Iron limitation induces motility in uropathogenic E. coli CFT073 partially through action of LpdA.

More than half of women will experience a urinary tract infection (UTI) with most cases caused by uropathogenic Escherichia coli (UPEC). Bacterial swimming motility enhances UPEC pathogenicity, resulting in more severe disease outcomes including kidney infection. Surprisingly, the connection between motility and iron limitation is mostly unexplored despite the lack of free iron available in the host. We sought to investigate a potential connection between iron restriction and regulation of motility in UPEC. We cultured E. coli CFT073, a prototypical UPEC strain, under iron limitation and observed that CFT073 had elevated fliC (flagella) promoter activity, and this iron-specific response was repressed by the addition of exogenous iron. We confirmed increased flagellar expression in CFT073 by measuring fliC transcript, FliC protein, and surface-expressed flagella under iron-limited conditions. Interestingly, known motility regulator flhDC did not have altered transcription under these conditions. To define the regulatory mechanism of this response, we constructed single knockouts of eight master regulators and found the iron-regulated response was lost in crp, arcA, and fis mutants. Thus, we focused on the five genes regulated by all three regulators. Of the five genes knocked out, the iron-regulated motility response was most strongly dysregulated in the lpdA mutant, which also resulted in significantly lowered fitness in the murine model of ascending UTI, both against the WT and a non-motile fliC mutant. Collectively, we demonstrated that iron-mediated motility in CFT073 is partially regulated by lpdA, which contributes to the understanding of how uropathogens differentially regulate motility mechanisms in the iron-restricted host. IMPORTANCE: Urinary tract infections (UTIs) are ubiquitous and responsible for over five billion dollars in associated health care costs annually. Both iron acquisition and motility are highly studied virulence factors associated with uropathogenic Escherichia coli (UPEC), the main causative agent of uncomplicated UTI. This work is innovative by providing mechanistic insight into the synergistic relationship between these two critical virulence properties. Here, we demonstrate that iron limitation has pleiotropic effects with consequences that extend beyond metabolism and impact other virulence mechanisms. Indeed, targeting iron acquisition as a therapy may lead to an undesirable enhancement of UPEC pathogenesis through increased motility. It is vital to understand the full breadth of UPEC pathogenesis to adequately respond to this common infection, especially with the increase of antibiotic-resistant pathogens.

Altered motility in response to iron-limitation is regulated by lpdA in uropathogenic E. coli CFT073.

More than half of all women will experience a urinary tract infection (UTI) in their lifetime with most cases caused by uropathogenic Escherichia coli (UPEC). Bacterial motility enhances UPEC pathogenicity, resulting in more severe disease outcomes including kidney infection. Surprisingly, the connection between motility and iron limitation is mostly unexplored, despite the lack of free iron available in the host. Therefore, we sought to explore the potential connection between iron restriction and regulation of motility in UPEC. We cultured E. coli CFT073, a prototypical UPEC strain, in media containing an iron chelator. Under iron limitation, CFT073 had elevated fliC (flagella) promoter activity, driving motility on the leading edge of the colony. Furthermore, this iron-specific response was repressed by the addition of exogenous iron. We confirmed increased flagella expression in CFT073 by measuring fliC transcript, FliC protein, and surface-expressed flagella under iron-limited conditions. To define the regulatory mechanism, we constructed single knockouts of eight master regulators. The iron-regulated response was lost in crp, arcA, and fis mutants. Thus, we focused on the five genes regulated by all three transcription factors. Of the five genes knocked out, the iron-regulated motility response was most strongly dysregulated in an lpdA mutant, which also resulted in significantly lowered fitness in the murine model of ascending UTI. Collectively, we demonstrated that iron-mediated motility in CFT073 is regulated by lpdA , which contributes to the understanding of how uropathogens differentially regulate motility mechanisms in the iron-restricted host. Importance: Urinary tract infections (UTIs) are ubiquitous and responsible for over five billion dollars in associated health care costs annually. Both iron acquisition and motility are highly studied virulence factors associated with uropathogenic E. coli (UPEC), the main causative agent of uncomplicated UTI. This work is innovative by providing mechanistic insight into the synergistic relationship between these two critical virulence properties. Here, we demonstrate that iron limitation has pleiotropic effects with consequences that extend beyond metabolism, and impact other virulence mechanisms. Indeed, targeting iron acquisition as a therapy may lead to an undesirable enhancement of UPEC pathogenesis through increased motility. It is vital to understand the full breadth of UPEC pathogenesis to adequately respond to this common infection, especially with the increase of antibiotic resistant pathogens.

