Molecular Characterisation of Bacteriophage K Towards Applications for the Biocontrol of Pathogenic Staphylococci

End of project reportThe aim of this work was to characterise staphylococcal bacteriophage (a bacterial virus) and to assess their potential as therapeutic agents against pathogenic strains of Staphylococcus aureus, particularly mastitis-causing strains. The project included the use of two newly isolated phage CS1 and DW2, and an existing polyvalent phage. The new phage were isolated from the farmyard and characterised by electron microscopy and restriction analysis. Both phage were shown to belong to the Siphoviridae family and were lytic for representatives of all three clonal groups of Irish mastitis-associated staphylococci. A cocktail of three phage (CS1, DW2 and K) at 108 (plaque forming units) PFU/ml was infused into cows teats in animal trials. The lack of an increase in somatic cell counts in milks indicated strongly that the phage did not irritate the animal. In addition, the most potent phage used in this study, phage K, was further studied by genome sequencing, which revealed a linear DNA genome of 127,395 base pairs, which encodes 118 putative ORFs (open reading frames)

Identification of EOMES-expressing spermatogonial stem cells and their regulation by PLZF.

Long-term maintenance of spermatogenesis in mammals is supported by GDNF, an essential growth factor required for spermatogonial stem cell (SSC) self-renewal. Exploiting a transgenic GDNF overexpression model, which expands and normalizes the pool of undifferentiated spermatogonia between Plzf +/+ and Plzf lu/lu mice, we used RNAseq to identify a rare subpopulation of cells that express EOMES, a T-box transcription factor. Lineage tracing and busulfan challenge show that these are SSCs that contribute to steady state spermatogenesis as well as regeneration following chemical injury. EOMES+ SSCs have a lower proliferation index in wild-type than in Plzf lu/lu mice, suggesting that PLZF regulates their proliferative activity and that EOMES+ SSCs are lost through proliferative exhaustion in Plzf lu/lu mice. Single cell RNA sequencing of EOMES+ cells from Plzf +/+ and Plzf lu/lu mice support the conclusion that SSCs are hierarchical yet heterogeneous

Somatostatin-expressing parafacial neurons are CO2/H+ sensitive and regulate baseline breathing.

Glutamatergic neurons in the retrotrapezoid nucleus (RTN) function as respiratory chemoreceptors by regulating breathing in response to tissue CO2/H+. The RTN and greater parafacial region may also function as a chemosensing network composed of CO2/H+-sensitive excitatory and inhibitory synaptic interactions. In the context of disease, we showed that loss of inhibitory neural activity in a mouse model of Dravet syndrome disinhibited RTN chemoreceptors and destabilized breathing (Kuo et al., 2019). Despite this, contributions of parafacial inhibitory neurons to control of breathing are unknown, and synaptic properties of RTN neurons have not been characterized. Here, we show the parafacial region contains a limited diversity of inhibitory neurons including somatostatin (Sst)-, parvalbumin (Pvalb)-, and cholecystokinin (Cck)-expressing neurons. Of these, Sst-expressing interneurons appear uniquely inhibited by CO2/H+. We also show RTN chemoreceptors receive inhibitory input that is withdrawn in a CO2/H+-dependent manner, and chemogenetic suppression of Sst+ parafacial neurons, but not Pvalb+ or Cck+ neurons, increases baseline breathing. These results suggest Sst-expressing parafacial neurons contribute to RTN chemoreception and respiratory activity

Cellular taxonomy and spatial organization of the murine ventral posterior hypothalamus.

The ventral posterior hypothalamus (VPH) is an anatomically complex brain region implicated in arousal, reproduction, energy balance, and memory processing. However, neuronal cell type diversity within the VPH is poorly understood, an impediment to deconstructing the roles of distinct VPH circuits in physiology and behavior. To address this question, we employed a droplet-based single-cell RNA sequencing (scRNA-seq) approach to systematically classify molecularly distinct cell populations in the mouse VPH. Analysis of \u3e16,000 single cells revealed 20 neuronal and 18 non-neuronal cell populations, defined by suites of discriminatory markers. We validated differentially expressed genes in selected neuronal populations through fluorescence in situ hybridization (FISH). Focusing on the mammillary bodies (MB), we discovered transcriptionally-distinct clusters that exhibit neuroanatomical parcellation within MB subdivisions and topographic projections to the thalamus. This single-cell transcriptomic atlas of VPH cell types provides a resource for interrogating the circuit-level mechanisms underlying the diverse functions of VPH circuits

Competitive drivers of interspecific deviations of crown morphology from theoretical predictions measured with Terrestrial Laser Scanning

