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This prospective, descriptive study assessed caring practices of pet cat and dog owners in Northern Ireland with a survey questionnaire, focusing primarily on feeding and exercising. Owners of both pet cats and dogs reported that they fed their pets either twice or three times a day (53 and 78% for cats and dogs, respectively). However, in the case of nearly 40% of pet cat owners it transpired that they fed their cat(s) as often as the animal demanded (18.8%) or that food was available all the time (20.7%). Cat owners reported that their pets had access to outdoors (84%) and were not provided with any play time (53.1%). The highest number of pet dogs (46.1%) were reported as being walked daily for less than 1 h, and the highest number (38.4%) were played with daily for less than 1 h. Both cat (60%) and dog (61%) owners perceived the body condition of their animal ‘as it should be’ for animal age/sex. However, both pet cat (76%) and dog (63%) owners reported that neither their pet's bodyweight nor its body condition was monitored. Our results suggest not all pet cat and dog owners are aware of the fundamentals of their animals’ feeding requirements as regards health and many provide insufficient physical activity required for their animals’ health and welfare. The major strengths of the study include the size and geographic distribution of the study population allowing the generalisation of the results to other locations.

Filter feeding is a biotic process that brings waterborne bacteria in close contact with each other and may thus support the horizontal transfer of their antimicrobial resistance genes. This laboratory study investigated whether the freshwater sponge Ephydatia fluviatilis supported the transfer of vancomycin resistance between two Enterococcus faecalis strains that we previously demonstrated to exhibit pheromone responsive plasmid conjugation. Microcosm experiments exposed live and dead colonies of laboratory-grown sponges to a vancomycin-resistant donor strain and a rifampicin-resistant recipient strain of Ent. faecalis. Enterococci with both resistance phenotypes were detected on double selection plates. In comparison to controls, abundance of these presumed transconjugants increased significantly in water from sponge microcosms. Homogenized suspensions of sponge cells also yielded presumed transconjugants; however, there was no significant difference between samples from live or dead sponges. Fluorescent in situ hybridization analysis of the sponge cell matrix using species-specific probes revealed the presence of enterococci clusters with cells adjacent to each other. The results demonstrated that sponge colonies can support the horizontal transfer of antimicrobial resistance although the mechanism underlying this process, such as binding of the bacteria to the sponge collagen matrix, has yet to be fully elucidated.

Clostridiodes difficile (C. difficile) was ranked an “urgent threat” by the Centers for Disease Control and Prevention (CDC) in 2019. C. difficile infection (CDI) is the most common healthcare-associated infection (HAI) in the United States of America as well as the leading cause of antibiotic-associated gastrointestinal disease. C. difficile is a gram-positive, rod-shaped, spore-forming, anaerobic bacterium that causes infection of the epithelial lining of the gut. CDI occurs most commonly after disruption of the human gut microflora following the prolonged use of broad-spectrum antibiotics. However, the recurrent nature of this disease has led to the hypothesis that biofilm formation may play a role in its pathogenesis. Biofilms are sessile communities of bacteria protected from extracellular stresses by a matrix of self-produced proteins, polysaccharides, and extracellular DNA. Biofilm regulation in C. difficile is still incompletely understood, and its role in disease recurrence has yet to be fully elucidated. However, many factors have been found to influence biofilm formation in C. difficile, including motility, adhesion, and hydrophobicity of the bacterial cells. Small changes in one of these systems can greatly influence biofilm formation. Therefore, the biofilm regulatory system would need to coordinate all these systems to create optimal biofilm-forming physiology under appropriate environmental conditions. The coordination of these systems is complex and multifactorial, and any analysis must take into consideration the influences of the stress response, quorum sensing (QS), and gene regulation by second messenger molecule cyclic diguanosine monophosphate (c-di-GMP). However, the differences in biofilm-forming ability between C. difficile strains such as 630 and the “hypervirulent” strain, R20291, make it difficult to assign a “one size fits all” mechanism to biofilm regulation in C. difficile. This review seeks to consolidate published data regarding the regulation of C. difficile biofilms in order to identify gaps in knowledge and propose directions for future study., Author summary Clostridioides difficile (C. difficile) is an opportunistic bacterial pathogen that causes infection of the human gut epithelium following disruption of the normal gut microflora, usually by broad-spectrum antibiotics. C. difficile infection (CDI) is recurrent in 20% to 30% of cases and can lead to significant health-related complications such as pseudomembranous colitis and, in severe cases, death. The impact and cost of this pathogen on healthcare systems are significant, and some aspects of the pathogen’s lifestyle in the host are, as yet, unknown. It is hypothesised that C. difficile exists in the gut as a biofilm due to the infection’s severity and recurrent nature. The biofilm mode of bacterial growth can protect the cells from external factors such as antibiotic treatment, physiological processes, and the immune system. However, biofilm regulation in C. difficile is not yet fully characterised, and in this review, we consolidate published primary research on C. difficile biofilm regulation to gain a comprehensive overview of the factors involved and how they may interact to enable biofilm development within a host.