Examination of the exposome in an animal model: The impact of high fat diet and rat strain on local and systemic immune markers following occupational welding fume exposure.

The goal of this study was to investigate the impact of multiple exposomal factors (genetics, lifestyle factors, environmental/occupational exposures) on pulmonary inflammation and corresponding alterations in local/systemic immune parameters. Accordingly, male Sprague-Dawley (SD) and Brown Norway (BN) rats were maintained on either regular (Reg) or high fat (HF) diets for 24wk. Welding fume (WF) exposure (inhalation) occurred between 7 and 12wk. Rats were euthanized at 7, 12, and 24wk to evaluate local and systemic immune markers corresponding to the baseline, exposure, and recovery phases of the study, respectively. At 7wk, HF-fed animals exhibited several immune alterations (blood leukocyte/neutrophil number, lymph node B-cell proportionality)-effects which were more pronounced in SD rats. Indices of lung injury/inflammation were elevated in all WF-exposed animals at 12wk; however, diet appeared to preferentially impact SD rats at this time point, as several inflammatory markers (lymph node cellularity, lung neutrophils) were further elevated in HF over Reg animals. Overall, SD rats exhibited the greatest capacity for recovery by 24wk. In BN rats, resolution of immune alterations was further compromised by HF diet, as many exposure-induced alterations in local/systemic immune markers were still evident in HF/WF animals at 24wk. Collectively, HF diet appeared to have a greater impact on global immune status and exposure-induced lung injury in SD rats, but a more pronounced effect on inflammation resolution in BN rats. These results illustrate the combined impact of genetic, lifestyle, and environmental factors in modulating immunological responsivity and emphasize the importance of the exposome in shaping biological responses.

Zinc hydroxychloride supplementation improves tibia bone development and intestinal health of broiler chickens.

This study was conducted to investigate the effects of zinc (Zn), as a combination of oxide (ZnO) and sulfate (ZnSO4), compared with incremental levels of zinc hydroxychloride (ZH) on tibia traits, intestinal integrity, expression of selected jejunal genes, cecal short chain fatty acids and microbial composition in broilers. Day-old male Ross 308 chicks (n = 784) were randomly allocated to seven dietary treatments, each replicated seven times with 16 chicks per replication. The dietary treatments included a negative control diet (NC) with no supplemental Zn, a positive control (PC) with 100 mg/kg supplemental Zn from an ionic bound source combination (50 mg/kg ZnO + 50 mg/kg ZnSO4), and the NC diet supplemented with one of 20, 40, 60, 80, or 100 mg/kg Zn as ZH. The diets were fed over starter (1-14 d) and grower (14-35 d) phases, with tissue and digesta samples collected from 3 birds per replicate on days 14 and 35. The results showed that dietary Zn level had a significant effect on tibia breaking strength on d 35 (P < 0.05), and tibia Zn concentration both on d 14 and d 35 (P < 0.01). Dietary Zn levels linearly (P < 0.01) increased cecal lactic acid production, increased Lactobacillus, and decreased Bacillus and total bacteria counts (P < 0.05). Inclusion of 80 and 100 mg/kg Zn as ZH tended to upregulate the expression of claudin-1 (P = 0.088) and tight junction protein-1 (P = 0.086). The results obtained in this study suggest that a non-Zn supplemented diet can negatively influence tibia development and gut microbiota composition in broiler chickens. Higher supplemental Zn in the diet alters cecal microbiota population in favor of Lactobacillus and can decrease the total bacterial load. Supplemental Zn level in the feed have the potential to manipulate the jejunal gut integrity at a molecular level.

Copper hydroxychloride is more efficacious than copper sulfate in improving broiler chicken's growth performance, both at nutritional and growth-promoting levels.