Abstract: Tree crown morphology is a key driver of forest dynamics, determining not only the competitiveness of an individual but also the competitive effect exerted on neighbouring trees. Multiple ecological theories, including metabolic scaling theory (MST), predict crown morphology from first principles, but typically lack consideration of competition. The accurate quantification of crown morphology to test theoretical predictions, and the canopy interactions that could alter them, has historically been limited by the simplicity and associated error of traditional crown measurements. In this study, we calculate high‐resolution two‐ and three‐dimensional crown metrics from Terrestrial Laser Scanning data for 1,441 Pinus sylvestris, P. nigra, Quercus faginea and Q. ilex trees from a water‐limited forest community in central Spain and test height‐crown metric scaling relationships. We demonstrate new TLS methods to define symmetric and asymmetric neighbourhood metrics based on tree height, crown size and neighbour projected crown area, and test the importance of neighbourhood genus diversity on crown morphology by separating competition from congeneric and heterogeneric neighbours. Competition negatively impacted all crown metrics except crown depth where only P. nigra showed sensitivity. Asymmetric competition was the strongest driver of pine crown morphology, but oaks were more sensitive to symmetric competition, in line with shade tolerance expectations. Congeneric competition reduced Q. faginea crown size and changed its shape, but we found no significant effects of heterogeneric neighbours. Most species and crown dimensions had height‐crown scaling exponents below those predicted by MST, which may be due to water limitation effects. Pines and oaks showed large differences in crown depth to height scaling, with the former shallower and the latter deeper, in contrast to theoretical predictions. Synthesis. Our study is the first to demonstrate the ability of TLS to characterise crown morphology from leaf‐wood separated clouds and competitive neighbour effects in a water‐limited forest community, and to use TLS metrics to test ecological crown scaling theory. Most crown metrics scaled below theoretical predictions. Pines were more sensitive to competition by larger neighbours and oaks to crowding from all neighbours, with competition from neighbours of the same genus having a consistent negative effect

Non-thermal Plasma Exposure Rapidly Attenuates Bacterial AHL-Dependent Quorum Sensing and Virulence.

The antimicrobial activity of atmospheric pressure non-thermal plasma has been exhaustively characterised, however elucidation of the interactions between biomolecules produced and utilised by bacteria and short plasma exposures are required for optimisation and clinical translation of cold plasma technology. This study characterizes the effects of non-thermal plasma exposure on acyl homoserine lactone (AHL)-dependent quorum sensing (QS). Plasma exposure of AHLs reduced the ability of such molecules to elicit a QS response in bacterial reporter strains in a dose-dependent manner. Short exposures (30–60 s) produce of a series of secondary compounds capable of eliciting a QS response, followed by the complete loss of AHL-dependent signalling following longer exposures. UPLC-MS analysis confirmed the time-dependent degradation of AHL molecules and their conversion into a series of by-products. FT-IR analysis of plasma-exposed AHLs highlighted the appearance of an OH group. In vivo assessment of the exposure of AHLs to plasma was examined using a standard in vivo model. Lettuce leaves injected with the rhlI/lasI mutant PAO-MW1 alongside plasma treated N-butyryl-homoserine lactone and n-(3-oxo-dodecanoyl)-homoserine lactone, exhibited marked attenuation of virulence. This study highlights the capacity of atmospheric pressure non-thermal plasma to modify and degrade AHL autoinducers thereby attenuating QS-dependent virulence in P. aeruginosa

Prospectus, October 21, 1987

https://spark.parkland.edu/prospectus_1987/1022/thumbnail.jp

Biodiversity as a multidimensional construct: a review, framework and case study of herbivory's impact on plant biodiversity

Biodiversity is inherently multidimensional, encompassing taxonomic, functional, phylogenetic, genetic, landscape and many other elements of variability of life on the Earth. However, this fundamental principle of multidimensionality is rarely applied in research aimed at understanding biodiversity's value to ecosystem functions and the services they provide. This oversight means that our current understanding of the ecological and environmental consequences of biodiversity loss is limited primarily to what unidimensional studies have revealed. To address this issue, we review the literature, develop a conceptual framework for multidimensional biodiversity research based on this review and provide a case study to explore the framework. Our case study specifically examines how herbivory by whitetail deer (Odocoileus virginianus) alters the multidimensional influence of biodiversity on understory plant cover at Black Rock Forest, New York. Using three biodiversity dimensions (taxonomic, functional and phylogenetic diversity) to explore our framework, we found that herbivory alters biodiversity's multidimensional influence on plant cover; an effect not observable through a unidimensional approach. Although our review, framework and case study illustrate the advantages of multidimensional over unidimensional approaches, they also illustrate the statistical and empirical challenges such work entails. Meeting these challenges, however, where data and resources permit, will be important if we are to better understand and manage the consequences we face as biodiversity continues to decline in the foreseeable future