Enterococci, which are on the WHO list of priority pathogens, are commonly encountered in hospital acquired infection and are becoming increasing significant due to the development of strains resistant to multiple antibiotics. Enterococci are also important microorganisms in the environment, and their presence is frequently used as an indicator of faecal pollution. Their success is related to their ability to survive within a broad range of habitats and the ease by which they acquire mobile genetic elements, including plasmids, from other bacteria. The enterococci are frequently present within a bacterial biofilm, which provides stability and protection to the bacterial population along with an opportunity for a variety of bacterial interactions. Enterococci can accept extrachromosomal DNA both from within its own species and from other bacterial species, and this is enhanced by the proximity of the donor and recipient strains. It is this exchange of genetic material that makes the role of biofilms such an important aspect of the success of enterococci. There remain many questions regarding the most suitable model systems to study enterococci in biofilms and regarding the transfer of genetic material including antibiotic resistance in these biofilms. This review focuses on some important aspects of biofilm in the context of horizontal gene transfer (HGT) in enterococci.

Enterococci and biofilm-associated infections are a growing problem worldwide, given the rise in antibiotic resistance in environmental and clinical settings. The increasing incidence of antibiotic resistance and its propagation potential within enterococcal biofilm is a concern. This requires a deeper understanding of how enterococcal biofilm develops, and how antibiotic resistance transfer takes place in these biofilms. Enterococcal biofilm assays, incorporating the study of antibiotic resistance transfer, require a system which can accommodate non-destructive, real-time experimentation. We adapted a Gene Frame® combined with fluorescence microscopy as a novel non-destructive platform to study the conjugal transfer of vancomycin resistance in an established enterococcal biofilm.A multi-purpose fluorescent in situ hybridisation (FISH) probe, in a novel application, allowed the identification of low copy number mobile elements in the biofilm. Furthermore, a Hoechst stain and ENU 1470 FISH probe identified Enterococcus faecium transconjugants by excluding Enterococcus faecalis MF06036 donors. Biofilm created with a rifampicin resistant E. faecalis (MW01105Rif) recipient had a transfer efficiency of 2.01 × 10−3, double that of the biofilm primarily created by the donor (E. faecalis MF06036). Conjugation in the mixed enterococcal biofilm was triple the efficiency of donor biofilm. Double antibiotic treatment plus lysozyme combined with live/dead imaging provided fluorescent micrographs identifying de novo enterococcal vancomycin resistant transconjugants inside the biofilm. This is a model system for the further study of antibiotic resistance transfer events in enterococci. Biofilms promote the survival of enterococci and reduce the effectiveness of drug treatment in clinical settings, hence giving enterococci an advantage. Enterococci growing in biofilms exchange traits by means of horizontal gene transfer, but currently available models make study difficult. This work goes some way to providing a non-destructive, molecular imaging-based model system for the detection of antibiotic resistance gene transfer in enterococci.

A combined reinforcement and drainage geogrid (herein referred to as “draining geogrid”) has been developed for the construction of steep slopes and walls with low-permeability fills. With such fills, drainage is necessary to minimize pore pressure buildup. The draining geogrids provide drainage thanks to channels in their longitudinal strips. The design of reinforced slopes and walls using draining geogrids consists in: estimating the magnitude and time for dissipation of excess pore water pressures in the reinforced fill; evaluating the stability of the reinforced fill; and evaluating if the hydraulic transmissivity of the draining geogrids is sufficient to ensure rapid dissipation of excess pore water pressure. Draining geogrids have been used for a decade. However, a significant increase in the number of applications of draining geogrids is expected due to the increased scarcity of draining fill and the increasing use of local soils for environmental purposes. An updated design method for reinforced slopes and walls using draining geogrids is presented.

