Transposon mutagenesis in a hyper-invasive clinical isolate of Campylobacter jejuni reveals a number of genes with potential roles in invasion

Transposon mutagenesis has been applied to a hyper-invasive clinical isolate of Campylobacter jejuni, 01/51. A random transposon mutant library was screened in an in vitro assay of invasion and 26 mutants with a significant reduction in invasion were identified. Given that the invasion potential of C. jejuni is relatively poor compared to other enteric pathogens, the use of a hyper-invasive strain was advantageous as it greatly facilitated the identification of mutants with reduced invasion. The location of the transposon insertion in 23 of these mutants has been determined; all but three of the insertions are in genes also present in the genome-sequenced strain NCTC 11168. Eight of the mutants contain transposon insertions in one region of the genome (∼14 kb), which when compared with the genome of NCTC 11168 overlaps with one of the previously reported plasticity regions and is likely to be involved in genomic variation between strains. Further characterization of one of the mutants within this region has identified a gene that might be involved in adhesion to host cells

Cytolethal distending toxin (CDT)-negative Campylobacter jejuni strains and anti-CDT neutralising antibodies induced during human infection but not chicken colonisation

The cytolethal distending toxin (CDT) of Campylobacter jejuni was detectable, using an in vitro assay, in most but not all of 24 strains tested. The reason for the absence of toxin activity in these naturally occurring CDT-negative C. jejuni strains was then investigated at the genetic level. CDT is encoded by three highly conserved genes, cdtA, -B, and -C. In the CDT-negative strains, two types of mutation were identified. The CDT activities of C. jejuni strains possessing both types of mutation were successfully complemented with the functional genes of C. jejuni 11168. The first type of mutation comprised a 667-bp deletion across cdtA and cdtB and considerable degeneration in the remainder of the cdt locus. Using a PCR technique to screen for this deletion, this mutation occurred in fewer than 3% of 147 human, veterinary, and environmental strains tested. The second type of mutation involved at least four nonsynonymous nucleotide changes, but only the replacement of proline with serine at CdtB position 95 was considered important for CDT activity. This was confirmed by site-directed mutagenesis. This type of mutation also occurred in fewer than 3% of strains as determined using a LightCycler biprobe assay. The detection of two CDT-negative clinical isolates raised questions about the role of CDT in some cases of human campylobacteriosis. To determine if anti-CDT antibodies are produced in human infection, a toxin neutralization assay was developed and validated using rabbit antisera. Pooled human sera from infected patients neutralized the toxin, indicating expression and immunogenicity during infection. However, no neutralizing antibodies were detected in colonized chickens despite the expression of CDT in the avian gut as indicated by reverse transcription-PCR

Campylobacter

Background: The objective of this study was to characterise Campylobacter growth in enrichment broths (Bolton broth, brain heart infusion broth), caecal material (in vitro), and in the naturally infected live broilers (in vivo) in terms of mean lag periods and generation times as well as maximum growth rates and population (cell concentration) achieved. Methods: Bolton and brain heart infusion broths and recovered caecal material were inoculated with 10 poultry strains of Campylobacter (eight Campylobacter jejuni and two Campylobacter coli), incubated under microaerobic conditions, and Campylobacter concentrations determined periodically using the ISO 10272:2006 method. Caeca from 10 flocks, infected at first thinning, were used to characterise Campylobacter growth in the live birds. Mean generation times (G) (early lag to exponential phase) were calculated using the formula: G=t/3.3 logb/B. Mean lag times and µmax were calculated using the Micro Fit© Software (Version 1.0, Institute of Food Research). Statistical comparison was performed using GENSTAT ver. 14.1 (VSN International Ltd., Hemel, Hempstead, UK). Results: The mean lag periods in Bolton broth, brain heart infusion broth, caecal material, and in the live bird were estimated to be 6.6, 6.7, 12.6, and 31.3 h, respectively. The corresponding mean generation times were 2.1, 2.2, 3.1, and 6.7 h, respectively; maximum growth rates were 0.7, 0.8, 0.4, and 2 generations h−1 and the maximum populations obtained in each matrix were 9.6, 9.9, 7.8, and 7.4 log10 CFU/g, respectively. Conclusion: This study provides data on the growth of Campylobacter in a range of laboratory media, caecal contents, and in broilers which may be used to develop predictive models and/or inform science-based control strategies such as the maximum time between flock testing and slaughter, logistical slaughter, and single-stage depopulation of broiler units

Detection of emphysema progression in alpha 1-antitrypsin deficiency using CT densitometry; Methodological advances

<p>Abstract</p> <p>Background</p> <p>Computer tomography (CT) densitometry is a potential tool for detecting the progression of emphysema but the optimum methodology is uncertain. The level of inspiration affects reproducibility but the ability to adjust for this variable is facilitated by whole lung scanning methods. However, emphysema is frequently localised to sub-regions of the lung and targeted densitometric sampling may be more informative than whole lung assessment.</p> <p>Methods</p> <p>Emphysema progression over a 2-year interval was assessed in 71 patients (alpha 1-antitrypsin deficiency with PiZ phenotype) with CT densitometry, using the 15^thpercentile point (Perc15) and voxel index (VI) -950 Hounsfield Units (HU) and -910 HU (VI -950 and -910) on whole lung, limited single slices, and apical, central and basal thirds. The relationship between whole lung densitometric progression (ΔCT) and change in CT-derived lung volume (ΔCT_Vol) was characterised, and adjustment for lung volume using statistical modelling was evaluated.</p> <p>Results</p> <p>CT densitometric progression was statistically significant for all methods. ΔCT correlated with ΔCT_Voland linear regression indicated that nearly one half of lung density loss was secondary to apparent hyperinflation. The most accurate measure was obtained using a random coefficient model to adjust for lung volume and the greatest progression was detected by targeted sampling of the middle third of the lung.</p> <p>Conclusion</p> <p>Progressive hyperinflation may contribute significantly to loss of lung density, but volume effects and absolute tissue loss can be identified by statistical modelling. Targeted sampling of the middle lung region using Perc15 appears to be the most robust measure of emphysema progression.</p