This study was designed to compare the effects of nutritional and growth-promoting levels of copper hydroxychloride (CH) with copper sulfate (CuSO4) on growth, carcass characteristics, tibia traits and mineral concentration in broilers fed a conventional wheat-soybean meal-based diet. Day-old Ross 308 male chicks (n = 864) were randomly assigned into 8 dietary treatments with 6 replicates of 18 chicks per treatment. The dietary treatments included a basal diet containing no supplemental copper (Cu) serving as the negative control (NC); basal diet supplemented with 15 or 200 mg/kg Cu as CuSO4; basal diet supplemented with either 15, 50, 100, 150, or 200 mg/kg Cu from CH. Diets were fed over the starter (day 1-14) and grower (day 14-35) phases. Birds in the NC group gained the same body weight and had similar feed conversion ratio (FCR) to birds receiving 15 mg/kg Cu as CuSO4, but birds receiving 15 mg/kg Cu as CH had a lower FCR than the NC birds (day 0-35; P < 0.05). Birds fed 200 mg/kg Cu as CH gained more weight (77 g/bird) and had a lower FCR (3.2 point) compared with those fed 200 mg/kg Cu as CuSO4 (P < 0.01). Based on broken-line regression models, the optimum inclusion level of Cu as CH in the diet for optimal body weight gain and FCR were estimated to be 109.5 and 72.3 mg/kg, respectively (P < 0.001). Carcass characteristics were not affected by dietary Cu sources or levels (P > 0.05). The highest and lowest tibia ash content were observed in birds fed diet with 150 mg/kg Cu as CH and 200 mg/kg Cu as CuSO4, respectively (P < 0.05). Supplementation with 200 mg/kg Cu as CH resulted in higher duodenal mucosa Cu content compared with the diet containing 200 mg/kg Cu as CuSO4 (P < 0.001). In conclusion, supplementation of Cu from CH was more efficacious than CuSO4 in promoting growth performance, both at nutritional and pharmacological levels.

Evaluation of the skin-sensitizing potential of gold nanoparticles and the impact of established dermal sensitivity on the pulmonary immune response to various forms of gold.

Gold nanoparticles (AuNP) are largely biocompatible; however, many studies have demonstrated their potential to modulate various immune cell functions. The potential allergenicity of AuNP remains unclear despite the recognition of gold as a common contact allergen. In these studies, AuNP (29 nm) dermal sensitization potential was assessed via Local Lymph Node Assay (LLNA). Soluble gold (III) chloride (AuCl3) caused lymph node (LN) expansion (SI 10.9), whereas bulk particles (Au, 942 nm) and AuNP did not. Next, the pulmonary immune effects of AuNP (10, 30, 90 µg) were assessed 1, 4, and 8 days post-aspiration. All markers of lung injury and inflammation remained unaltered, but a dose-responsive increase in LN size was observed. Finally, mice were dermally-sensitized to AuCl3 then aspirated once, twice, or three times with Au or AuNP in doses normalized for mass or surface area (SA) to assess the impact of existing contact sensitivity to gold on lung immune responses. Sensitized animals exhibited enhanced responsivity to the metal, wherein subsequent immune alterations were largely conserved with respect to dose SA. The greatest increase in bronchoalveolar lavage (BAL) lymphocyte number was observed in the high dose group - simultaneous to preferential expansion of BAL/LN CD8+ T-cells. Comparatively, the lower SA-based doses of Au/AuNP caused more modest elevations in BAL lymphocyte influx (predominantly CD4+ phenotype), exposure-dependent increases in serum IgE, and selective expansion/activation of LN CD4+ T-cells and B-cells. Overall, these findings suggest that AuNP are unlikely to cause sensitization; however, established contact sensitivity to gold may increase immune responsivity following pulmonary AuNP exposure.

Evaluation of an intervention targeting loneliness and isolation for older people in North Wales.

AIMS: Age Cymru Gwynedd and Môn provided a service, Cadwyn Môn, which aimed to reduce loneliness and isolation of older people living on Anglesey, North Wales. It involved working with individuals to identify goals, beginning on a one-to-one basis, with the aim of integrating into a wider social setting. The aim of this study was evaluate the impact of Cadwyn Môn on those who received the service. METHODS: The evaluation adopted a mixed-methods design. Standardized measures of loneliness, social isolation and well-being were obtained at baseline and follow-up time points, and a qualitative element explored client experiences further. RESULTS: Improvements were observed in all standardized measures. Qualitative interviews also revealed important psychological and lifestyle changes. CONCLUSIONS: Given the anticipated rise in loneliness and social isolation alongside an ageing population, this evaluation contributes to understanding the qualities required for interventions to be effective. The findings provide support for the importance of developing individualized interventions that aim to gradually re-integrate people into their communities.

Effects of pulmonary exposure to chemically-distinct welding fumes on neuroendocrine markers of toxicity.

