Research Note: Effect of a phlorotannin extract of the brown seaweed Ascophyllum nodosum as a potential control strategy against Campylobacter in broilers.

Poultry is generally recognized as the main source of human campylobacteriosis and Campylobacter is highly prevalent at the farm level. To reduce the relative risk of human campylobacteriosis attributable to broiler meat, it is necessary to reduce Campylobacter loads in broiler ceca but to date, no effective, reliable and practical strategy is available. The marine environment is a rich source of original natural compounds exhibiting different biological activities. The objective of this study was to test a phlorotannin extract of the brown seaweed Ascophyllum nodosum as a potential control strategy against Campylobacter in broilers. Bactericidal activity has been demonstrated in vitro, on several Campylobacter spp. strains at a range of 0.06 to 0.47 mg/mL. Therefore, an in vivo trial in experimental facilities was performed to evaluate addition of 0.2% (w/w) of an A. nodosum extract to feed distributed at the end of rearing from day 31 to day 35, and to assess the effect on artificial Campylobacter jejuni colonization. No statistical differences in Campylobacter enumeration were observed between the treated and control groups. Another trial was performed in a commercial broiler flock. Feed containing the extract at 0.2% (w/w) (2 kg/t) was distributed during the last 5 days of rearing (day 33-day 38). No significant effects on Campylobacter colonization and on growth parameters were observed compared to the control group. Additional studies are needed to assess whether active polyphenols are found in the cecum.

Prevalence and genetic diversity of Campylobacter spp. in the production chain of broiler chickens in Lebanon and its association with the intestinal protozoan Blastocystis sp.

Campylobacter jejuni is recognized as the most common foodborne pathogen associated with human gastroenteritis worldwide. Broilers are frequently infected by the bacteria and are considered the main source of exposure to humans. However, despite its public health impact, no recent data are currently available in Lebanon about Campylobacter spp. in poultry and human population. Therefore, this study aimed to determine the prevalence and genetic diversity of Campylobacter spp. in 227 ceca and on 227 carcasses of broiler chickens collected in Lebanese slaughterhouses. Overall, the prevalence of Campylobacter was shown to reach 67.0% in ceca and 17.2% on carcasses of Lebanese poultry. The only 2 Campylobacter species identified were C. jejuni and C. coli, with a slightly higher prevalence of C. coli in ceca and of C. jejuni on carcasses. A high level of genetic diversity was reported among the 51 C. jejuni isolates selected, since 25 distinct profiles were identified according to the comparative genomic fingerprinting typing method based on a subset of 40 genes using the 90% similarity threshold. Predominant clusters observed in Lebanese poultry isolates were also frequently found among French human clinical cases, highlighting that broiler chickens represent a potential reservoir for human campylobacteriosis. In addition, a significantly higher prevalence of Campylobacter spp. was found in slaughterhouse workers than in a cohort of hospitalized patients with no contact with poultry, confirming that contaminated broiler chickens in slaughterhouse appeared to be a non-negligible source of Campylobacter spp. transmission. Interestingly, a significant association between Campylobacter spp. and Blastocystis sp. has been observed. This correlation suggested that the presence of Campylobacter spp. would be favored when Blastocystis sp. is present and, similarly, the absence of one would favor the absence of the other. This is the first large-scale investigation focusing on the impact of Campylobacter spp. in broiler chickens in Lebanon and confirmed the need to implement prevention and control measures in the poultry production to reduce the burden of campylobacteriosis in the human population.

NLRP3 as a potentially novel biomarker for the management of osteoarthritis.

Osteoarthritis (OA) was previously thought of as 'wear and tear' as humans age, however there is increasing evidence to support an inflammatory theory. The nucleotide-binding and oligomerization domain-like receptor containing protein 3 (NLRP3) inflammasome has been implicated in the pathogenesis of a number of arthritic disorders, producing proinflammatory cytokines and degradative enzymes such as Interleukin-1 beta (IL-1ß), Tumour necrosis factor alpha (TNF-α) and Matrix metalloproteinase-3 (MMP-3) which drive cartilage degeneration and synovial inflammation. This review aims to summarise the evidence of NLRP3 involvement in OA. Currently, treatment options focus on management of the disease and to date there is no cure. The development of novel biomarkers for OA could improve diagnosis, treatment and management. Importantly, this review provides detail on the involvement of the NLRP3 inflammasome in OA pathology and how its members could act as potential biomarkers to assist clinical decisions.

Randomized control trial to test the effect of a feed additive on Campylobacter contamination in commercial broiler flocks up to slaughter.

