The efficacy of a commercial competitive exclusion product on Campylobacter colonization in broiler chickens in a 5-week pilot-scale study.

The efficacy of the commercial competitive exclusion product Broilact against Campylobacter jejuni was evaluated in broiler chickens in a 5-week pilot-scale study. Newly-hatched broiler chicks were brought from a commercial hatchery. After arrival 50 seeder chicks were challenged orally with approximately 10(3) cfu of C. jejuni, wing marked, and placed back in a delivery box and moved to a separate room. The rest of the chicks (contact chicks) were placed in floor pens, 100 chicks per pen. Birds in two pens were treated orally on the day of hatch with the commercial competitive exclusion (CE) product Broilact, and three pens were left untreated. The following day 10 seeder chicks were introduced into the Broilact treated and untreated control pens. One pen was left both untreated and unchallenged (0-control). Each week the ceca of 10 contact chicks and one seeder chick were examined quantitatively for Campylobacter The treatment prevented or significantly reduced the colonization of the challenge organism in the ceca during the two first weeks; the percentage of colonized birds being 0% after the first week and 30% after the second week in the Broilact treated groups but was 100% in the control groups the entire 5-week rearing period. During the third rearing week the proportion of Campylobacter positive birds started to increase in the treated pens, being 80% after the third week and 95 and 90% after the fourth and fifth rearing weeks, respectively. Similarly the average count of Campylobacter in the cecal contents of the Broilact treated chicks started to increase, the difference between the treated and control chicks being 1.4 logs at the end of the rearing period. Although the protective effect was temporary and occurred only during the first two weeks of the rearing period, the results of this study support the earlier observations that CE flora designed to protect chicks from Salmonella may also reduce Campylobacter colonization of broiler chickens.

Raw grated beetroot linked to several outbreaks of sudden-onset gastrointestinal illness, Finland 2010.

In 2010, 7/44 (16%) reported foodborne outbreaks in Finland were linked with raw beetroot consumption. We reviewed data from the national outbreak registry in order to hypothesize the aetiology of illness and to prevent further outbreaks. In the seven outbreaks, 124 cases among 623 respondents were identified. Consumption of raw beetroot was strongly associated with gastrointestinal illness (relative risk 8∙99, 95% confidence interval 6∙06-13∙35). The illness was characterized by sudden onset of gastrointestinal symptoms; the median incubation time was 40 min and duration of illness 5 h. No common foodborne pathogens or toxins were found in either clinical or beetroot samples, but all tested beetroot samples were of poor quality according to total bacterial counts. Beta-haemolytic Pseudomonas fluorescens was detected in several beetroot samples but its effect on human health is unknown. No outbreaks were reported after the Finnish Food Safety Authority Evira advised against serving raw beetroot in institutional canteens.

Factors related to the prevalence of pathogenic Yersinia enterocolitica on pig farms.

A survey of 788 pigs from 120 farms was conducted to determine the within-farm prevalence of pathogenic Yersinia enterocolitica and a questionnaire of management conditions was mailed to the farms afterwards. A univariate statistical analysis with carriage and shedding as outcomes was conducted with random-effects logistic regression with farm as a clustering factor. Variables with a P value <0·15 were included into the respective multivariate random-effects logistic regression model. The use of municipal water was discovered to be a protective factor against carriage and faecal shedding of the pathogen. Organic production and buying feed from a certain feed manufacturer were also protective against total carriage. Tonsillar carriage, a different feed manufacturer, fasting pigs before transport to the slaughterhouse, higher-level farm health classification, and snout contacts between pigs were risk factors for faecal shedding. We concluded that differences in management can explain different prevalences of Y. enterocolitica between farms.

Humoral immune response to keyhole limpet haemocyanin, the protein carrier in cancer vaccines.

