene transfer agents: Another step towards a mechanistic understanding of expression
Authors: Sherlock David , Benn Millie , Fogg Paul
Published: July 2020

Gene transfer agents (GTAs) are small viruses that package and transfer random pieces of the producing cell’s genome but are unable to transfer all the genes required for their own production. GTAs are able to spread any DNA in the host cell and so their potential impact upon bacterial evolution and antimicrobial resistance is immense.

Our discovery that the product of gene rcc01865 is a specific GTA activation factor (GafA) for the model Rhodobacter capsulatus GTA (RcGTA) and that GafA is essential for RcGTA production, has provided the link between GTA production and host regulatory pathways. However, while GafA has significantly improved our understanding of GTA regulation the complete mechanism is unclear. Our goal was to investigate the GafA mechanism of action in more detail. We demonstrate direct protein-protein interaction between GafA and the RNA polymerase omega subunit (RNAP Ω) using bacterial-two-hybrid and pull down assays. Further evidence for the interaction has come from random and site directed mutagenesis of gafA and targeted truncations. GafA mutants were also tested to assess their impact on RcGTA production. RNAP Ω is thought to recruit alternative sigma factors to the RNAP holoenzyme. Regions of GafA also share sequence homology with known sigma factor proteins, and we propose that GafA acts as an alternative sigma factor to co-ordinate expression of disparate RcGTA genes. Our results advance our understanding of this fascinating mode of horizontal gene transfer, not only in the model species but also in other potential GTA producing species that contain gafA homologues.
Impact of air pollution on buff-tailed bumblebees (Bombus terrestris) and their gut microbiome
Authors: Sampson Hannah , Ketley Julian , Mallon Eamonn , Morrissey Julie
Published: July 2020

Bumblebees play a major role in global pollination. Consequently, their health is of high importance for food security worldwide. Yet, recent population estimates show that their numbers are declining. This decline has been attributed to habitat loss, infection and use of pesticides. An important factor for bee health that contributes to population survival is the gut microbiome composition. The bee gut microbiome provides protection from pathogens, is specific to the host and helps break down food. Without a balanced gut microbiome, the health of the bee is threatened through increased infection and mortality. The bee gut microbiome is relatively simple, being dominated by 8 core bacterial species providing a convenient study system. Previous published data shows that air pollution has an impact on bacterial behaviour. Therefore, our hypothesis is exposure to air pollution causes an imbalance in the bee gut microbiome. To test this, we exposed bees to black carbon (BC), a major component of air pollution particulate matter. We assessed the effects on bee behaviour, microbiome composition and gut bacteria treated in vitro. Bees treated with BC showed a significant increase in viable bacterial cells in their faecal community. Independent culture of gut commensals showed that BC significantly alters the structure of their biofilms, which are important for colonisation in vivo. This supports the hypothesis that air pollution can cause an imbalance in the bee gut microbiome, and may adversely influence bee health and pollinator populations.
MORF: An online tool for exploring microbial cell responses using multi-omics analysis
Authors: Springthorpe Vicki , Leaman Rosalyn , Sifouna Despoina , Bennett Joyce , Thomas Gavin
Published: July 2020

With continuing improvements and reducing costs of high-throughput technologies, microbiologists are increasingly collecting multi-omics datasets. However, the tools and techniques used to analyse these kinds of data are often highly specialised and require bioinformatics, statistics and often coding experience. Many studies also tend to report on a single aspect of the data whilst overlooking other potentially interesting phenomena. Consequently, many of these multi-omics data sets are not being used to their full potential. MORF was created as a solution to these problems by providing access to multi-omics datasets through an online interface which presents the data in a user-friendly and accessible way. No coding experience or specialist statistical knowledge is required, and users are free to explore the data using interactive graphics and simple analysis tools.

Here we demonstrate MORF using multi-omics datasets from two experiments using bacteria in industrial fermentation processes. First, Escherichia coli engineered to produce styrene, a valuable chemical used in the manufacture of polymers, and secondly a Clostridium which produces the biofuel butanol. A key outcome was the identification of targets believed to be involved in responding to membrane stress, which we identified using MORF’s differential gene and protein analysis tools. Work is underway to further characterise and engineer these targets to improve product yields. In conclusion, MORF provides a framework for omics analysis that can be applied to any organism or set of experimental conditions, and will help researchers and collaborators to make the most of their data.
Studying the surface-tethered AaaA as a virulence factor and anti-P. aeruginosa drug target
Authors: S. Laxton Claire , Jordana-Lluch Elena , Luckett Jeni , Heeb Stephan , Hardie Kim
Published: July 2020