A randomized controlled trial (RCT) was carried to evaluate the effect of a feed additive on Campylobacter contamination of broilers reared in commercial conditions. Twenty-four broiler flocks naturally contaminated with Campylobacter were enrolled in the RCT: 12 were assigned to a control group (C) fed with a conventional finishing feed from 4 weeks of age to slaughter (around 35 days), and the other group of 12 flocks (S) was fed with a finishing feed supplemented with 250 ppm of a patented feed additive (an ion-exchanged clay compound) previously proven to reduce Campylobacter contamination in broiler caeca under experimental conditions. Enumeration of Campylobacter colonies in caeca (8 per flock) was carried out following ISO standards before feed distribution and at slaughter. Before treatment, the caecal Campylobacter load tended to be lower in C flocks (7.1 ± 1.9 log CFU/g, CI95% [6.6-7.5]) than in S flocks (7.7 ± 1.0 log UFC/g, CI95% [7.5-7.9]) (p = .05). At slaughter, the bacterial load was similar in the S (7.7 ± 1.0 log CFU/g, CI95% [7.5-7.9]) and C groups (7.5 ± 1.2 log CFU/g, CI95% [7.2-7.8]) (p = .73). Therefore, the feed additive had no significant effect on the caecal Campylobacter load at slaughter under the tested conditions. The logistical constraints inherent in field trials and the natural variability of Campylobacter contamination in naturally infected broiler flocks make it difficult to reproduce experimental results in in situ farm conditions. RCT testing of an intervention strategy in commercial situation is therefore a key step in evaluating pre-harvest interventions against food-borne pathogens.

A bovine botulism outbreak associated with a suspected cross-contamination from a poultry farm.

In October 2014, an outbreak of botulism type D/C occurred on two cattle farms in close proximity. A poultry farm located nearby with no history of botulism had transferred poultry manure to both bovine farms before the beginning of the outbreak. Given this context, epidemiological investigation was conducted to determine if the poultry farm was a reservoir of C. botulinum type D/C and to identify the source of contamination on the cattle farms. Environmental samples were collected at three houses on the poultry farm (boot swabs from the surroundings, swabs from the ventilation system, boot swabs from the poultry litter and darkling beetles samples), and on the two cattle farms (silage samples, boot swabs from the cattle stalls, boot swabs from the cattle pasture and poultry manure samples). These samples were analyzed using real-time PCR after an enrichment step to detect C. botulinum type D/C. On the poultry farm, three boot swabs from the surroundings, two swabs from the ventilation system, one boot swab from the litter and one sample of darkling beetles were detected positive. On one cattle farm, C. botulinum type D/C was identified in a sample of silage made from grass grown on a field on which the poultry manure had previously been stored and in a boot swab from a pasture. On the other cattle farm, C. botulinum type D/C was detected in a sample of poultry manure stored on the cattle farm and in a boot swab from a pasture. This investigation shows that the healthy poultry farm might have been the reservoir of C. botulinum type D/C and that cross-contamination between poultry and cattle likely occurred, resulting in the botulism outbreak on the two cattle farms.

Investigation of a type C/D botulism outbreak in free-range laying hens in France.

In 2014, a botulism outbreak in a flock of laying hens was investigated in France. In the flock of 5020 hens, clinical signs of botulism occurred at 46 weeks of age. A type C/D botulism outbreak was confirmed using the mouse lethality assay for detection of botulinum toxin in serum and a real-time PCR test to detect Clostridium botulinum in intestinal contents. The disease lasted one week with a mortality rate of 2.6% without recurrence. Botulism in laying hens has rarely been reported. Five monthly visits were made to the farm between December 2014 and May 2015 for a longitudinal study of the persistence of C. botulinum in the poultry house after the outbreak, and to assess egg contamination by C. botulinum. Several samples were collected on each visit: in the house (from the ventilation circuit, the egg circuit, water and feed, droppings) and the surrounding area. Thirty clean and 30 dirty eggs were also swabbed at each visit. In addition, 12 dirty and 12 clean eggs were collected to analyse eggshell and egg content. The samples were analysed using real-time PCR to detect type C/D C. botulinum. The bacterium was still detected in the house more than 5 months after the outbreak, mostly on the walls and in the egg circuit. Regarding egg contamination, the bacteria were detected only on the shell but not in the content of the eggs. Control measures should therefore be implemented throughout the egg production period to avoid dissemination of the bacteria, particularly during egg collection.

Influence of enrichment and isolation media on the detection of Campylobacter spp. in naturally contaminated chicken samples.