Keyhole limpet haemocyanin (KLH) appears to be a promising protein carrier for tumor antigens in numerous cancer vaccine candidates. The humoral immune response to KLH was characterized at the single-cell level with ELISPOT combined with separations of cell populations according to their expression of homing receptors (HRs). The analysis of HR expressions is expected to reveal the targeting of the immune response in the body. Eight orally primed and four nonprimed volunteers received KLH-vaccine subcutaneously. Circulating KLH-specific plasmablasts were found in all volunteers, 60 KLH-specific plasmablasts/10(6) PBMC in the nonprimed and 136/10(6) in the primed group. The proportion of L-selectin(+) plasmablasts proved high and integrin α(4)ß(7) (+) low. KLH serving as protein carrier in several vaccines, the homing profile of KLH-specific response may be applicable to the cancer antigen parts in the same vaccines. The present data reflect a systemic homing profile, which appears advantageous for the targeting of immune response to cancer vaccines.

Decreasing trend of overlapping multilocus sequence types between human and chicken Campylobacter jejuni isolates over a decade in Finland.

We describe the long-term multilocus sequence typing (MLST) analysis of the population structure and dynamics of 454 Finnish human Campylobacter jejuni isolates, as well as 208 chicken isolates, collected during the mid-1990s to 2007. The sequence type clonal complexes (ST CC) ST-45 CC, ST-21 CC, and ST-677 CC were the most common ones found among all isolates, and they covered 73.9% of all isolates. The ST-283 CC also was found frequently among chicken isolates (8.2%). The predominant STs among all isolates were ST-45, ST-50, and ST-677. ST-137 and ST-230 were common among human isolates, and ST-267 was found more frequently among chicken isolates than human isolates. The ST-45 CC was significantly associated with chicken isolates (P < 0.01), whereas the ST-21 CC was associated with human isolates (P < 0.001). The ST-677 CC was not associated with any host (P = 0.5), and an opposite temporary trend of this complex was seen among chicken and human isolates, with an increase in the former and a decrease in the latter during the study period. Furthermore, the ST-22 and ST-48 CCs were significantly associated with human isolates (P < 0.01), but neither of the CCs was found in chicken isolates. The annual overlap between STs from human and chicken isolates decreased from 76% at the beginning of the study to 58% at the end. Our results suggest that the importance of chicken as a reservoir for strains associated with human infections has declined despite the consumption of domestic chicken meat increasing during the follow-up period by 83%.

Evaluation of isolation methods for pathogenic Yersinia enterocolitica from pig intestinal content.

AIMS: The aim of this study was to evaluate the efficiency of four isolation methods for the detection of pathogenic Yersinia enterocolitica from pig intestinal content. METHODS AND RESULTS: The four methods comprised of 15 isolation steps using selective enrichments (irgasan-ticarcillin-potassium chlorate and modified Rappaport broth) and mildly selective enrichments at 4 or 25 degrees C. Salmonella-Shigella-desoxycholate-calcium chloride agar, cefsulodin-irgasan-novobiocin agar were used as plating media. The most sensitive method detected 78% (53/68) of the positive samples. Individual isolation steps using cold enrichment as the only enrichment or as a pre-enrichment step with further selective enrichment showed the highest sensitivities (55-66%). All isolation methods resulted in high numbers of suspected colonies not confirmed as pathogenic Y. enterocolitica. CONCLUSIONS: Cold enrichment should be used in the detection of pathogenic Y. enterocolitica from pig intestinal contents. In addition, more than one parallel isolation step is needed. SIGNIFICANCE AND IMPACT OF THE STUDY: The study shows that depending on the isolation method used for Y. enterocolitica, the detected prevalence of Y. enterocolitica in pig intestinal contents varies greatly. More selective and sensitive isolation methods need to be developed for pathogenic Y. enterocolitica.

Shedding of Campylobacter spp. in Finnish cattle on dairy farms.

AIMS: The aim of this study was to determine variation of prevalence throughout a year, colonization levels and genotypes of Campylobacter jejuni in Finnish dairy cattle herds. METHODS AND RESULTS: Faecal samples and tank milk samples from three dairy cattle herds were taken five times, and swab samples from drinking troughs once during a 1-year sampling period. The samples were enriched in Bolton broth and subsequently spread on mCCDA. Isolates were then subtyped by pulsed-field gel electrophoresis using SmaI. Campylobacter jejuni was detected in 169 of the 340 faecal samples and in one drinking trough sample. Prevalences between herds and sampling times varied widely. The faecal levels of C. jejuni were mainly low. Between one and four SmaI subtypes were identified from each herd per sampling. Two SmaI subtypes persisted in two of the herds throughout the study. CONCLUSIONS: Dairy cattle can be a long-term reservoir of C. jejuni subtypes similar to clinical isolates. Differences in the colonization potential among C. jejuni strains as well as in the resistance to campylobacter colonization among animals are possible. SIGNIFICANCE AND IMPACT OF THE STUDY: The study provides data on contamination dynamics, colonization levels and the persistence of C. jejuni in dairy cattle.

