Quantitative trait loci and transcriptome signatures associated with avian heritable resistance to Campylobacter.

Campylobacter is the leading cause of bacterial foodborne gastroenteritis worldwide. Handling or consumption of contaminated poultry meat is a key risk factor for human campylobacteriosis. One potential control strategy is to select poultry with increased resistance to Campylobacter. We associated high-density genome-wide genotypes (600K single nucleotide polymorphisms) of 3000 commercial broilers with Campylobacter load in their caeca. Trait heritability was modest but significant (h2 = 0.11 ± 0.03). Results confirmed quantitative trait loci (QTL) on chromosomes 14 and 16 previously identified in inbred chicken lines, and detected two additional QTLs on chromosomes 19 and 26. RNA-Seq analysis of broilers at the extremes of colonisation phenotype identified differentially transcribed genes within the QTL on chromosome 16 and proximal to the major histocompatibility complex (MHC) locus. We identified strong cis-QTLs located within MHC suggesting the presence of cis-acting variation in MHC class I and II and BG genes. Pathway and network analyses implicated cooperative functional pathways and networks in colonisation, including those related to antigen presentation, innate and adaptive immune responses, calcium, and renin-angiotensin signalling. While co-selection for enhanced resistance and other breeding goals is feasible, the frequency of resistance-associated alleles was high in the population studied and non-genetic factors significantly influenced Campylobacter colonisation.

Characterising the Influence of Genetics on Breast Muscle Myopathies in Broiler Chickens.

This report provides the first estimates of the genetic basis of all key breast muscle myopathies (BMM) in broiler chickens [Deep pectoral myopathy, wooden breast, white striping and spaghetti breast] and their relationship with body weight and breast yield. Data from a pure bred high yielding commercial broiler line were analysed to estimate the genetic parameters using a multivariate animal model with the appropriate fixed effects and permanent environmental effect of the dam. Heritabilities of the BMM ranged from 0.04 to 0.25 and the genetic correlation of the BMM with body weight and breast yield ranged from -0.06 to 0.41. Here we highlight that the genetic variance of BMM accounts for a low proportion of the phenotypic variance and the BMM have a low genetic relationship with performance traits. The large contribution of residual variance to the phenotypic variance for the BBM was >71.5% which indicates the importance of the non-genetic effects on BMM. The data presented also show that the moderate to low genetic influence for the development of BMM can be used, through balanced selection, to reduce the myopathy incidence in the long term. The impact of genetic selection against BMM was tested empirically by comparing the incidence of WB and % breast yield of a commercial broiler with a high generation (HG) broiler. The HG broiler used represents 2 years of genetic improvement compared to the commercial broiler; the HG broiler had an 18.4% relative decrease in WB and a 1.02% relative increase in breast yield compared to the commercial broiler. This paper describes the relationship between the genetic and non-genetic factors influencing BMM highlighting the importance of understanding the non-genetic effects on myopathy incidence. It also shows that the genetic component of BMM can be reduced whilst at the same time improving breast yield as part of balanced breeding goals.

Highly multiplexed quantitative PCR-based platform for evaluation of chicken immune responses.

To address the need for sensitive high-throughput assays to analyse avian innate and adaptive immune responses, we developed and validated a highly multiplexed qPCR 96.96 Fluidigm Dynamic Array to analyse the transcription of chicken immune-related genes. This microfluidic system permits the simultaneous analysis of expression of 96 transcripts in 96 samples in 6 nanolitre reactions and the 9,216 reactions are ready for interpretation immediately. A panel of 89 genes was selected from an RNA-seq analysis of the transcriptional response of chicken macrophages, dendritic cells and heterophils to agonists of innate immunity and from published transcriptome data. Assays were confirmed to be highly specific by amplicon sequencing and melting curve analysis and the reverse transcription and preamplification steps were optimised. The array was applied to RNA of various tissues from a commercial line of broiler chickens housed at two different levels of biosecurity. Gut-associated lymphoid tissues, bursa, spleen and peripheral blood leukocytes were isolated and transcript levels for immune-related genes were defined. The results identified blood cells as a potentially reliable indicator of immune responses among all the tissues tested with the highest number of genes significantly differentially transcribed between birds housed under varying biosecurity levels. Conventional qPCR analysis of three differentially transcribed genes confirmed the results from the multiplex qPCR array. A highly multiplexed qPCR-based platform for evaluation of chicken immune responses has been optimised and validated using samples from commercial chickens. Apart from applications in selective breeding programmes, the array could be used to analyse the complex interplay between the avian immune system and pathogens by including pathogen-specific probes, to screen vaccine responses, and as a predictive tool for immune robustness.

