Biotechnological applications of functional metagenomics in the food and pharmaceutical industries

peer-reviewedMicroorganisms are found throughout nature, thriving in a vast range of environmental conditions. The majority of them are unculturable or difficult to culture by traditional methods. Metagenomics enables the study of all microorganisms, regardless of whether they can be cultured or not, through the analysis of genomic data obtained directly from an environmental sample, providing knowledge of the species present, and allowing the extraction of information regarding the functionality of microbial communities in their natural habitat. Function-based screenings, following the cloning and expression of metagenomic DNA in a heterologous host, can be applied to the discovery of novel proteins of industrial interest encoded by the genes of previously inaccessible microorganisms. Functional metagenomics has considerable potential in the food and pharmaceutical industries, where it can, for instance, aid (i) the identification of enzymes with desirable technological properties, capable of catalyzing novel reactions or replacing existing chemically synthesized catalysts which may be difficult or expensive to produce, and able to work under a wide range of environmental conditions encountered in food and pharmaceutical processing cycles including extreme conditions of temperature, pH, osmolarity, etc; (ii) the discovery of novel bioactives including antimicrobials active against microorganisms of concern both in food and medical settings; (iii) the investigation of industrial and societal issues such as antibiotic resistance development. This review article summarizes the state-of-the-art functional metagenomic methods available and discusses the potential of functional metagenomic approaches to mine as yet unexplored environments to discover novel genes with biotechnological application in the food and pharmaceutical industries.Science Foundation Ireland(SFI)Grant Number 13/SIRG/215

New Weapons to Fight Old Enemies: Novel Strategies for the (Bio)control of Bacterial Biofilms in the Food Industry

peer-reviewedBiofilms are microbial communities characterized by their adhesion to solid surfaces and the production of a matrix of exopolymeric substances, consisting of polysaccharides, proteins, DNA and lipids, which surround the microorganisms lending structural integrity and a unique biochemical profile to the biofilm. Biofilm formation enhances the ability of the producer/s to persist in a given environment. Pathogenic and spoilage bacterial species capable of forming biofilms are a significant problem for the healthcare and food industries, as their biofilm-forming ability protects them from common cleaning processes and allows them to remain in the environment post-sanitation. In the food industry, persistent bacteria colonize the inside of mixing tanks, vats and tubing, compromising food safety and quality. Strategies to overcome bacterial persistence through inhibition of biofilm formation or removal of mature biofilms are therefore necessary. Current biofilm control strategies employed in the food industry (cleaning and disinfection, material selection and surface preconditioning, plasma treatment, ultrasonication, etc.), although effective to a certain point, fall short of biofilm control. Efforts have been explored, mainly with a view to their application in pharmaceutical and healthcare settings, which focus on targeting molecular determinants regulating biofilm formation. Their application to the food industry would greatly aid efforts to eradicate undesirable bacteria from food processing environments and, ultimately, from food products. These approaches, in contrast to bactericidal approaches, exert less selective pressure which in turn would reduce the likelihood of resistance development. A particularly interesting strategy targets quorum sensing systems, which regulate gene expression in response to fluctuations in cell-population density governing essential cellular processes including biofilm formation. This review article discusses the problems associated with bacterial biofilms in the food industry and summarizes the recent strategies explored to inhibit biofilm formation, with special focus on those targeting quorum sensing.Science Foundation Irelan

Observational constraints on an inflation model with a running mass

We explore a model of inflation where the inflaton mass-squared is generated at a high scale by gravity-mediated soft supersymmetry breaking, and runs at lower scales to the small value required for slow-roll inflation. The running is supposed to come from the coupling of the inflaton to a non-Abelian gauge field. In contrast with earlier work, we do not constrain the magnitude of the supersymmetry breaking scale, and we find that the model might work even if squark and slepton masses come from gauge-mediated supersymmetry breaking. With the inflaton and gaugino masses in the expected range, and $\alpha = g^2/4\pi$ in the range $10^{-2}$ to $10^{-3}$ (all at the high scale) the model can give the observed cosmic microwave anisotropy, and a spectral index in the observed range. The latter has significant variation with scale, which can confirm or rule out the model in the forseeable future.Comment: Latex, 19 pages, 14 figures, uses epsf.st

Pseudo-planar Ge-on-Si Single-photon Avalanche Diode Detector with Record Low Noise-equivalent Power