Investigating Campylobacter epidemiology requires adequate technique and media to ensure optimal culturing and accurate detection and isolation of Campylobacter strains. In the present study, we investigated the performances of three enrichment durations in Bolton broth (0, 24 and 48h) and compared four isolation media (mCCDA, Karmali, Butzler no. 2 and CampyFood agar (CFA)) for the detection of Campylobacter positive samples and the identification of Campylobacter species, from naturally contaminated broiler chicken samples (caeca, neck skin from carcasses, and skin from thighs). We compared our local results to those we obtained with samples from a European survey (caeca and neck skin) and a national survey (neck skin, thigh skin, and breast). Direct plating favored the detection of positive samples highly contaminated by Campylobacter (caeca and neck skin from carcasses) whatever the media. A longer enrichment reduced the rates of Campylobacter recovery except when using Butzler no. 2, more particularly for neck skin which background microflora was less important than in caeca. As a matter of fact, enrichment allowed a higher detection rate of positive samples with low Campylobacter contamination levels (breast, thigh skin), this detection being enhanced when using Butzler no. 2. When comparing the 3 other selective media, CFA was the 2nd most efficient media prior to mCCDA and Karmali. Interestingly, enrichment promoted the growth of Campylobacter coli but this promotion was least with Butzler no. 2 agar. Our study has confirmed the need to adapt the method to the types of samples for improving the detection of Campylobacter and that the method may affect the prevalence of the species.

Efficacy of feed additives against Campylobacter in live broilers during the entire rearing period: Part B.

A total of 636 day-of-hatch Ross 308 broilers chicks were used in 4 independent trials carried out to screen the effect of 12 feed additives on reducing cecal colonization of Campylobacterin broilers. The tested additives were probiotics based on B. subtilis and S. cerevisae, a garlic extract, a blend of herbal substances and essential oils, two different combinations of essential oils and organic acids (OA), two mixtures of flavoring compounds, medium chain fatty acids (MCFA), monoglycerides (MG) of MCFA and MG-MCFA+OA. At 14 days of age, all the birds were orally infected with 0.1 mL of a bacterial suspension of C. jejuni ST-45 diluted at 10(5) cfu/mL in tryptone salt broth. In each trial, there was a positive control group and 2 (Trials 1 and 2) or 4 (Trials 3 and 4) additional treatment groups supplemented with additives, which were added to feed or water only to the finisher (21 to 42 d) diet (Trials 1 and 2) or to the starter (0 to 21 d) and finisher diets (Trials 3 and 4). Feed and water were available ad libitum. On days 35 and 42 of age in Trials 1 and 2, and on days 21, 35 and 42 of age in Trials 3 and 4, 10 (Trials 1 and 2) or 12 birds (Trials 3 and 4) per group were euthanized for cecal sampling. In Trial 1, birds fed with MCFA and MG-MCFA had a significant (P<0.05) reduction in cecal Campylobacter colonization compared to control at 35 d, but only the group treated with MG-MCFA maintained the reduction at 42 d. In Trials 2 to 4, no significant differences (P>0.05) in cecal Campylobacter counts were found between the treated and control animals. In conclusion, although none of the treatments were able to completely prevent the colonization of chickens with C. jejuni, MCFA and MG-MCFAs could reduce the pathogen counts when supplemented from 21 days onwards.

Control strategies against Campylobacter at the poultry production level: biosecurity measures, feed additives and vaccination.

Campylobacteriosis is the most prevalent bacterial foodborne gastroenteritis affecting humans in the European Union, and ranks second in the United States only behind salmonellosis. In Europe, there are about nine million cases of campylobacteriosis every year, making the disease a major public health issue. Human cases are mainly caused by the zoonotic pathogen Campylobacter jejuni. The main source of contamination is handling or consumption of poultry meat. Poultry constitutes the main reservoir of Campylobacter, substantial quantities of which are found in the intestines following rapid, intense colonization. Reducing Campylobacter levels in the poultry chain would decrease the incidence of human campylobacteriosis. As primary production is a crucial step in Campylobacter poultry contamination, controlling the infection at this level could impact the following links along the food chain (slaughter, retail and consumption). This review describes the control strategies implemented during the past few decades in primary poultry production, including the most recent studies. In fact, the implementation of biosecurity and hygiene measures is described, as well as the immune strategy with passive immunization and vaccination trials and the nutritional strategy with the administration of organic and fatty acids, essential oil and plant-derived compound, probiotics, bacteriocins and bacteriophages.

Occurrence of C. botulinum in healthy cattle and their environment following poultry botulism outbreaks in mixed farms.