Does selection for growth rate in broilers affect their resistance and tolerance to Eimeria maxima?

Chickens exhibit varied responses to infection with Eimeria parasites. We hypothesise that broilers selected for increased growth rate will show lower resistance and tolerance to a coccidian challenge. 288 chickens of fast (F) or slow (S) growing lines were inoculated with 0 (control), 2500 (low-dose), or 7000 (high-dose) sporulated E. maxima oocysts at 13 days of age in two consecutive rounds. Gain and Intake were measured daily and their values relative to BW at the point of infection were calculated over the pre-patent (days 1-4 post-infection), acute (d5-8 pi), and recovery (d9-12 pi) phases of infection to assess the impact of infection. Levels of plasma carotenoids, vitamins E and A, long bone mineralisation, caecal microbiota diversity indices, and histological measurements were assessed at the acute (d6 pi) and recovery stage (d13 pi). In addition, we measured the levels of nitric oxide metabolites and the number of parasite genome copies in the jejunumat d6pi. In absolute terms F birds grew 1.42 times faster than S birds when not infected. Infection significantly reduced relative daily gain and intake (P < 0.001), with the effects being most pronounced during the acute phase (P < 0.001). Levels of all metabolites were significantly decreased, apart from NO which increased (P < 0.001) in response to infection on d6pi, and were accompanied by changes in histomorphometric features and the presence of E. maxima genome copies in infected birds, which persisted to d13pi. Furthermore, infection reduced tibia and femur mineralisation, which also persisted to d13pi. Reductions in measured variables were mostly independent of dose size, as was the level of parasite replication. The impact of infection was similar for S and F-line birds for all measured parameters, and there were no significant interactions between line x dose size on any of these parameters. In conclusion, our results suggest that line differences in productive performance do not influence host responses to coccidiosis when offered nutrient adequate diets.

Colonization of a commercial broiler line by Campylobacter is under limited genetic control and does not significantly impair performance or intestinal health.

Campylobacter is the leading bacterial cause of foodborne diarrheal illness in humans and source attribution studies unequivocally identify handling or consumption of poultry meat as a key risk factor. Campylobacter colonizes the avian intestines in high numbers and rapidly spreads within flocks. A need therefore exists to devise strategies to reduce Campylobacter populations in poultry flocks. There has been a great deal of research aiming to understand the epidemiology and transmission characteristics of Campylobacter in poultry as a means to reduce carriage rates in poultry and reduce infection in humans. One potential strategy for control is the genetic selection of poultry for increased resistance to colonization by Campylobacter. The potential for genetic control of colonization has been demonstrated in inbred populations following experimental challenge with Campylobacter where quantitative trait loci associated with resistance have been identified. Currently in the literature there is no information of the genetic basis of Campylobacter colonization in commercial broiler lines and it is unknown whether these QTL are found in commercial broiler lines. The aim of this study was to estimate genetic parameters associated with Campylobacter load and genetic correlations with gut health and production traits following natural exposure of broiler chickens to Campylobacter.The results from the analysis show a low but significant heritability estimate (0.095 ± 0.037) for Campylobacter load which indicates a limited genetic basis and that non-genetic factors have a greater influence on the level of Campylobacter found in the broiler chicken.Furthermore, through examination of macroscopic intestinal health and absorptive capacity, our study indicated that Campylobacter has no detrimental effects on intestinal health and bird growth following natural exposure in the broiler line under study. These data indicate that whilst there is a genetic component to Campylobacter colonization worthy of further investigation, there is a large proportion of phenotypic variance under the influence of non-genetic effects. As such the control of Campylobacter will require understanding and manipulation of non-genetic host and environmental factors.