End-to-end 6-DoF Object Pose Estimation through Differentiable Rasterization

Here we introduce an approximated differentiable renderer to refine a 6-DoF pose prediction using only 2D alignment information. To this end, a two-branched convolutional encoder network is employed to jointly estimate the object class and its 6-DoF pose in the scene. We then propose a new formulation of an approximated differentiable renderer to re-project the 3D object on the image according to its predicted pose; in this way the alignment error between the observed and the re-projected object silhouette can be measured. Since the renderer is differentiable, it is possible to back-propagate through it to correct the estimated pose at test time in an online learning fashion. Eventually we show how to leverage the classification branch to profitably re-project a representative model of the predicted class (i.e. a medoid) instead. Each object in the scene is processed independently and novel viewpoints in which both objects arrangement and mutual pose are preserved can be rendered. Differentiable renderer code is available at:https://github.com/ndrplz/tensorflow-mesh-renderer

Hand and Eye Dominance in Sport: Are Cricket Batters Taught to Bat Back-to-Front?

Background: When first learning to bimanually use a tool to hit a target (e.g., when chopping wood or hitting a golf ball), most people assume a stance that is dictated by their dominant hand. By convention, this means that a ‘right-handed’ or ‘left-handed’ stance that places the dominant hand closer to the striking end of the tool is adopted in many sports. Objective: The aim of this study was to investigate whether the conventional stance used for bimanual hitting provides the best chance of developing expertise in that task. Methods: Our study included 43 professional (international/first-class) and 93 inexperienced (<5 years’ experience) cricket batsmen. We determined their batting stance (plus hand and eye dominance) to compare the proportion of batters who adopted a reversed stance when batting (that is, the opposite stance to that expected based on their handedness). Results: We found that cricket batsmen who adopted a reversed stance had a stunning advantage, with professional batsmen 7.1 times more likely to adopt a reversed stance than inexperienced batsmen, independent of whether they batted right or left handed or the position of their dominant eye. Conclusion: Findings imply that batsmen who adopt a conventional stance may inadvertently be batting ‘back-to-front’ and have a significant disadvantage in the game. Moreover, the results may generalize more widely, bringing into question the way in which other bimanual sporting actions are taught and performed

The effect of the timing of exposure to Campylobacter jejuni on the gut microbiome and inflammatory responses of broiler chickens

Background Campylobacters are an unwelcome member of the poultry gut microbiota in terms of food safety. The objective of this study was to compare the microbiota, inflammatory responses, and zootechnical parameters of broiler chickens not exposed to Campylobacter jejuni with those exposed either early at 6 days old or at the age commercial broiler chicken flocks are frequently observed to become colonized at 20 days old. Results Birds infected with Campylobacter at 20 days became cecal colonized within 2 days of exposure, whereas birds infected at 6 days of age did not show complete colonization of the sample cohort until 9 days post-infection. All birds sampled thereafter were colonized until the end of the study at 35 days (mean 6.1 log10 CFU per g of cecal contents). The cecal microbiota of birds infected with Campylobacter were significantly different to age-matched non-infected controls at 2 days post-infection but generally the composition of the cecal microbiota were more affected by bird age as the time post infection increased. The effects of Campylobacter colonization on the cecal microbiota were associated with reductions in the relative abundance of OTUs within the taxonomic family Lactobacillaceae and the Clostridium cluster XIVa. Specific members of the Lachnospiraceae and Ruminococcaceae families exhibit transient shifts in microbial community populations dependent upon the age at which the birds become colonized by C. jejuni. Analysis of ileal and cecal chemokine/cytokine gene expression revealed increases in IL-6, IL-17A and Il-17F consistent with a Th17 response but the persistence of the response was dependent on the stage/time of C. jejuni colonization that coincide with significant reductions in the abundance of Clostridium cluster XIVa. Conclusions This study combines microbiome data, cytokine/chemokine gene expression with intestinal villus and crypt measurements to compare chickens colonized early or late in the rearing cycle to provide insights into the process and outcomes of Campylobacter colonization. Early colonization results in a transient growth rate reduction and pro-inflammatory response but persistent modification of the cecal microbiota. Late colonization produces pro-inflammatory responses with changes in the cecal microbiota that will endure in market ready chickens

Genotypic characterisation and cluster analysis of Campylobacter jejuni isolates from domestic pets, human clinical cases and retail food

The genetic similarity of Campylobacter jejuni isolates from pets, compared to human clinical cases and retail food isolates collected in Ireland over 2001-2006 was investigated by cluster analysis of pulsed-field gel electrophoresis (PFGE) fingerprinting profiles. Comparison of the PFGE profiles of 60 pet isolates and 109 human isolates revealed that seven (4.1%) profiles were grouped in clusters including at least one human and one pet C. jejuni isolate. In total six (1.6%) of 60 pet and 310 food profiles were in clusters with at least one food and one pet C. jejuni isolate. The detection of only a small number of genetically indistinguishable isolates by PFGE profile cluster analysis from pets and from humans with enteritis in this study suggests that pets are unlikely to be an important reservoir for human campylobacteriosis in Ireland. However, genetically indistinguishable isolates were detected and C. jejuni from pets may circulate and may contribute to clinical infections in humans. In addition, contaminated food fed to pets may be a potential source of Campylobacter infection in pets, which may subsequently pose a risk to humans