Single-photon avalanche diode (SPAD) detectors are of significant interest for numerous applications, including light detection and ranging (LIDAR), and quantum technologies such as quantum-key distribution and quantum information processing. Here we present a record low noise-equivalent-power (NEP) for Ge-on-Si SPADs using a pseudo-planar design, showing high detection efficiency in the short-wave infrared; a spectral region which is key for quantum technologies and hugely beneficial for LIDAR. These devices can leverage the benefits of Si avalanche layers, with lower afterpulsing compared to InGaAs/InP, and reduced cost due to Si foundry compatibility. By scaling the SPAD pixels down to 26μm diameter, a step change in performance has been demonstrated, with significantly reduced dark count rates (DCRs), and low jitter (134ps). Ge-on-Si SPADs were fabricated using photolithography techniques and characterised using time-correlated single-photon counting. The DCR reaches as low as kilocount/s at 100K for excess bias up to ~5%. This reduction in DCR enables higher temperature operation; e.g. the DCR of a 26μm diameter pixel at 150 K is approximately equivalent to a 100 μm diameter pixel at 77 K (100s of kilocounts/s). These low values of DCR, coupled with the relatively temperature independent single photon detection efficiencies (SPDE) of ~29% (at 1310nm wavelength) leads to a record low NEP of 7.7×10−17WHz−1/2. This is approximately 2 orders of magnitude lower than previous similarly sized mesa-geometry Ge-on-Si SPADs. This technology can potentially offer a lowcost, Si foundry compatible SPAD operating at short-wave infrared wavelengths, with potential applications in quantum technologies and autonomous vehicle LIDAR

Defining a Novel Role for the Coxsackievirus and Adenovirus Receptor in Human Adenovirus Serotype 5 Transduction In Vitro in the Presence of Mouse Serum

Human adenoviral serotype 5 (HAdV-5) vectors have predominantly hepatic tropism when delivered intravascularly, resulting in immune activation and toxicity. Coagulation FX binding to HAdV-5 mediates liver transduction and provides protection from virion neutralisation in mice. FX is dispensable for liver transduction in mice lacking IgM antibodies or complement, suggesting alternative transduction pathways exist. To identify novel factor(s) mediating HAdV-5 FX-independent entry, we investigated HAdV-5 transduction in vitro in the presence of serum from immunocompetent C57BL/6 or immunocompromised mice lacking IgM antibodies (Rag 2-/- and NSG). Sera from all three mouse strains enhanced HAdV-5 transduction of A549 cells. While inhibition of HAdV-5:FX interaction with X-bp inhibited transduction in the presence of C57BL/6 serum, it had negligible effect on the enhanced transduction observed in the presence of Rag 2-/- or NSG serum. Rag 2-/- serum also enhanced transduction of the FX-binding deficient AdT*. Interestingly, Rag 2-/- serum enhanced HAdV-5 transduction in a FX-independent manner in CHO-CAR and SKOV3-CAR cells. Additionally, blockade of CAR with soluble HAdV-5 fiber knob inhibited mouse serum-enhanced transduction in A549 cells, suggesting a potential role for CAR. Transduction of HAdV-5 KO1 and HAdV-5/F35 (CAR-binding deficient) in the presence of Rag 2-/- serum was equivalent to that of HAdV-5, indicating that direct interaction between HAdV-5 and CAR is not required. These data suggest that FX may protect HAdV-5 from neutralization but has minimal contribution to HAdV-5 transduction in the presence of immunocompromised mouse serum. Alternatively, transduction occurs via an unidentified mouse serum protein capable of bridging HAdV-5 to CAR

Relationships between Challenge Tour golfers’ clubhead velocity and force producing capabilities during a countermovement jump and isometric mid-thigh pull

A number of field-based investigations have evidenced practically significant relationships between clubhead velocity (CHV), vertical jump performance and maximum strength. Unfortunately, whilst these investigations provide a great deal of external validity, they are unable to ascertain vertical ground reaction force (vGRF) variables that may relate to golfers’ CHVs. This investigation aimed to assess if the variance in European Challenge Tour golfers’ CHVs could be predicted by countermovement jump (CMJ) positive impulse (PI), isometric mid-thigh pull (IMTP) peak force (PF) and rate of force development (RFD) from 0–50 ms, 0–100 ms, 0–150 ms and 0–200 ms. Thirty-one elite level European Challenge Tour golfers performed a CMJ and IMTP on dual force plates at a tournament venue, with CHV measured on a driving range. Hierarchical multiple regression results indicated that the variance in CHV was significantly predicted by all four models (model one R2 = 0.379; model two R2 = 0.392, model three R2 = 0.422, model four R2 = 0.480), with Akaike’s information criterion indicating that model one was the best fit. Individual standardised beta coefficients revealed that CMJ PI was the only significant variable, accounting for 37.9% of the variance in European Challenge Tour Golfers’ CHVs