The genetic basis of pectoralis major myopathies in modern broiler chicken lines.

This is the first report providing estimates of the genetic basis of breast muscle myopathies (BMM) and their relationship with growth and yield in broiler chickens. In addition, this paper addresses the hypothesis that genetic selection for increase breast yield has contributed to the onset of BMM. Data were analyzed from ongoing recording of BMM within the Aviagen breeding program. This study focused on three BMM: deep pectoral myopathy (DPM; binary trait), white striping (WS; 4 categories) and wooden breast (WB; 3 categories). Data from two purebred commercial broiler lines (A and B) were utilized providing greater than 40,000 meat quality records per line. The difference in selection history between these two lines has resulted in contrasting breast yield (BY): 29% for Line A and 21% for Line B. Data were analyzed to estimate genetic parameters using a multivariate animal model including six traits: body weight (BW), processing body weight (PW), BY, DPM, WB, and WS, in addition to the appropriate fixed effects and permanent environmental effect of the dam. Results indicate similar patterns of heritability and genetic correlations for the two lines. Heritabilities (h2) of BW, PW and BY ranged from 0.271-0.418; for DPM and WB h2<0.1; and for WS h2≤0.338. Genetic correlations between the BMM and BW, PW, or BY were ≤0.132 in Line A and ≤0.248 in Line B. This paper demonstrates the polygenic nature of these traits and the low genetic relationships with BW, PW, and BY, which facilitates genetic improvement across all traits in a balanced breeding program. It also highlights the importance of understanding the environmental and/or management factors that contribute greater than 65% of the variance in the incidence of white striping of breast muscle and more than 90% of the variance of the incidence of wooden breast and deep pectoral myopathy in broiler chickens.

Campylobacter colonization and proliferation in the broiler chicken upon natural field challenge is not affected by the bird growth rate or breed.

The zoonotic association between Campylobacter bacteria in poultry and humans has been characterized by decades of research which has attempted to elucidate the epidemiology of this complex relationship and to reduce carriage within poultry. While much work has focused on the mechanisms facilitating its success in contaminating chicken flocks (and other animal hosts), it remains difficult to consistently exclude Campylobacter under field conditions. Within the United Kingdom poultry industry, various bird genotypes with widely varying growth rates are available to meet market needs and consumer preferences. However, little is known about whether any differences in Campylobacter carriage exist across this modern broiler range. The aim of this study was to establish if a relationship exists between growth rate or breed and cecal Campylobacter concentration after natural commercial flock Campylobacter challenge. In one investigation, four pure line genotypes of various growth rates were grown together, while in the second, eight different commercial broiler genotypes were grown individually. In both studies, the Campylobacter concentration was measured in the ceca at 42 days of age, revealing no significant difference in cecal load between birds of different genotypes both in mixed- and single-genotype pens. This is important from a public health perspective and suggests that other underlying reasons beyond genotype are likely to control and affect Campylobacter colonization within chickens. Further studies to gain a better understanding of colonization dynamics and subsequent proliferation are needed, as are novel approaches to reduce the burden in poultry.

The effect of helical computed tomography on diagnostic and treatment strategies in patients with suspected pulmonary embolism.