Geosynthetic reinforced piled embankments (GRPE’s) have become an increasingly popular means of constructing on unsuitable foundation soils. However the design of GRPE’s is extremely complex and relies on determining the magnitude of arching in the embankment fill and the tension in a geosynthetic reinforcement layer at the base of the embankment. Several design methods are available for estimating the magnitude of arching and the tension in the geosynthetic reinforcement. Naughton (2007) showed that the magnitude of arching and therefore the tension in the geosynthetic reinforcement can be estimated based on the concept of critical height. The critical height is a function of the frictional characteristics of the embankment fill and pile-pile cap spacing at the base of the embankment. An instrumented laboratory 1:3 scaled model of typical piled embankment geometries was used to investigate the influence of the embankment fill material properties on the location the critical height. Three sand types with different strength and gradation characteristics were examined. The model was used to quantify the unrestrained arching in the model embankment. The influence of the angle of internal friction on the location of the critical height was investigated and the critical height was found to increase as the angle of internal friction increased. It was observed that when the peak angle of friction was used in the method suggested by Naughton (2007) that good agreement was found with the experimental data.

A method is presented for the design of steep slopes constructed from low-permeability fill reinforced with draining geogrids. A parametric variation of the drainage characteristics of the fill and the geometry of the steep slope indicates that the vertical spacing and the length of the reinforcement are important design considerations. For typical values of the coefficients of volume compressibility and consolidation in low-permeability fills an optimum vertical spacing of the reinforcement of 0.5 m was determined. At a vertical spacing of 0.5 m a draining geogrid will dissipate pore water pressures, over the full range of likely drainage properties encountered in low-permeability fills, inside 24 hours. Furthermore, at this vertical spacing the required transmissivity of the geogrids is independent of the length of the draining geogrid. Finally it is economically viable to manufacture draining geogrids with the drainage characteristics required to dissipate pore water pressures in low-permeability fills.

Determining the strength of soil containing fibres is problematic. The presence of discrete fibres reinforces the soil to such a degree that the determined value is not a true representation of the global strength. In this study cone penetration and plate bearing testing in a geotechnical centrifuge were used to determine the undrained shear strength of fibre reinforced clay. Plate bearing testing was performed on foundations of different shape and geometry. The undrained shear strength was back calculated from the foundation bearing resistance. Both inflight cone penetration and plate bearing test results showed an increase in undrained shear strength with fibre content. No optimum fibre content was found in the testing undertaken. Once a threshold fibre content of between 3-4% was exceeded a two ‒ threefold increase in strength was observed. Overall, good correlation was found between the undrained shear strength measured with the cone penetrator and that back calculated from the plate bearing testing.

Research has shown that geotechnical engineers and the construction industry in general like to think of themselves as working to benefit the environment and providing for future generations in tandem, if not always in harmony, with nature. However, engineers still prefer to work with high quality and expensive imported granular fill, rather than seriously consider using recycled spoil generated as part of the construction activity. Engineers can design reinforced soil structures using marginal poorly drained fills without the need to import granular fill, which has both cost and environmental implications. Marginal fills typically have high silt and/or clay contents which, when loaded, have the potential to generate excess pore water pressures in the structural backfill. Poor drainage in the structural fill reduces the available strength of the fill, thus reducing the bond between the fill and the geogrid reinforcement. Therefore, to use marginal fill efficiently, adequate drainage must be provided in the reinforced soil structure. By using a novel geocomposite that combines reinforcement and draining into a geogrid sustainable and environmentally friendly slopes and walls can be designed and constructed. During 2015, 2016 and 2017 an innovative design methodology and approach for the construction of reinforced slopes and walls, up to 15m in height, with low-permeability fills was successfully constructed in the UK. The design and construction experience gained in the UK is presented to confirm the effectiveness of the system and given engineers confidence in the use of marginal fills in reinforced soil systems.