Tandem amplification of SCCmec can drive high level methicillin resistance in MRSA

Hospital-associated methicillin-resistant Staphylococcus aureus (MRSA) strains typically express high-level, homogeneous (HoR) beta-lactam resistance, whereas community-associated MRSA (CA-MRSA) more commonly express low-level heterogeneous (HeR) resistance. Expression of the HoR phenotype typically requires both increased expression of the mecA gene, carried on the staphylococcal cassette chromosome mec element (SCCmec), and additional mutational event(s) elsewhere on the chromosome. Here the oxacillin concentration in a chemostat culture of the CA-MRSA strain USA300 was increased from 8 mu g/ml to 130 mu g/ml over 13 days to isolate highly oxacillin-resistant derivatives. A stable, small-colony variant, designated HoR34, which had become established in the chemostat culture was found to have acquired mutations in gdpP, clpX, guaA, and camS. Closer inspection of the genome sequence data further revealed that reads covering SCCmec were similar to 10 times overrepresented compared to other parts of the chromosome. Quantitative PCR (qPCR) confirmed >10-fold-higher levels of mecA DNA on the HoR34 chromosome, and MinION genome sequencing verified the presence of 10 tandem repeats of the SCCmec element. qPCR further demonstrated that subculture of HoR34 in various concentrations of oxacillin (0 to 100 mu g/ml) was accompanied by accordion-like contraction and amplification of the SCCmec element. Although slower growing than strain USA300, HoR34 outcompeted the parent strain in the presence of subinhibitory oxacillin. These data identify tandem amplification of the SCCmec element as a new mechanism of high-level methicillin resistance in MRSA, which may provide a competitive advantage for MRSA under antibiotic selection

Mitochondrial physiology

As the knowledge base and importance of mitochondrial physiology to evolution, health and disease expands, the necessity for harmonizing the terminology concerning mitochondrial respiratory states and rates has become increasingly apparent. The chemiosmotic theory establishes the mechanism of energy transformation and coupling in oxidative phosphorylation. The unifying concept of the protonmotive force provides the framework for developing a consistent theoretical foundation of mitochondrial physiology and bioenergetics. We follow the latest SI guidelines and those of the International Union of Pure and Applied Chemistry (IUPAC) on terminology in physical chemistry, extended by considerations of open systems and thermodynamics of irreversible processes. The concept-driven constructive terminology incorporates the meaning of each quantity and aligns concepts and symbols with the nomenclature of classical bioenergetics. We endeavour to provide a balanced view of mitochondrial respiratory control and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. Uniform standards for evaluation of respiratory states and rates will ultimately contribute to reproducibility between laboratories and thus support the development of data repositories of mitochondrial respiratory function in species, tissues, and cells. Clarity of concept and consistency of nomenclature facilitate effective transdisciplinary communication, education, and ultimately further discovery

Mitochondrial physiology

As the knowledge base and importance of mitochondrial physiology to evolution, health and disease expands, the necessity for harmonizing the terminology concerning mitochondrial respiratory states and rates has become increasingly apparent. The chemiosmotic theory establishes the mechanism of energy transformation and coupling in oxidative phosphorylation. The unifying concept of the protonmotive force provides the framework for developing a consistent theoretical foundation of mitochondrial physiology and bioenergetics. We follow the latest SI guidelines and those of the International Union of Pure and Applied Chemistry (IUPAC) on terminology in physical chemistry, extended by considerations of open systems and thermodynamics of irreversible processes. The concept-driven constructive terminology incorporates the meaning of each quantity and aligns concepts and symbols with the nomenclature of classical bioenergetics. We endeavour to provide a balanced view of mitochondrial respiratory control and a critical discussion on reporting data of mitochondrial respiration in terms of metabolic flows and fluxes. Uniform standards for evaluation of respiratory states and rates will ultimately contribute to reproducibility between laboratories and thus support the development of data repositories of mitochondrial respiratory function in species, tissues, and cells. Clarity of concept and consistency of nomenclature facilitate effective transdisciplinary communication, education, and ultimately further discovery