Exposure to welding fumes may result in disorders of the pulmonary, cardiovascular, and reproductive systems. Welders are also at a greater risk of developing symptoms similar to those seen in individuals with idiopathic Parkinson's disease. In welders, there are studies that suggest that alterations in circulating prolactin concentrations may be indicative of injury to the dopamine (DA) neurons in the substantia nigra. The goal of these studies was to use an established model of welding particulate exposure to mimic the effects of welding fume inhalation on reproductive functions. Since previous investigators suggested that changes in circulating prolactin may be an early marker of DA neuron injury, movement disorders, and reproductive dysfunction, prolactin, hypothalamic tyrosine hydroxylase (TH) levels (a marker of DA synthesis), and other measures of hypothalamic-pituitary-gonadal (HPG) function were measured after repetitive instillation of welding fume particulates generated by flux core arc-hard surfacing (FCA-HS), manual metal arc-hard surfacing (MMA-HS) or gas metal arc-mild steel (GMA-MS) welding, or manganese chloride (MnCl2). Exposure to welding fume particulate resulted in the accumulation of various metals in the pituitary and testes of rats, along with changes in hypothalamic TH and serum prolactin levels. Exposure to particulates with high concentrations of soluble manganese (Mn) appeared to exert the greatest influence on TH activity levels and serum prolactin concentrations. Thus, circulating prolactin levels may serve as a biomarker for welding fume/Mn-induced neurotoxicity. Other reproductive measures were collected, and these data were consistent with epidemiological findings that prolactin and testosterone may serve as biomarkers of welding particulate induced DA neuron and reproductive dysfunction.

Egg production and egg quality in free-range laying hens housed at different outdoor stocking densities.

Free-range laying hen systems are increasing in number within Australia. Variation in outdoor stocking densities has led to development of a national information standard on free-range egg labeling, including setting a maximum density of 10,000 hens per hectare. However, there are few data on the impacts of differing outdoor densities on production and egg quality. ISA Brown hens in small (150 hens) flocks were housed in identical indoor pens, each with access (from 21 weeks) to different sized ranges simulating one of three outdoor stocking densities (2 replicates each: 2,000 hens/hectare (ha), 10,000 hens/ha, 20,000 hens/ha). Hen-day production was tracked from 21 through 35 weeks with eggs visually graded daily for external deformities. All eggs laid on one day were weighed each week. Eggs were collected from each pen at 25, 30, and 36 weeks and analyzed for egg quality. There were no effects of outdoor stocking density on average hen-day percentage production (P = 0.67), egg weight (P = 0.09), percentages of deformed eggs (P = 0.30), shell reflectivity (P = 0.74), shell breaking strength (P = 0.07), shell deformation (P = 0.83), or shell thickness (P = 0.24). Eggs from hens in the highest density had the highest percentage shell weight (P = 0.004) and eggs from the lowest density had the highest yolk color score (P < 0.001). The amount of cuticle present did not differ between densities (P = 0.95) but some aspects of shell colors (P ≤ 0.01) and location of protoporphyrin IX (P = 0.046) varied. Hen age affected the majority of measurements. Stocking density differences may be related to hen diet as previous radio-frequency identification tracking of individual hens in these flocks showed birds used the range for longer in the lowest density and the least in the highest density, including depleting the range of vegetation sooner in the smaller ranges. An additional study assessing the relationship between individual hen range use, nutrition, and egg quality is warranted.

The challenge of tetradic relationships in medically interpreted pediatric primary care visits: A descriptive study of communication practices.

OBJECTIVE: To examine spoken interactions between pediatricians and community-based interpreters speaking with adolescents and parents with Limited English proficiency (LEP) in primary care to identify the challenges of interpreting in a four-person or tetradic visit, its sources of co-constructed errors, and specific practices for educational intervention. METHODS: As part of a larger study of vaccine decision-making at six clinical sites in two states, this descriptive study used discourse analysis to examine 20 routine primary care visits in a Latino Clinic in interactions between adolescents, parents, community-based interpreters, and pediatricians. Specific patterns of communication practices were identified that contributed to inaccuracies in medical interpretation RESULTS: Practices needing improvement were tallied for simple frequencies and included: omissions; false fluency; substitutions; editorializing; added clarification, information, or questions; medical terminology; extra explanation to mother; and, cultural additions. Of these speaking practices, omissions were the most common (123 out of 292 total) and the most affected by pediatricians. CONCLUSION: The dynamics of both pediatricians and interpreters contributed to identification of areas for improvement, with more adolescent participation in bilingual than monolingual visits. PRACTICE IMPLICATIONS: These observations provide opportunities for mapping a communication skills training intervention based on observations for future testing of an evidence-based curriculum.

