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Maintaining a healthy gut environment is a prerequisite for sustainable animal production. The gut plays a key role in the digestion and absorption of nutrients and constitutes an initial organ exposed to external factors influencing bird’s health. The intestinal epithelial barrier serves as the first line of defense between the host and the luminal environment. It consists of a continuous monolayer of intestinal epithelial cells connected by intercellular junctional complexes which shrink the space between adjacent cells. Consequently, free passing of solutes and water via the paracellular pathway is prevented. Tight junctions (TJs) are multi-protein complexes which are crucial for the integrity and function of the epithelial barrier as they not only link cells but also form channels allowing permeation between cells, resulting in epithelial surfaces of different tightness. Tight junction’s molecular composition, ultrastructure, and function are regulated differently with regard to physiological and pathological stimuli. Both in vivo and in vitro studies suggest that reduced tight junction integrity greatly results in a condition commonly known as “leaky gut”. A loss of barrier integrity allows the translocation of luminal antigens (microbes, toxins) via the mucosa to access the whole body which are normally excluded and subsequently destroys the gut mucosal homeostasis, coinciding with an increased susceptibility to systemic infection, chronic inflammation and malabsorption. There is considerable evidence that the intestinal barrier dysfunction is an important factor contributing to the pathogenicity of some enteric bacteria. It has been shown that some enteric pathogens can induce permeability defects in gut epithelia by altering tight junction proteins, mediated by their toxins. Resolving the strategies that microorganisms use to hijack the functions of tight junctions is important for our understanding of microbial pathogenesis, because some pathogens can utilize tight junction proteins as receptors for attachment and subsequent internalization, while others modify or destroy the tight junction proteins by different pathways and thereby provide a gateway to the underlying tissue. This review aims to deliver an overview of the tight junction structures and function, and its role in enteric bacterial pathogenesis with a special focus on chickens. A main conclusion will be that the molecular mechanisms used by enteric pathogens to disrupt epithelial barrier function in chickens needs a much better understanding, explicitly highlighted for Campylobacter jejuni, Salmonella enterica and Clostridium perfringens. This is a requirement in order to assist in discovering new strategies to avoid damages of the intestinal barrier or to minimize consequences from infections.

Background M. morganii is a bacterium frequently associated with urinary infections in humans. While many human strains are sequenced, only the genomes of few poultry strains are available. Here, we performed a detailed characterization of five highly resistant Morganella morganii strains isolated in association with Escherichia coli from diseased domestic Austrian poultry flocks, namely geese, turkeys and chicken layers. Additionally, we sequenced the genomes of these strains by NGS and analyzed phylogenetic clustering, resistance and virulence genes in the context of host-specificity. Results Two strains were identified to be Extended Spectrum Beta Lactamase (ESBL) and one as AmpC beta-lactamases (AMP-C) phenotype, while two were ESBL negative. By integrating the genome sequences of these five poultry strains with all the available M. morganii genomes, we constructed a phylogenetic tree that clearly separates the Morganella genus into two clusters (M1 and M2), which approximately reflect the proposed subspecies classification ( morganii and sibonii ). Additionally, we found no association between phylogenetic structure and host, suggesting interspecies transmission. All five poultry strains contained genes for resistance to aminocoumarins, beta-lactams, colistin, elfamycins, fluoroquinolones, phenicol, rifampin and tetracycline. A comparative genomics analysis of virulence genes showed acquisition of novel virulence genes involved in secretion system and adherence in cluster M2. We showed that some of these genes were acquired by horizontal gene transfer from closely related Morganellaceae species and propose that novel virulence genes could be responsible for expansion of tissue tropism in M. morganii . Finally, we detected variability in copy number and high sequence divergence in toxin genes and provided evidence for positive selection in insecticidal toxins genes, likely reflecting host-related adaptations. Conclusions In summary, this study describes i) the first isolation and characterization of M. morganii from goose and turkey, ii) a large-scale genetic analysis of M. morganii and an attempt to generate a global picture of the M. morganii intraspecific phylogenetic structure.