BACKGROUND: Helical computed tomography (CT) has been proposed as a first-line test for the diagnosis of pulmonary embolism. How the test affects the diagnostic evaluation of patients with suspected pulmonary embolism is unknown. METHODS: We examined a cohort of 360 patients evaluated for pulmonary embolism at a teaching hospital in the 4 years following the introduction of the helical CT scan. We collected patient demographic and clinical data to calculate the pretest likelihood of pulmonary embolism; we then read the test results and determined rates of further testing and treatment for pulmonary embolism. RESULTS: After the helical CT scan became available, the number of patients referred for pulmonary embolism testing increased markedly from 170 to 624 total evaluations during 1997 to 2000 (P <0.01). This rise was due to increased use of the helical CT scan (9% to 83% of evaluations, P <0.01) as the use of ventilation-perfusion scanning (79% to 17%, P = 0.03) and pulmonary angiography (12% to <1%, P <0.01) fell. There was no change in the pre-test likelihood of disease over time, but the percentage of scans that were positive for pulmonary embolism rose (14% to 32%, P =0.02). Clinicians treated all patients who had a positive CT scan, but became less likely over time to order further testing for patients who had a negative scan (30% to 12%, P = 0.02). CONCLUSION: At this academic medical center, introduction of the helical CT scan had a profound effect on the evaluation of pulmonary embolism, resulting in more frequent use of the CT scan, and more frequent diagnosis and treatment of pulmonary embolism, despite no change in the pretest probability of disease. Future studies should confirm our findings and determine whether increased detection of pulmonary emboli results in improved outcomes.

Helical CT pulmonary angiography predictors of in-hospital morbidity and mortality in patients with acute pulmonary embolism.

PURPOSE: To determine if CT variables predict in-hospital morbidity and mortality in patients with pulmonary embolism (PE). MATERIALS AND METHODS: CT scans and charts of 173 patients with CT scans positive for PE were reviewed. CT scans were reviewed for leftward ventricular septal bowing, increased right ventricle (RV) to left ventricle (LV) diameter ratio, clot burden, increased pulmonary artery to aorta diameter ratio, and oligemia. Charts were reviewed for severe morbidity and mortality outcomes: death from pulmonary emboli or any cause, and cardiac arrest. Charts were also reviewed for milder morbidity outcomes: intubation, vasopressor use, or admission to an intensive care unit (ICU) and for multiple comorbidities. RESULTS: No CT predictor was significantly associated with severe morbidity or mortality outcomes. Ventricular septal bowing and increased RV/LV diameter ratio were both associated with subsequent admission to an ICU (P = 0.004 and P = 0.025, respectively). Oligemia (either lung) was associated with subsequent intubation; right lung oligemia was associated with the subsequent use of vasopressors. After controlling for history of congestive heart failure, ischemic heart disease, and pulmonary disease, both septal bowing and an increased RV/LV diameter ratio remained associated with admission to an ICU. CONCLUSION: No CT variables predicted severe in-hospital morbidity and mortality (death from pulmonary embolism, death from any cause, or cardiac arrest) in patients with PE. However, ventricular septal bowing and increased RV/LV diameter ratio were both strongly predictive of less severe morbidity, namely, subsequent ICU admission, and oligemia was associated with subsequent intubation and vasopressor use.

Are chest radiographs routinely necessary following thoracostomy tube removal?

OBJECTIVE: Chest radiographs (CXRs) are routinely obtained at many institutions in all pediatric patients following thoracostomy tube removal to search for pneumothorax (PTX). To aid in evaluating the necessity of this practice, this study investigates whether clinical signs and symptoms may be a sensitive predictor of PTX in such patients. MATERIALS AND METHODS: Reports from CXRs obtained following chest tube removal in all pediatric patients (374 patients) who underwent cardiac surgery with chest tube placement over 1 year were reviewed. For cases with reported PTX, the PTX was quantified and chart review was performed to assess whether signs and symptoms of PTX preceded the CXR result. RESULTS: Fifty-one of 374 children (13.6%) had a radiographically defined PTX within 6 h after thoracostomy tube removal. The PTX was large (>40%) in 2 children, moderate (20-40%) in 5 children, and small (<20%) in 44 children. Symptoms (dyspnea, tachypnea, respiratory distress) or signs (increased oxygen requirement, worsening arterial blood gas and/or hypotension) of respiratory distress were present at the time of the initial CXR in six of seven patients, who later underwent a major clinical intervention, and in one patient who did not. Major clinical interventions were performed in all patients with a large PTX, four of five patients with a moderate PTX, and one patient with a small PTX that later enlarged. CONCLUSIONS: Clinical signs and symptoms identified nearly all patients with significant pneumothoraces. Future prospective investigations may examine reserving chest radiography following chest tube removal for select groups, such as symptomatic patients or those with tenuous cardiovascular status.