Cosmetic and personal care products are globally used and often applied directly on the human skin. According to a recent survey in Europe, the market value of cosmetic and personal care products in Western Europe reached about 84 billion euros in 2018 and are predicted to increase by approximately 6% by the end of 2020. With these significant sums of money spent annually on cosmetic and personal care products, along with chemical surfactants being the main ingredient in a number of their formulations, of which many have been reported to have the potential to cause detrimental effects such as allergic reactions and skin irritations to the human skin; hence, the need for the replacement of chemical surfactants with other compounds that would have less or no negative effects on skin health. Biosurfactants (surfactants of biological origin) have exhibited great potential such as lower toxicity, skin compatibility, protection and surface moisturizing effects which are key components for an effective skincare routine. This review discusses the antimicrobial, skin surface moisturizing and low toxicity properties of glycolipid and lipopeptide biosurfactants which could make them suitable substitutes for chemical surfactants in current cosmetic and personal skincare pharmaceutical formulations. Finally, we discuss some challenges and possible solutions for biosurfactant applications.

One current strategy to deal with the serious issue of antibiotic resistance is to use biosurfactants, weak antimicrobials in their own right, with antibiotics in order to extend the efficacy of antibiotics. Although an adjuvant effect has been observed, the underlying mechanisms are poorly understood. To investigate the nature of the antibiotic and biosurfactant interaction, we undertook a scanning electron microscopy (SEM) and atomic force microscopy (AFM) microscopic study of the effects of the tetracycline antibiotic, combined with sophorolipid and rhamnolipid biosurfactants, on Methicillin-resistant Staphylococcus aureus using tetracycline concentrations below and above the minimum inhibitory concentration (MIC). Control and treated bacterial samples were prepared with an immersion technique by adsorbing the bacteria onto glass substrates grafted with the poly-cationic polymer polyethyleneimine. Bacterial surface morphology, hydrophobic and hydrophilic surface characters as well as the local bacterial cell stiffness were measured following combined antibiotic and biosurfactant treatment. The sophorolipid biosurfactant stands alone insofar as, when used with the antibiotic at sub-MIC concentration, it resulted in bacterial morphological changes, larger diameters (from 758 ± 75 to 1276 ± 220 nm, p-value = 10-4) as well as increased bacterial core stiffness (from 205 ± 46 to 396 ± 66 mN/m, p-value = 5 × 10-5). This investigation demonstrates that such combination of microscopic analysis can give useful information which could complement biological assays to understand the mechanisms of synergy between antibiotics and bioactive molecules such as biosurfactants.

Enterococci are robust gram-positive bacteria that are found in a variety of surroundings and that cause a significant number of healthcare-associated infections. The genus possesses a high-efficiency pheromone-responsive plasmid (PRP) transfer system for genetic exchange that allows antimicrobial-resistance determinants to spread within bacterial populations. The pCF10 plasmid system is the best characterised, and although other PRP systems are structurally similar, they lack exact functional homologues of pCF10-encoded genes. In this review, we provide an overview of the enterococcal PRP systems, incorporating functional details for the less-well-defined systems. We catalogue the virulence-associated elements of the PRPs that have been identified to date, and we argue that this reinforces the requirement for elucidation of the less studied systems.

Salmonella enterica serovar Rissen has been recognised as a common serovar in humans and pigs around the world. This study investigated S. Rissen prevalence in pigs slaughtered in Northern Ireland additionally looking at antibiotic susceptibility profiles, genetic profiles and plasmid profiles to provide information on an emerging non-typhoid Salmonella serotype with the potential to cause disease in humans. S. Rissen were isolated on five separate sampling occasions from both the boning hall and slaughter line of a randomly selected single pig abattoir in Northern Ireland (NI). Following antibiotic susceptibility testing against 16 antibiotics, all S. Rissen isolates were identified as susceptible to 15 antibiotics but resistant to tetracycline (R-type: Te). Of the 29 S. Rissen, 27 were isolated from pigs originating in NI and two S. Rissen were isolated from pigs originating in the Republic of Ireland (RoI). The combined results of the PFGE and plasmid profiling analyses were capable of subdividing the S. Rissen isolates into three distinct groups. The data suggests that S. Rissen is an emerging serovar in Northern Ireland and continued surveillance of this serovar is warranted as it has the potential to cause disease in humans.