Eggshell color in brown-egg laying hens - a review.

The major pigment in eggshells of brown-egg laying hens is protoporphyrin IX, but traces of biliverdin and its zinc chelates are also present. The pigment appears to be synthesized in the shell gland. The protoporphyrin IX synthetic pathway is well defined, but precisely where and how it is synthesized in the shell gland of the brown-egg laying hen is still ambiguous. The pigment is deposited onto all shell layers including the shell membranes, but most of it is concentrated in the outermost layer of the calcareous shell and in the cuticle. Recently, the genes that are involved in pigment synthesis have been identified, but the genetic control of synthesis and deposition of brown pigment in the commercial laying hen is not fully understood. The brown coloration of the shell is an important shell quality parameter and has a positive influence on consumer preference. The extent of pigment deposition is influenced by the housing system, hen age, hen strain, diet, stressors, and certain diseases such as infectious bronchitis. In this article, the physiological and biochemical characteristics of the brown pigment in commercial brown-egg layers are reviewed in relation to its various functions in the poultry industry.

Eggshell penetration by Salmonella Typhimurium in table eggs: Examination of underlying eggshell structures by micro-computed tomography and scanning electron microscopy.

Horizontal infection of table eggs by food borne, human infection causative agents such as Salmonella is a serious concern for consumers and industry. In this study, we investigated the relationship between eggshell translucency, mammillary layer abnormalities and pore structure using Computed Tomography (CT) and scanning electron microscopy. The effects of eggshell pore structure, size and number on Salmonella Typhimurium penetration was also investigated. The eggs were infected with S. Typhimurium and were incubated at 37°C for 3 or 6days. Micro CT results comparing shell features to shell translucency found that there was a significantly increased incidence of externally branching pores found in the high translucency score eggshell group, and more straight pores found in the low translucency score group. Different pore structures, the total number of pores and the shell thickness do not appear to play a role in the horizontal infection of eggs by the S. Typhimurium strain used in this study. While it is likely that the presence of shell pores is responsible for shell penetration, other unknown shell factors must also play a role, and eggshells with a higher incidence of shell pores are not penetrated at a higher rate.

The location of protoporphyrin in the eggshell of brown-shelled eggs.

Protoporphyrin has been identified as the main eggshell pigment in brown-shelled eggs. However, there has been some uncertainty concerning the distribution of the pigment within the shell (and cuticle) in brown-shelled eggs. Most previous studies have suggested that the bulk of the shell pigment is deposited in the cuticle of the shell. The present study measured the levels of protoporphyrin in intact eggshells and in shells from which the cuticle had been removed, using eggs from flocks at 3 different ages. This enabled the calculation of the relative amount of protoporphyrin in the calcareous eggshell and the cuticle layer of the eggshell. The majority of the protoporphyrin pigment was located in the calcareous part of the eggshell (80-87%) with a minority contained within the cuticle (13-20%). These findings suggest that studies focused on maintenance of shell color in brown-shelled eggs need to consider the stage of egg formation at which the reduction in pigment deposition is occurring.

Effects of egg shell quality and washing on Salmonella Infantis penetration.

The vast majority of eggs in Australia are washed prior to packing to remove dirt and fecal material and to reduce the microbial contamination of the egg shell. The egg contents can be an ideal growth medium for microorganisms which can result in human illness if eggs are stored improperly and eaten raw or undercooked, and it is estimated that egg-related salmonellosis is costing Australia $44 million per year. Egg shell characteristics such as shell thickness, amount of cuticle present, and thickness of individual egg shell layers can affect the ease with which bacteria can penetrate the egg shell and washing could partially or completely remove the cuticle layer. The current study was conducted to investigate the effects of egg washing on cuticle cover and effects of egg shell quality and cuticle cover on Salmonella Infantis penetration of the egg shell. A higher incidence of unfavorable ultrastructural variables of the mammillary layer such as late fusion, type B bodies, type A bodies, poor cap quality, alignment, depression, erosion and cubics were recorded in Salmonella penetrated areas of egg shells. The influence of egg washing on the ability of Salmonella Infantis on the egg shell surface to enter the egg internal contents was also investigated using culture-based agar egg penetration and real-time qPCR based experiments. The results from the current study indicate that washing affected cuticle cover. There were no significant differences in Salmonella Infantis penetration of washed or unwashed eggs. Egg shell translucency may have effects on Salmonella Infantis penetration of the egg shell. The qPCR assay was more sensitive for detection of Salmonella Infantis from egg shell wash and internal contents than traditional microbiological methods. The agar egg and whole egg inoculation experiments indicated that Salmonella Infantis penetrated the egg shells. Egg washing not only can be highly effective at removing Salmonella Infantis from the egg shell surface, but also allows subsequent trans-shell and trans-membrane penetration into the egg. Consequently, it is important to prevent recontamination of the egg after washing.