Colisepticaemia caused by avian pathogenic Escherichia coli (APEC) is a challenging disease due to its high economic importance in poultry, dubious pathogenesis and potential link with zoonosis and food safety. The existing in vitro studies can’t define hallmark traits of APEC isolates, suggesting a paradigm shift towards host response to understand pathogenesis. This study investigated the comprehensive pathological and microbial progression of colisepticaemia, and transmission of E. coli into eggs using novel tools. In total 48 hens were allocated into three groups and were inoculated intratracheally with ilux2 - E. coli PA14/17480/5­/ovary (bioluminescent strain), E. coli PA14/17480/5­/ovary or phosphate buffered saline. Infection with both strains led to typical clinical signs and lesions of colibacillosis as in field outbreaks. Based on lung histopathology, colisepticaemia progression was divided into four disease stages as: stage I (1–3 days post infection (dpi)), stage II (6 dpi), stage III (9 dpi) and stage IV (16 dpi) that were histologically characterized by predominance of heterophils, mixed cells, pyogranuloma, and convalescence, respectively. As disease progressed, bacterial colonization in host organs also decreased, revealed by the quantification of bacterial bioluminescence, bacteriology, and quantitative immunohistochemistry. Furthermore, immunofluorescence, immunohistochemistry, and bacteria re-isolation showed that E. coli colonized the reproductive tract of infected hens and reached to egg yolk and albumen. In conclusion, the study provides novel insights into the pathogenesis of colisepticemia by characterizing microbial and pathological changes at different disease stages, and of the bacteria transmission to table eggs, which have serious consequences on poultry health and food safety.

The mycotoxin deoxynivalenol (DON) is of high importance among feed contaminants because of its frequent occurrence in toxicologically relevant concentrations worldwide. Cereal crops, the main component of chicken diet, are commonly contaminated with DON, resulting in frequent exposure of chickens to DON. Likewise, Campylobacter ( C. ), a pathogen of major public and animal health concern, is frequently found in chicken flocks and poses a threat to the One Health approach. Campylobacter colonizes the gastrointestinal (GI) tract of poultry with a high bacterial load in the caeca. However, the mechanism of C. jejuni colonization in chickens is still not understood albeit it is well known that C. jejuni resides primarily in the mucosal layer of the chicken intestine. Therefore, in the actual study we focused on the effect of exposure to DON and/or C. jejuni on expression profiles of intestinal mucins (MUC1, MUC2), β-defensins (Gallinacin (GAL) 10, 12), cytokines (Toll-like receptor 2 (TLR2), Interleukin (IL) 6, 8, Interferon-γ (IFN)-γ), inducible nitric oxide synthase 2 (iNOS2), as well as selected tight junction proteins (Claudin 5 (CLDN5), Occludin (OCLN), and zonula occludens-1 (ZO1) via RT-qPCR. For this, a total of 150 one-day-old Ross 308 broiler chickens were randomly allocated to six different groups (n = 25 with 5 replicates/group) and were fed for 5 weeks with either contaminated diets (5 or 10 mg DON/kg feed) or basal diets (control). Following oral infection of birds with C. jejuni NCTC 12744 at 14 days of age, several changes in gene expression patterns were demonstrated. A significant ( P  ≤ 0.05) downregulation of MUC2 mRNA expression was observed in birds fed DON5 and DON10 diet, as well as in birds co-exposed to DON5 and C. jejuni at 7 dpi. Furthermore, at 14 dpi, MUC2 mRNA expression was significantly ( P  ≤ 0.05) downregulated in birds fed DON (5 mg and 10 mg/kg diet) with and without C. jejuni and in birds infected solely with C. jejuni . The actual study also demonstrated that co-exposure of broiler chickens to DON and C. jejuni resulted in a decreased barrier function via downregulation of OCLD mRNA expression. In addition, Campylobacter infection induced an increased expression of the antimicrobial peptide GAL12 and the IL8 gene, indicating that C. jejuni can initiate an immune response in the chicken gut in a proinflammatory manner. Similarly, DON with and without C. jejuni induced upregulation of GAL10 and GAL12 mRNA expression at 7 dpi. Moreover, no change in iNOS2 mRNA expression was observed in both the jejunum and the cecum at either 7 dpi or 14 dpi, suggesting unchanged NO production during exposure/infection. In conclusion, we confirmed that DON contamination corresponding to the currently applicable EU guidance value of 5 mg DON/kg feed affects the intestinal gene expression profiles of broilers, mainly in a dose-independent manner . Furthermore, DON exposure interacted synergistically with C. jejuni challenge regarding mucins, innate immunity gene expression in either the jejunum or the cecum, suggesting immunomodulatory activity of both foodborne agents (DON and C. jejuni ).