Severe Outbreak of Sorbitol-Fermenting Escherichia coli O157 via Unpasteurized Milk and Farm Visits, Finland 2012.

Shiga toxin-producing, sorbitol-fermenting Escherichia coli O157 (SF O157) has emerged as a cause of severe human illness. Despite frequent human findings, its transmission routes and reservoirs remain largely unknown. Foodborne transmission and reservoir in cattle have been suspected, but with limited supporting evidence. This study describes the outbreak of SF O157 that occurred in Finland in 2012. The outbreak originated from a recreational farm selling unpasteurized milk, as revealed by epidemiologic and microbiological investigations, and involved six hospitalized children and two asymptomatic adults with culture-confirmed infection. An identical strain of SF O157 was isolated from patients, cattle and the farm environment, and epidemiologic analysis suggested unpasteurized milk as the vehicle of transmission. This study reports the first milkborne outbreak of SF O157, provides supporting evidence of cattle as a reservoir and highlights the health risks related to the consumption of unpasteurized milk.

Prevalence of Pathogenic Yersinia enterocolitica in Finnish Slaughter Pigs.

The prevalence of human pathogenic Yersinia enterocolitica was determined in tonsil and intestinal content samples from 388 healthy fattening pigs at the four biggest Finnish slaughterhouses. These slaughterhouses process 73% of pigs in Finland. Tonsil samples were tested by PCR targeted for yadA, and intestinal samples were cultured. All pathogenic Y. enterocolitica isolates represented bioserotype 4/O:3. The prevalence of Y. enterocolitica in tonsil samples was 60% (95% confidence limit, 55.4 to 65.1%), and its prevalence in intestinal samples was 26% (95% confidence limit, 22.1 to 31.2%). The prevalence of Y. enterocolitica in tonsil and intestinal samples varied between the four slaughterhouses. The tonsil prevalence of Y. enterocolitica was higher in slaughterhouse B, and the prevalence in intestinal content was higher in slaughterhouse C. There were more positive results in both tonsil and intestinal samples in pigs coming from fattening farms than in pigs coming from farrowing-and-fattening farms. A seasonal variation was observed in the prevalence of Y. enterocolitica in intestinal samples, with the highest prevalence during July and August, but no seasonal variation was detected in tonsil samples.

Frictional work in double-sided tablet compression.

The aim of this study was to evaluate the friction during double-sided tablet compression. Dicalcium phosphate dihydrate and lactose were tabletted with a compaction simulator with symmetrical and asymmetrical double-sided sawtooth punch displacement profiles. The estimation of force transmission in a powder column was based on an exponential equation, including the material parameter consisting of both the friction coefficient and Poisson's ratio. This parameter was predetermined from a single-sided compression. A novel equation was derived from a previously presented equation for friction work in single-sided tablet compression. The basic assumption was drawn from the linearly decreasing movement of infinitely thin particle layers, which are produced as the compressing punch surface approaches the other punch. This calculation was also based on the assumption that the equilibrium point, where the particles do not move, is halfway between the punches in the symmetrical profile and at a distance proportional to the amplitudes of the asymmetrical upper and lower sawtooth profiles. The tensile strength of tablets compressed with single-double-sided profiles was identical, and thus the behavior of the materials studied under compression was independent of the compression profiles. The friction work values that were calculated with the proposed expression for double-sided profiles were close to the theoretical values, as estimated by calculations based on compressions with single-sided profiles. In conclusion, the novel mathematical expression opens new possibilities for the evaluation of friction in double-sided compression; for example, in rotary press tabletting.

Epidemiological evaluation of the salmonella-controlling effect of a nationwide use of a competitive exclusion culture in poultry.