Ten cattle farms located in an area with a recent history of poultry botulism outbreaks were investigated to evaluate the occurrence of toxigenic C. botulinum in healthy cattle. Environmental samples in the 10 cattle farms and bovine fecal contents in farms with a confirmed environmental contamination were collected. Detection of C. botulinum toxin genes C, D, C/D, D/C and E was performed using real-time PCR. 4.9% (7/143) of the environmental samples collected in the 10 investigated cattle farms were positive for C. botulinum type C/D. Theses samples (boot-swabs in stalls and on pasture and water of a stream) were collected in 3 different farms. One cow dung sample and 3 out of 64 fecal contents samples collected in a single farm were also positive for C. botulinum type C/D. This study demonstrates that cattle are probably indirectly contaminated via poultry botulism in the area and that they can be intermittent carrier of C. botulinum type C/D after poultry botulism outbreaks in mixed farms.

Investigation of Clostridium botulinum in commercial poultry farms in France between 2011 and 2013.

Between 2011 and 2013, 17 poultry botulism outbreaks were investigated in France. All cases were associated with Clostridium botulinum type C-D. Presence of C. botulinum was studied in seven areas: poultry house, changing room, ventilation system, surroundings, animal reservoirs, water, and feed. Swabs, litter, soil, darkling beetles, rodents and wild bird droppings, feed and water samples were collected. The presence of C. botulinum type C-D in the environment of affected flocks was detected in 39.5% of the 185 samples analysed by real-time polymerase chain reaction. C. botulinum type C-D was reported in each area. Four areas were more frequently contaminated, being found positive in more than one-half of farms: darkling beetles (9/11), poultry house (14/17), water (13/16) and surroundings (11/16). After cleaning and disinfection, the ventilation system and/or the soil (in the houses and the surroundings) returned positive results in four out of eight poultry farms. Consequently, darkling beetles, the drinking water, the ventilation system and the soil in the surroundings and the houses were identified as the main critical contaminated areas to consider in poultry farms to prevent recurrence of botulism outbreaks.

Prevalence of and risk factors for Campylobacter colonisation in broiler flocks at the end of the rearing period in France.

1. A study was conducted to estimate the prevalence and quantification by species of Campylobacter infection in broiler flocks at the end of the rearing period and to identify associated risk factors. 2. A questionnaire about the rearing period was completed and caecal samples were collected from 121 broiler flocks in Brittany, France, during 2008. 3. Campylobacter was isolated in 87 out of 121 flocks--a prevalence of 71.9% (95% CI, 63.7-80.1%), including 40.5% of Campylobacter jejuni and 29.8% of Campylobacter coli. 4. The average concentration, in positive flocks, was 7.96 log10 cfu/g and ranged from 3.15 to 10.32 log10 cfu/g. 5. The average concentration by species was: 7.57 log10 cfu/g for C. jejuni and 8.44 log10 cfu/g for C. coli. 6. There was a seasonal effect, with increased risk of Campylobacter colonisation in June, July and August (odds ratio (OR) = 9.59, 95% CI 1.15-79.75). 7. The other factors, associated with lower risk of Campylobacter colonisation, were the acidification of drinking water (OR = 0.33, 95% CI 0.13-0.86), antibiotic treatment at the beginning of the rearing period (OR = 0.20, 95% CI 0.07-0.55) and rodent control around the house (OR = 0.18, 95% CI 0.03-0.95). 8. The results show that hygiene practices and biosecurity measures could lead to a reduction in Campylobacter colonisation.

The Bayesian microbial subtyping attribution model: robustness to prior information and a proposition.

Attributing foodborne illnesses to food sources is essential to conceive, prioritize, and assess the impact of public health policy measures. The Bayesian microbial subtyping attribution model by Hald et al. is one of the most advanced approaches to attribute sporadic cases; it namely allows taking into account the level of exposure to the sources and the differences between bacterial types and between sources. This step forward requires introducing type and source-dependent parameters, and generates overparameterization, which was addressed in Hald's paper by setting some parameters to constant values. We question the impact of the choices made for the parameterization (parameters set and values used) on model robustness and propose an alternative parameterization for the Hald model. We illustrate this analysis with the 2005 French data set of non-typhi Salmonella. Mullner's modified Hald model and a simple deterministic model were used to compare the results and assess the accuracy of the estimates. Setting the parameters for bacterial types specific to a unique source instead of the most frequent one and using data-based values instead of arbitrary values enhanced the convergence and adequacy of the estimates and led to attribution estimates consistent with the other models' results. The type and source parameters estimates were also coherent with Mullner's model estimates. The model appeared to be highly sensitive to parameterization. The proposed solution based on specific types and data-based values improved the robustness of estimates and enabled the use of this highly valuable tool successfully with the French data set.