This paper presents and discusses a student-focussed, small group learning and teaching, and school-wide initiative designed for first year undergraduate students in support of their transition into higher education. The details of the underlying pedagogic strategy, as well as the organisation (small group teaching environment), the content of applied learning and teaching approaches (including assignment and assessment), and the outcomes of the initiative evaluation (including required human resources) based on data collected from five consecutive academic years are presented. This paper aims to serve as an example of a pedagogic approach that facilitates student transition into Higher Education aimed at practitioners, especially those involved in delivery and/or those who are engaged or responsible for undergraduate curricula development. The findings are discussed in terms of implications for curricular changes to support students’ engagement at this early stage of their university career.

The Enterococci are a resilient collection of species, found in the human intestine, river sediment and even certain cheeses. Human infection by this genus is dominated by E. faecalis and E. faecium. Vancomycin resistant enterococci (VRE) are associated with higher mortality rates over non-VRE strains. Enterococci can utilise the highly efficient pheromone responsive plasmid (PRP) system to transfer plasmid DNA between cells. Plasmid containing donor cells respond to small peptide pheromones (7–8 amino acids) and transfer plasmid DNA to pheromone-producing plasmid-free recipient cells. PRP can encode antibiotic resistance (including vancomycin) and virulence enhancing factors. Investigation into the PRP system between donor and recipient E. faecalis environmental isolates has indicated a 40 % decrease in PRP transfer in colder environments. Additionally, PRP efficiencies under other conditions, including in presence of synthetic pheromone peptides, have been calculated. Future assays will utilise pheromone imitative fluorescently labelled synthetic peptides to visualise the pheromone binding receptor (PrgZ) on the E. faecalis donor cell membrane. Later experiments will focus on varying the synthetic pheromone amino acid composition so to interfere with the PRP system machinery, with the aim of reducing PRP transfer efficiency or preventing PRP transfer completely.

Synthetic surfactants are becoming increasingly unpopular in many applications due to previously disregarded effects on biological systems and this has led to a new focus on replacing such products with biosurfactants that are biodegradable and produced from renewal resources. Microbially derived biosurfactants have been investigated in numerous studies in areas including: increasing feed digestibility in an agricultural context, improving seed protection and fertility, plant pathogen control, antimicrobial activity, antibiofilm activity, wound healing and dermatological care, improved oral cavity care, drug delivery systems and anticancer treatments. The development of the potential of biosurfactants has been hindered somewhat by the myriad of approaches taken in their investigations, the focus on pathogens as source species and the costs associated with large-scale production. Here, we focus on various microbial sources of biosurfactants and the current trends in terms of agricultural and biomedical applications.

This case report presents an evaluation of the effect of a work-based placement on the achievements in the final year of the undergraduate degree. We also aimed to assess whether “better” students were opting to take up a placement year and if such a con-founder would have an influence on the effect of placement on final year performance. Retrospective data were collected for ten consecutive cohorts of students who had read for the same full time BSc Hons degree programme with a broad curriculum that offered an optional work-based placement year that was not closely aligned to their academic studies. The results have shown that “better” students opt to take up an optional work-based placement year; however, taking up the placement has not affected students’ final year attainment. This case report supports the notion that for a work-based placement to have a positive effect on students’ final year attainment, the placement must be closely aligned to the discipline of academic study.

This study aimed to assess whether a daily supplementation with Lactobacillus rhamnosus GG (LGG) can alter the body composition as well as bone turnover and the mechanical properties of the bones in systemic immunodeficiency. For this, we use an in vivo model of severe combined immunodeficient (scid) mice supplemented daily with LGG over 8 weeks period. After 8 weeks of the treatment, the animals were assessed by DEXA and then the blood was collected from the hepatic portal vein for bone turnover biomarkers; post-mortem, the caecum was removed for isolation and enumeration of the gut bacteria, while femurs/tibiae were excised for the assessment of their mechanical properties. LGG treatment affected the composition of the gut microbiota at the order level, and it decreased (P = 0.002) total adipose tissue content. Moreover, LGG treatment decreased the stiffness (N/mm) of both femurs (P = 0.008) and tibiae (P = 0.011). To conclude, this study shows that dietary supplementation with LGG in immunodeficient animals can alter mechanical properties of the long bones and concomitantly it can modify host’s total adiposity level.