Survey of Enterobacteriaceae contamination of table eggs collected from layer flocks in Australia.

In the present study, eggs from commercial caged layer flocks at different stages of lay in Australia were collected. Enterobacteriaceae populations from eggshell surface and eggshell pore were enumerated and these populations characterized using API® Rapid 20E strips. The eggshell surface, eggshell pore and egg internal content samples were also processed for the isolation of Salmonella and these isolates were tested for the presence or absence of several virulence genes (prgH, sopB, spiC, orfL, invA, sifA, sitC, misL). Results indicated that there was no significant difference in total Enterobacteriaceae count on the eggs of the flock from early, mid or late lay flocks. Enterobacteriaceae isolates were of 11 different genera which included: Cedecea, Citrobacter, Enterobacter, Escherichia, Klebsiella, Kluyvera, Leclercia, Pantoea, Salmonella, Serratia and Yersinia. Out of all 153 identified Enterobacteriaceae isolates, the Escherichia genus was reported most frequently (60.78%). Results also indicated that overall there were 4.51% (14/310) Salmonella positive pooled samples. In this study, 14 Salmonella strains were isolated, serotyping confirmed that 12 out of them were Salmonella Infantis and the 2 others were Salmonella enterica subsp. enterica serovar 4,12:d: Polymerase chain reaction results indicated that all Salmonella Infantis isolates harboured invA, misL, orfL, prgH, sifA, sitC, sopB and spiC genes which suggests that Salmonella Infantis strains isolated from eggshell surface may have the capacity to invade and survive in macrophages.

Recovery of Salmonella from eggshell wash, eggshell crush, and egg internal contents of unwashed commercial shell eggs in Australia.

The experiment was conducted to study the prevalence of Salmonella spp. on the eggshell surface, eggshell membranes or pores, and in egg internal contents from unwashed eggs collected from commercial caged layer farms in Australia. Eggshell rinsate, shell crush, and egg internal contents (yolk and albumen) of eggs were processed for Salmonella spp. Salmonella Infantis and Salmonella subspecies 1, serotype 4,12:d were isolated from the eggshell surface. Salmonella spp. were not isolated from any eggshell crush or egg internal contents. It would appear that the occurrence of Salmonella in the Australian egg industry is low.

Prevalence of antibodies to Mycoplasma synoviae in laying hens and possible effects on egg shell quality.

Mycoplasma synoviae (M. synoviae) can cause respiratory disease, synovitis, or result in a silent infection in chickens and turkeys. The importance of M. synoviae is well established in broilers but only a few studies have been conducted in layers. In the present study, the prevalence of M. synoviae in commercial layer flocks was estimated using ELISA. For this study, 19 commercial layer flocks were selected randomly from New South Wales and Queensland region of Australia from producers who were willing to participate in the survey. Sixty eggs per flocks were randomly collected, out of these 30 eggs were used for ELISA and remaining 30 eggs were used to estimate various egg shell quality parameters. Subsequently, association between the serological status of eggs for M. synoviae and egg shell quality was studied. In the flocks under study, seroprevalence of M. synoviae was found to be high at 69% (95% confidence interval (CI)=41.3-89.0). Statistical analysis showed an association between serological status for M. synoviae and egg quality parameters such as translucency, shell breaking strength, % shell reflectivity and shell deformation. On the other hand, there was no significant association between serological status for M. synoviae and other egg quality parameters such egg weight, egg shell weight, % egg shell or shell thickness.

Extraesophageal gastroesophageal reflux disease (GERD) symptoms are not more frequently associated with proximal esophageal reflux than typical GERD symptoms.