Prevalence and risk factors for Salmonella spp. contamination in French broiler-chicken flocks at the end of the rearing period.

A nation-wide survey was carried out in 370 randomly chosen French commercial broiler chicken flocks from October 2005 to September 2006 to determine Salmonella spp. prevalence and to identify risk factors for contamination, at the end of the rearing period. The Salmonella status of the flocks was assessed from five faecal samples (litter swabs) analysed by classical bacteriological methods. A flock with at least one contaminated sample was classified as a Salmonella-positive flock. The apparent prevalence of Salmonella was 8.6% (95% CI: 5.7, 11.5%). The most prevalent serovar was S. hadar followed by S. anatum and S. mbandaka. Logistic regression methods were used to analyse the associations between husbandry practices, farm characteristics, general hygiene and the Salmonella status of the sample. The risk for Salmonella contamination of the flock at the end of the rearing period increased when neighbours helped in the placement of day-old chicks. On the contrary, the risk decreased when mobile equipment was dismantled before cleaning and disinfection, when the farm had a specific container for dead-bird disposal and when acetic acid was added to the drinking water.

Endoscopic resection of sporadic duodenal adenomas: an efficient technique with a substantial risk of delayed bleeding.

INTRODUCTION: Data on endoscopic resection of sporadic duodenal adenoma (SDA) are sparse; we present our results concerning safety and efficacy in a retrospective analysis of saline-assisted endoscopic resection of SDA. PATIENTS AND METHODS: The cases of all patients who underwent endoscopic resection for SDA between May 1998 and May 2006 were analyzed. Endoscopic resection was carried out using standard injection and cut methods. In some patients hemoclips and argon plasma coagulation were used, either for prophylaxis or for the treatment of procedure-related bleeding. RESULTS: Thirty-six patients with a total of 37 lesions (mean size 19 mm, range 4 - 50 mm) were analyzed. Lesions larger than 20 mm were more frequently resected piecemeal ( P = 0.022). Intraprocedural bleeding occurred in 14 % of cases, without any significant association with lesion size or the resection technique. One fatal perforation occurred. Macroscopically complete resection was achieved in 97 % of cases, as confirmed by at least one negative control endoscopy with biopsies after a mean follow-up of 15 months. Although clipping and argon plasma coagulation were not applied in any systematic way, it was noticeable that in the 20 cases in which they were used for hemostasis or prevention, no delayed bleeding was observed after the procedures. In contrast, bleeding occurred in 22 % of the 23 procedures performed without these additional techniques. CONCLUSION: Endoscopic resection is an efficient and acceptably safe technique for treating SDA. Further studies need to assess whether systematic bleeding prophylaxis will reduce the incidence of delayed hemorrhage after endoscopic resection.

Miniprobe EUS in the pretherapeutic assessment of early esophageal neoplasia.

BACKGROUND AND STUDY AIMS: The high-frequency endosonography miniprobe has been advocated prior to endoscopic treatment of superficial esophageal neoplasm. We conducted a retrospective study, which summarized our clinical experience with various miniprobe techniques in the assessment of early squamous cell carcinoma (SCC) and superficial adenocarcinoma on Barrett's mucosa (SAB). PATIENTS AND METHODS: A total of 106 superficial and naive esophageal lesions were included in this series (52 SAB, 54 SCC). The results of the miniprobe staging were compared with pathology in order to determine the accuracy of this technique in differentiating mucosal from submucosal infiltration. Secondary aims were a comparison between the frequencies (20- vs. 30-MHz) and coupling methods (water-filled lumen vs. balloon-sheathed catheter) used. RESULTS: Overall accuracy, sensitivity, and specificity to differentiate T1sm from T1 m tumors were 73.5 %, 62 %, and 76.5 %, respectively. Overdiagnosis occurred in 18.6 % of the lesions. No statistically significant difference was found in the accuracy of the miniprobe staging between the two lesion types (SCC and SAB) ( P = 0.65), and results were significantly poorer in the distal part of the esophagus ( P = 0.00039). No significant difference was found between the two frequencies ( P = 0.51). Water-filled lumen proved to be superior to the balloon-sheathed catheter ( P = 0.015). CONCLUSION: High-frequency miniprobe examination has a limited accuracy in the detection of submucosal invasion in early esophageal cancer. Further improvements in acoustic coupling and ultrasound technique are required to improve the miniprobe accuracy before its implementation into clinical routine.