Background Enterococci are now well recognised for their ability to transfer antibiotic resistance and for their association with nosocomial infections, but less is known regarding their relevance in the wider environment. Enterococcus faecalis and Enterococcus faecium were isolated from a range of agrarian associated sources (low-flow water, septic tank, poultry litter, high flow water, slurry/soil) and were assessed for latent ability to transfer antimicrobial resistance. Results The isolates were tested for phenotypic clumping in the presence of cell-free supernatant from other isolates. Some isolates were identified which demonstrated clumping, indicating that they possessed peptide sex pheromone conjugal machinery. All isolates were also tested for antibiotic resistance phenotypes using both disc diffusion and minimum inhibitory concentration (MIC) assays. These tests revealed that the enterococci demonstrated both phenotypic clumping and antibiotic resistance phenotypes. Based on these selection criteria, the isolates were identified as having the potential for horizontal gene transfer and were used to investigate the transfer of multiple antibiotic resistance phenotypes. Conjugal transfer of antibiotic resistance phenotypes was determined using a solid agar mating method followed by a standard antibiotic selection test resulting in different transfer patterns. An interspecies conjugal transfer of vancomycin resistance from E. faecalis to E. faecium was identified while the remaining reactions were within the same species. Transfer efficiencies ranging from 2 × 10−1 to 2.3 × 10−5 were determined based on the reactions of three donor isolates (MF06036, MF0410 and MF06035) and two recipient isolates (MW01105Rif and ST01109Rif), with the transfer of vancomycin, erythromycin and tetracycline resistance genes. Conclusions The conjugation reactions and selection conditions used in this study resulted in a variety of co-transferred resistance phenotypes suggesting the presence of different mobile elements in the set of natural isolates. This study highlights the potential for extensive horizontal gene transfer in a previously neglected reservoir for enterococci.

In this study we hypothesized that an increase in numbers of beneficial bacteria in the large intestine can affect calcium (Ca) status in the elderly. Adult and aged rats were fed a diet with or without synbiotics for 21 days. Synbiotics increased the numbers of lactobacilli and bifidobacteria in large intestine in both adult and aged rats. The plasma Ca concentration was significantly increased while osteocalcin concentration was significantly decreased only in aged rats fed synbiotics.

Interdigestive intestinal motility, and especially phase III of the migrating myoelectric/motor complex (MMC), is responsible for intestinal clearance and plays an important role in prevention of bacterial overgrowth and translocation in the gut. Yet previous results from gnotobiotic rats have shown that intestinal microflora can themselves affect the characteristics of the myoelectric activity of the gut during the interdigestive state. Given that the composition of the intestinal microflora can be altered by dietary manipulations, we investigated the effect of supplementation of the diet with synbiotics on intestinal microflora structure and the duodenojejunal myoelectric activity in the rat. To reduce animal distress caused by restraint and handling, which can itself affect GI motility, we applied radiotelemetry for duodenojejunal EMG recordings in conscious, freely moving rats. Thirty 16-month-old Spraque-Dawley rats were used. The diet for 15 rats (E group) was supplemented with chicory inulin, Lactobacillus rhamnosus and Bifidobacterium lactis. The remaining 15 rats were fed control diet without supplements (C group). Three rats from each group were implanted with three bipolar electrodes positioned at 2, 14 and 28 cm distal to the pylorus. After recovery, two 6 h recordings of duodenojejunal EMG were carried out on each operated rat. Subsequently, group C rats received feed supplements and group E rats received only control diet for 1 week, and an additional two 6 h recordings were carried out on each of these rats. Non-operated C and E rats were killed and samples of GI tract were collected for microbiological analyses. Supplementation of the diet with the pro- and prebiotics mixture increased the number of bifidobacteria, whereas it decreased the number of enterobacteria in jejunum, ileum, caecum and colon. In both caecum and colon, the dietary supplementation increased the number of total anaerobes and lactobacilli. Treatment with synbiotics increased occurrence of phase III of the MMC at all three levels of the small intestine. The propagation velocity of phase III in the whole recording segment was also increased from 3.7 +/- 0.2 to 4.4 +/- 0.2 cm min(-1) by dietary treatment. Treatment with synbiotics increased the frequency of response potentials of the propagated phase III of the MMC at both levels of the jejunum, but not in the duodenum. In both parts of the jejunum, the supplementation of the diet significantly decreased the duration of phase II of the MMC, while it did not change the duration of phase I and phase III. Using the telemetry technique it was demonstrated that changes in the gastrointestinal microflora exhibited an intestinal motility response and, more importantly, that such changes can be initiated by the addition of synbiotics to the diet.