Extraesophageal (EE) symptoms such as cough and throat clearing are common in patients referred for reflux testing, but are less commonly associated with gastroesophageal reflux disease (GERD). Patients with reflux associated EE symptoms often lack typical GERD symptoms of heartburn and regurgitation. Our aim was to compare the frequency of proximal esophageal reflux between esophageal (typical) symptoms and EE (atypical) symptoms. Combined multichannel intraluminal impedance-pH (MII-pH) tracings were blinded by an investigator so that symptom markers were relabeled with a number without disclosure of symptom type. We selected 40 patients with at least five reflux-related symptom events for one of four symptoms (heartburn, regurgitation, cough, or throat clearing). A blinded investigator analyzed all 200 reflux episodes, reporting the proximal esophageal extent of the reflux for all symptoms. The percentage of symptom-related reflux extending proximally to 17 cm above the LES was similar among all four symptom types. At least 50% of all symptoms were associated with proximal esophageal reflux to 17 cm, with regurgitation having the highest frequency at 60%. Our data indicate that EE symptoms are not more frequently associated with proximal esophageal reflux than typical esophageal symptoms.

Recovery of Salmonella and Escherichia coli from commercial egg shells and effect of translucency on bacterial penetration in eggs.

This experiment was conducted to study the prevalence of Salmonella and Escherichia coli (E. coli). from the surface of egg shells, egg shell membranes or pores, and internal contents from unwashed eggs collected from commercial caged layer farms in Australia. Egg shell swabs, shell crush and egg internal contents (yolk and albumen) of an individual egg were processed for bacteriological examination. Salmonella spp. were not detected from any of the egg shell surfaces, egg shell crush or egg internal contents. Thirty five E. coli isolates were isolated from the egg shell surface. Ten E. coli strains were also isolated from shell crush. However, the internal contents of eggs appeared to be sterile. Polymerase chain reaction was performed on forty-five E. coli isolates using primers for heat stable enterotoxin genes A and B (STa and STb) and also for colicin V gene (cvaC). STa gene was detected in four E. coli isolates isolated from egg shell surfaces. All the E. coli isolates were negative for STb and cvaC genes. These data provide useful information regarding the prevalence of virulent E. coli and Salmonella spp. on and in unwashed eggs collected from layer farms. These data also suggest that unwashed eggs collected from caged layer farms are unlikely to be sources of Salmonella outbreaks. Egg shell translucency could be due to changes in the mammillary layer and mamillary cores during the early phases of egg shell formation and has the potential to increase the incidence of microcracks in egg shells, and hence, may be responsible for bacterial penetration. There was a significant correlation between egg shell translucency and egg shell penetration by Salmonella Infantis and E coli. Both strains of bacteria were able to penetrate the translucent egg shells even at very low doses. The penetration, however, was hindered in both translucent and non translucent eggs at 4 degrees C, as compared with room temperature which highlights the importance of storage of eggs at refrigerated temperatures.

Detection of infectious bronchitis virus strain N1/88 from the oviduct and feces of experimentally infected vaccinated and unvaccinated hens.

Hens were vaccinated during the rearing phase with infectious bronchitis virus (IBV) vaccines commercially available in Australia (Vic S and A3) or left unvaccinated and then challenged with the N1/88 strain of IBV at 30 wk of age. Oviduct and fecal samples were collected at regular intervals after N1/88 challenge. A locked nucleic acid probe-based reverse transcription real-time PCR test was designed and used to detect the IBV strain N1/88 from the oviduct and feces of unvaccinated and vaccinated laying hens. Using a recombinant plasmid standard, the detection limit of the reaction was found to be 100 copies and independent assay runs showed reproducible threshold cycle values. Viral RNA was detected in the oviduct of 12 unvaccinated then challenged hens and viral RNA increased sharply on d 10 and 12 postinfection (p.i.). By contrast, among the hens in the vaccinated group, N1/88 was detectable only in the oviduct of 2 hens at 8 and 12 d p.i. N1/88 challenge. Viral RNA was detected in feces of 2 unvaccinated hens up to 4 wk p.i. and in 1 vaccinated hen up to 3 wk p.i. This shows that rearing phase vaccination lowers the total viral RNA of the strain N1/88, even though this strain shows considerable antigenic and genetic variation from the vaccine strain. This new test will be useful for the rapid identification of the N1/88 strain of IBV from oviduct and fecal samples.