Purpose The purpose of this study is to investigate whether, how and why gender cues influence brand perception and subsequent purchasing behaviour. Design/methodology/approach Across four experimental studies conducted online with either a convenience sample (Studies 1a and 1b) or a representative sample of consumers (Studies 2 and 3), the authors empirically investigate whether gender cues impact brand perception along dimensions of warmth and competence and how other warmth and competence cues in a consumer environment moderate the effect of gender cues on consumer brand perceptions. Findings Gender cues (e.g. gender-typed colours and shapes) activate gender-stereotypical knowledge of warmth and competence, which spills over to the brand. This effect depends on the presence of other competence cues in a consumer’s environment. In contrast to conventional practice, in the presence of a high competence cue (e.g. reputable brands), feminine gender cues enhance purchase likelihood (via activation of warmth perceptions), whereas masculine cues actually decrease purchase likelihood. In contrast, in the presence of a low competence cue (e.g. new companies), masculine gender cues enhance purchase likelihood (via activation of competence perceptions), whereas feminine cues lower purchase likelihood. Research limitations/implications The authors used an experimental approach to explicitly test for causality and isolate the effect of gender cues in a controlled setting. Future research should further address the implication of gender cues using actual sales data. Practical implications Reputable companies often explicitly use cues to highlight their competence. The results of this research suggest that managers may want to reconsider this approach. That is, marketers of brands with established high competence should consider integrating more feminine cues to highlight their warmth, such as feminine shapes (e.g. circles and ovals) or feminine colours (e.g. a shade of pink) in their packaging and marketing communication. In contrast, companies that have not established their competence or not-for-profit organisations would be better off integrating masculine cues. Originality/value This is the first research to empirically investigate the effect of gender cues on brand perception and subsequent purchase behaviour. Not only does this research show that gender cues can alter brand perception along the warmth and competence perception but also the authors address the call to identify conditions under which warmth versus competence cues enhance brand perception and purchase likelihood (Aaker et al., 2010). In particular, this research demonstrates how multiple warmth and competence cues interact with each other.

Background Deoxynivalenol (DON) is a type B trichothecene mycotoxin that is commonly found in cereals and grains worldwide. The presence of this fungal secondary-metabolite raises public-health concerns at both the agriculture and food industry level. Recently, we have shown that DON has a negative impact on gut integrity, a feature also noticed for Campylobacter (C.) jejuni . We further demonstrated that DON increased the load of C. jejuni in the gut and inner organs. In contrast, feeding the less toxic DON metabolite deepoxy-deoxynivalenol (DOM-1) to broilers reduced the Campylobacter load in vivo. Consequently, it can be hypothesized that DON and DOM-1 have a direct effect on the growth profile of C. jejuni . The aim of the present study was to further resolve the nature of this interaction in vitro by co-incubation and RNA-sequencing. Results The co-incubation of C. jejuni with DON resulted in significantly higher bacterial growth rates from 30 h of incubation onwards. On the contrary, the co-incubation of C. jejuni with DOM-1 reduced the CFU counts, indicating that this DON metabolite might contribute to reduce the burden of C. jejuni in birds, altogether confirming in vivo data. Furthermore, the transcriptomic profile of C. jejuni following incubation with either DON or DOM-1 differed. Co-incubation of C. jejuni with DON significantly increased the expression of multiple genes which are critical for Campylobacter growth, particularly members of the Flagella gene family, frr (ribosome-recycling factor), PBP2 futA-like (Fe 3+ periplasmic binding family) and PotA (ATP-binding subunit). Flagella are responsible for motility, biofilm formation and host colonization, which may explain the high Campylobacter load in the gut of DON-fed broiler chickens. On the contrary, DOM-1 downregulated the Flagella gene family and upregulated ribosomal proteins. Conclusion The results highlight the adaptive mechanisms involved in the transcriptional response of C. jejuni to DON and its metabolite DOM-1, based on the following effects: (a) ribosomal proteins; (b) flagellar proteins; (c) engagement of different metabolic pathways. The results provide insight into the response of an important intestinal microbial pathogen against DON and lead to a better understanding of the luminal or environmental acclimation mechanisms in chickens.

Food-borne infections with Salmonella are among the most common causes of human diseases worldwide, and infections with the serovar Infantis are becoming increasingly important. So far, diverse phenotypes and genotypes of S. Infantis have been reported. Therefore, the present study aimed to investigate the infection dynamics of two different S. Infantis strains in broilers. For this purpose, 15 birds were infected on day 2 of life with 10 8  CFU/ml of a pESI+ or a pESI- S. Infantis strain, respectively. Ten uninfected birds served as in-contact birds to monitor transmission. In both groups, an increase of infection was observed from 7 days of age onwards, reaching its peak at 28 days. However, the pESI+ strain proved significantly more virulent being re-isolated from most cloacal swabs and organs by direct plating. In contrast, the pESI- strain could be re-isolated from cloacal swabs and caeca only when enrichment was applied. Although the excretion of this strain was limited, the transmission level to in-contact birds was similar to the pESI+ strain. Differences in infection dynamics were also reflected in the antibody response: whereas the pESI+ strain provoked a significant increase in antibodies, antibody levels following infection with the pESI- strain remained in the range of negative control birds. The actual findings provide for the first time evidence of S. Infantis strain-specific infectivity in broilers and confirm previous observations in the field regarding differences in persistence on farms and resistance against disinfectants.