The purpose of the study was to evaluate the salmonella contamination-controlling effect of a nationwide use of a competitive exclusion culture in poultry. During a 5-yr period, a bacterial flora from caecum of adult birds was given to broiler chickens in order to control salmonella infection. The culture was given on 144 occasions to a total of 2.86 million chickens. The base for the evaluation was the compulsory and regular control for salmonella contamination conducted according to the salmonella control program applied to Sweden. Epidemiological evaluation, especially in relation to the use of the culture during a period when salmonella was spread from a feed factory, strongly supports the conclusion that the culture has a salmonella contamination-preventing effect under field conditions, as earlier had been found in laboratory studies. Questionnaires answered by producers did not indicate significant adverse effects of the treatment.

Efficacy of a commercial competitive exclusion product against a chicken pathogenic Escherichia coli and E coli O157:H7.

The prevention of caecal colonisation of chicks by a poultry pathogenic Escherichia coli O20:K-:H8 and the human pathogenic E coli O157:H7 was studied in vivo in four one-week laboratory trials. Chicks were treated with a competitive exclusion product on the day of hatch and challenged one day later. The poultry pathogenic serotype showed higher levels of caecal colonisation than the human pathogenic serotype, but the protection against both pathogens was highly significant.

Trends in Campylobacter incidence in broilers and humans in six European countries, 1997-2007.

The objective of this study was to examine incidences of Campylobacter in broilers and humans, and to describe seasonal variation and long-term trends by comparing longitudinal surveillance data in six Northern European countries (Denmark, Finland, Iceland, Norway, Sweden and the Netherlands). Due to high degree of seasonality and autocorrelation, seasonally adjusted (de-seasonalized) and trend adjusted data (de-trended) were used for comparing incidences within and between the six countries. De-seasonalized time series were obtained by fitting the incidence time series to mean monthly temperature and then removing this effect from the data. Long-term trends were fitted to the de-seasonalized time series. The incidence of Campylobacter colonization in broiler flocks and incidence of campylobacteriosis in humans showed a concordant seasonality for all the countries. There was a strong association between the incidence in both broilers and humans in a given month and the mean temperature of the northern hemisphere in the same month, as well as the preceding month, as shown by the cross-correlations and the chosen Generalized Additive Model. Denmark and Sweden showed a steadily decreasing trend for Campylobacter in broilers and human campylobacteriosis in the period 2001-2007. In Iceland, there was a decreasing trend for campylobacteriosis in humans from 1999 to 2007, whilst the broiler trend for Campylobacter was stable from 2001 to 2004, then falling thereafter. In Norway, the human campylobacteriosis trend showed a steady increase throughout the period. On the other hand, the Norwegian broiler trend for Campylobacter showed a decrease from 2001 until 2004, but was thereafter stable. There was no significant decrease or increase in incidence for human campylobacteriosis in the Netherlands, and the trend for Campylobacter in broilers was close to stable. The seasonality seen in broiler and human closely follows the temperature, and was probably caused, at least partly, by temperature related factors.

Comparison of two different types of competitive exclusion products.

The efficacy of two different types of commercial competitive exclusion (CE) products against Salmonella was studied in three chicken assay trials. Chicks were treated on the day of hatch and challenged one day later either with Salm. infantis (Trials 1 and 2) or a combination of Salm. infantis and Salm. enteritidis (Trial 3). The caeca of the birds were examined for Salmonella five days after challenge. The mean logarithmic counts of Salmonella were from 3.4 to 5.7 in the groups treated with product. A derived from the whole caecal contents of an adult bird, from 0.0 to 1.2 in the groups treated with product B, a highly selected product that does not contain any clostridia, and from 6.3 to 7.6 in the control groups. None of the challenge organisms superseded the other in Trial 3, and neither did the double challenge affect the protective capacity of the treatment materials.

Efficacy of a commercial competitive exclusion product against Campylobacter jejuni.

1. Newly-hatched broiler chicks were treated orally with a commercial competitive exclusion product (Broilact) in 3 replicate trials 2. After 24 h the treated chicks and untreated control chicks were challenged orally with approximately 10(4) cfu of Campylobacter jejuni. 3. The caeca of the birds were examined quantitatively for campylobacter 12 d after the challenge. 4. In 3 separate trials, the treatment prevented or reduced colonisation of the challenge organisms in the caeca. The percentage of colonised birds varied from 0% to 62% in the treated groups and was 100% in the control groups. The average number of campylobacter was considerably lower in the treated groups than in the control groups.