P. aeruginosa is a leading cause of bacterial wound infections, and is associated with a disproportionally high level of mortality in burn patients especially. Because of its wide arsenal of virulence factors and biofilm formation ability, infections with P. aeruginosa are often chronic and extremely difficult to treat. This study uses an ex-vivo skin model to study the role of the virulence factor AaaA (PA0328) in chronic wound infections. AaaA, or the Arginine-specific aminopeptidase of Pseudomonas aeruginosa A, is a surface-tethered autotransporter which cleaves N-terminal arginine from peptides. In the oxygen and nutrient-limited environments of chronic wounds, this free arginine could serve as a nutrient source for P. aeruginosa via alternative metabolic pathways. Changes in local arginine concentrations may also alter host iNOS and Arginase immune responses which influence inflammation and wound healing, in order to favour a chronic infection. Being surface-tethered and immunogenic, AaaA is also of interest as a potential antimicrobial drug or vaccine target. This study aims to probe the role of AaaA on both pathogen survival and host response in the wound context, using a combination of in situ transcriptional reporters, immunofluorescence and laser-scanning microscopy, and RT-qPCR to localise and quantify P. aeruginosa survival and aaaA expression, as well as resident immune cell invasion, and expression of immune factor genes, in wounds over time. Here, we present preliminary data examining AaaA expression and function in P. aeruginosa biofilm cultures and ex-vivo skin wound infections, as well as the results of preliminary inhibitor screens.
Taking charge: Using electrochemical impedance spectroscopy to quantify biofilm formation in Pseudomonas spp.
Authors: Riordan Lily , Dunphy Raymond , Lasserre Perrine , Corrigan Damion , McCormick Christopher , Murray Paul , Duncan Katherine
Published: July 2020

The opportunistic pathogen, Pseudomonas aeruginosa, is infamous for its ability to rapidly form biofilms (<24 h) in inhospitable environments and the development of antimicrobial resistance (AMR). It has been seen to have resistance to nearly all known antibiotics, including last-line antibiotics colistin and carbapenems. AMR is currently considered one of the biggest threats to human health, causing 700,000 deaths annually, expected to rise to 10 million deaths per year by 2050. P. aeruginosa, alongside other opportunistic pathogens, has been implicated in infections following various surgical procedures. Such infections compromise patient recovery and, when a medical implant is present a biofilm can develop that will ultimately require a complex revision surgery to remove the infection.

In this study, impedance spectroscopy and differential pulse voltammetry were carried out in parallel to measure electrochemical and impedance properties of bacteria, allowing for identification and quantification of pyoverdine and pyocyanin; bacterial metabolites. Three Pseudomonas spp. (P. aeruginosa, P. fluorescens and P. putida) were assayed in liquid culture at OD600. The sensor was standardised with pyoverdine and pyocyanin, with an electrochemical reading taken every 30 minutes up to 4 hours. This assay was repeated with Pseudomonas spp. growing in biofilms in LB broth, with a screen-printed electrode as the solid surface. Readings were then used to correlate metabolite production to biofilm production in each Pseudomonas sp. Pyoverdine correlated with biofilm formation for all three assayed Pseudomonas, with variation in the quantity of metabolite produced between species. This allows the two metabolites to be used as indicators of biofilm mass on devices and surfaces.
Revisiting the role of PML protein targeting and disruption of PML bodies in human cytomegalovirus infection
Authors: Paulus Christina , Harwardt Thomas , Walter Bernadette , Marxreiter Andrea , Nevels Michael
Published: July 2020

Promyelocytic leukaemia (PML) bodies are nuclear organelles implicated in post-translational modification by small ubiquitin-like modifier (SUMO) proteins and in the antiviral host cell response to infection. The 72-kDa immediate-early protein 1 (IE1) is considered the principal antagonist of PML bodies encoded by the human cytomegalovirus, one of eight human herpesviruses. Previous work has suggested that the interaction between IE1 and PML proteins, the central organisers of PML bodies, and the subsequent disruption of these organelles serve a critical role in viral replication by counteracting intrinsic antiviral immunity and the induction of interferon (IFN)-stimulated genes. However, this picture has emerged largely from studying mutant IE1 proteins known or predicted to be globally misfolded und metabolically unstable. We systematically screened for stable IE1 mutants by clustered charge-to-alanine scanning. We identified a mutant protein (IE1cc172-176) selectively defective for PML interaction. Functional comparisons between the mutant and wild-type protein revealed that IE1 can undergo modification by mixed polymeric SUMO chains and that it targets PML and Sp100, the two main constituents of PML bodies, via distinct mechanisms. Unexpectedly, IE1cc172-176 supported viral replication almost as efficiently as wild-type IE1. Moreover, lower instead of higher (as expected) levels of tumor necrosis factor alpha, IFN-beta, IFN-lambda and IFN-stimulated gene expression were observed with the mutant compared to the wild-type protein and virus. These results suggest that the disruption of PML bodies is linked to induction rather than inhibition of antiviral gene expression. Our findings challenge current views regarding the role of PML bodies in viral infection.