Staphylococcus species are widespread in poultry environments and can cause various infections, often when the host’s defences are compromised. This manuscript reports on a co-infection of chickens with Staphylococcus lentus and Staphylococcus aureus associated with an outbreak of arthritis, synovitis, and osteomyelitis in an organic broiler breeder flock in Austria. Clinically, the affected flock showed weakness, lethargy, lameness, and increased mortality. Post-mortem examinations identified purulent arthritis and femoral head necrosis. Bacteriological analysis using MALDI-TOF MS identified both S. aureus and S. lentus in the affected joints. Antibiotic resistance testing revealed significant resistance, particularly in S. lentus. Histological analysis showed severe inflammation and bacterial colonies in the joints. While S. aureus is a common pathogen in poultry, S. lentus is less frequently reported. This study emphasises the need for detailed bacterial characterisation in outbreaks to better understand the role of less common pathogens like S. lentus. Further research is necessary to elucidate the impact of S. lentus on poultry health and its role in causing arthritis and synovitis, highlighting the importance of comprehensive investigation in such outbreaks.

Staphylococcus (S.) aureus is considered as a major mastitis pathogen, with considerable epidemiological information on such infections while the epidemiology of coagulase-negative staphylococci (CNS) is more controversial. The aim of this study was to use matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) technology for identification of staphylococci isolated from bovine milk at species level and to characterize them in reference to presentation, somatic cell count (SCC), bacterial shedding (cfu) and antimicrobial resistance patterns. A total of 200 staphylococcal isolates (S. aureus n = 100; CNS n = 100) originating from aseptically collected quarter milk samples from different quarters of dairy cows were included in the study. They originated from cases of clinical (CM) and subclinical mastitis (SCM) or were isolated from milk with SCC ≤ 100,000 cells/mL in pure culture. We found staphylococci predominantly in cases of SCM (n = 120). In low-SCC cows, 12 S. aureus and 32 CNS isolates were detected. Eighteen percent of each were associated with CM. Eleven CNS species were identified, S. chromogenes (n = 26) and S. xylosus (n = 40) predominated. CNS, particularly those in low-SCC cows, showed higher MIC90 (minimal inhibitory concentration) values for penicillin, ampicillin, cefoperazone, pirlimycin and marbofloxacin. Based on the present results, a careful interpretation of laboratory results is recommended to avoid antimicrobial therapy of staphylococci without clinical relevance and to ensure prudent use of antimicrobials.

Ornithobacterium rhinotracheale is one of the most important bacterial agents of respiratory diseases in poultry. For correct identification and characterization of this fastidious bacterium, reliable diagnostic tools are essential. Still, phenotypic tests are used to identify O. rhinotracheale and serotyping is the most common method for characterization, despite known drawbacks and disadvantages such as divergent results, cross-reactivity between strains, or the non-typeability of strains. The intention of the present study was to evaluate MALDI-TOF MS and whole genome sequencing for the identification and characterization of O. rhinotracheale. For this purpose, a selection of 59 well-defined reference strains and 47 field strains derived from outbreaks on Austrian turkey farms were investigated by MALDI-TOF MS. The field strains originated from different geographical areas in Austria with some of the isolates derived from multiple outbreaks on farms within a year, or recurrent outbreaks over several years. MALDI-TOF MS proved a suitable method for identification of O. rhinotracheale to genus or species level except for 3 strains representing serotypes M, K and F. Phylogenetic analysis showed that most strains grouped within one cluster even though they were comprised of different serotypes, while serotypes F, K, and M clearly formed a different cluster. All field isolates from turkey farms clustered together, independent of the origin of the isolates, e.g., geographical area, multiple outbreaks within a year or recurrent outbreaks over several years. Whole genome sequencing of serotype M, K and F strains confirmed the extraordinary status and deviation from known fully-sequenced strains due to a lack of sequence similarity. This was further confirmed by alignments of single genes (16S-RNA and rpoB) and multilocus sequence typing although the demarcation was less obvious. Altogether, the results indicate that these three serotypes belong to a different species than O. rhinotracheale, and might even be members of multiple new species.