Temporal and geographical distribution and overlap of Penner heat-stable serotypes and pulsed-field gel electrophoresis genotypes of Campylobacter jejuni isolates collected from humans and chickens in Finland during a seasonal peak.

The association of Penner heat-stable serotypes and pulsed-field gel electrophoresis genotypes of 208 human and 30 chicken Campylobacter jejuni isolates was studied. Overall, 46% of the human strains had overlapping sero- and genotype combinations with chicken strains. The percentage was reduced to 31% for strains that were considered temporally related. This suggests common environmental sources.

Stability of related human and chicken Campylobacter jejuni genotypes after passage through chick intestine studied by pulsed-field gel electrophoresis.

The genomic stability of 12 Campylobacter jejuni strains consisting of two groups of human and chicken isolates was studied by analysis of their PFGE (pulsed-field gel electrophoresis) patterns after passage through newly hatched chicks' intestines. The patterns of SmaI, SalI, and SacII digests remained stable after intestinal passage, except for those of two strains. One originally human strain, FB 6371, changed its genotype from II/A (SmaI/SacII) to I/B. Another strain, BTI, originally isolated from a chicken, changed its genotype from I/B to a new genotype. The genomic instability of the strains was further confirmed by SalI digestion and ribotyping of the HaeIII digests. In addition, heat-stable serotype 57 of strain FB 6371 changed to serotype 27 in all isolates with new genotypes but remained unchanged in an isolate with the original genotype. Serotype 27 of strain BTI remained stable. Our study suggests that during intestinal colonization, genomic rearrangement, as demonstrated by changed PFGE and ribopatterns, may occur.

Note: inhibition of the growth of Yersinia enterocolitica O:3 by the microflora of porcine caecum and ileum in an in vitro model.

The growth of Yersinia enterocolitica O:3 was tested in an in vitro model of the porcine intestine at the physiological temperature of 39 degrees C of growing pigs. The model supported a stable population of Y. enterocolitica at a level 10(8)-10(9) cells ml-1. Plasmid profile analysis and the Ca(2+)-dependent proportion of the population suggested that the great majority of the Y. enterocolitica population retained the 70 kb virulence plasmid, pYV, throughout the experimental period of 5 d. The growth of Y. enterocolitica was substantially inhibited by the ileal and the caecal flora compared to the growth of the bacterium alone. Yersinia enterocolitica was not isolated after 3 d of cultivation.

Differential homing commitments of antigen-specific T cells after oral or parenteral immunization in humans.

Animal experiments show that lymphocytes activated in the intestine circulate through mesenteric lymph nodes, lymphatics, and blood, returning to the gut. Homing into intestinal lamina propria is mediated by lymphocyte surface homing receptors, mainly the alpha4beta7-integrin. We studied in humans whether intestinal T cells entering the blood upon antigenic activation would exhibit homing commitments to the gut. Volunteers were immunized with keyhole limpet hemocyanin (KLH) first orally and then parenterally or only parenterally, and the expression of alpha4beta7 on T cells specific for KLH or tetanus toxoid was studied. Circulating T cells were depleted of alpha4beta7+ cells by immunomagnetic selection. This depletion removed a significant proportion of the KLH-specific cells (mean decrease in proliferative response of 71%) in the orally immunized volunteers. No difference in the KLH-induced proliferation was found between the total and the alpha4beta7-depleted populations in volunteers parenterally immunized with KLH, regardless of whether a preceding mucosal priming had taken place or not. In both immunization groups, the depletion of alpha4beta7+ cells had no influence on the proliferative response to tetanus toxoid. We conclude that, in contrast to T cells activated by parenteral immunization, gut-derived T cells have preferential homing commitments to the gut. This commitment was no longer observed after a subsequent parenteral Ag administration. Besides showing that the site of Ag encounter determines the expression of homing receptors, the present study is the first to provide evidence for a circulation of newly activated Ag-specific intestinal T cells back to the gut in humans.