A 2 x 2 factorial experiment with pigs was undertaken to investigate the effect of particle size (fine and coarse) and feed processing (pelleted and nonpelleted) on morphological characteristics in the small intestine, cecum, and colon of pigs and on the adhesion of Salmonella enterica serovar Typhimurium DT12 to the ileum in vitro. Ninety-six pigs (average BW = 33 +/- 7 kg) were fed the experimental diets. After 4 wk, 24 pigs were selected (six pigs per diet) and euthanized, and tissue samples were taken from the mid and distal small intestine, cecum, and distal colon. The effects of particle size and feed processing on villus height and crypt depth in the small intestine were minor. Feeding coarse diets increased (P = 0.05) the crypt depth in the colon. The crypt depth was 420 +/- 12 and 449 +/- 12 microm in pigs fed finely and coarsely ground feed, respectively. Pigs fed pelleted diets had a larger (P = 0.01) staining area for neutral mucins, as well as for acidic and sulfomucins on the villi of the distal small intestine than pigs fed nonpelleted diets. The area was 41, 46, and 33% larger for neutral, acidic, and sulfomucins, respectively. The mucin-staining areas of the crypts in the cecum and the colon were not affected by the experimental diets. Examination of lectin binding characteristics of the distal small intestine and the cecum did not reveal any differences between the experimental diets. Using a pig intestine organ culture model, Salmonella adhered less (P < 0.05) to the ileal tissue of pigs fed the nonpelleted diets than to those fed pelleted diets; the adherence was 60% less in these pigs. Results of this study suggest that pigs fed pelleted diets secrete mucins that are capable of binding Salmonella enterica serovar Typhimurium DT12 and thereby allowing for colonization. Therefore, pigs fed a nonpelleted diet are better protected against Salmonella infections than pigs fed a pelleted diet.

A two-by-two factorial experiment with pigs was conducted to study the effect of feed grinding (fine and coarse) and feed processing (pelleted and nonpelleted) on physicochemical properties, microbial populations, and survival of Salmonella enterica serovar Typhimurium DT12 in the gastrointestinal tracts of pigs. Results demonstrated a strong effect of diet on parameters measured in the stomachs of the pigs, whereas the effect was less in the other parts of the gastrointestinal tract. Pigs fed the coarse nonpelleted (C-NP) diet showed more solid gastric content with higher dry matter content than pigs fed the fine nonpelleted (F-NP), coarse pelleted (C-P), or fine pelleted (F-P) diet. Pigs fed the C-NP diet also showed significantly increased number of anaerobic bacteria ( P < 0.05), increased concentrations of organic acids, and reduced pH in the stomach. In addition, pigs fed the C-NP diet showed increased in vitro death rate of S. enterica serovar Typhimurium DT12 in content from the stomach ( P < 0.001). Pigs fed the C-NP diet had a significantly higher concentration of undissociated lactic acid in gastric content than pigs fed the other diets ( P < 0.001). A strong correlation between the concentration of undissociated lactic acid and the death rate of S. enterica serovar Typhimurium DT12 was found. In the distal small intestine, cecum, and midcolon, significantly lower numbers of coliform bacteria were observed in pigs fed the coarse diets than in pigs fed the fine diets ( P < 0.01). Pigs fed the C-NP diet showed the lowest number of coliform bacteria in these segments of the gastrointestinal tract. Pigs fed the coarse diets showed increased concentration of butyric acid in the cecum ( P < 0.05) and colon ( P < 0.10) compared with pigs fed the fine diets. It was concluded that feeding a coarsely ground meal feed to pigs changes the physicochemical and microbial properties of content in the stomach, which decreases the survival of Salmonella during passage through the stomach. In this way the stomach acts as a barrier preventing harmful bacteria from entering and proliferating in the lower part of the gastrointestinal tract.

Nitric oxide and S-nitrosothiols modulate a variety of important physiological activities. In vascular cells, agents that release NO and donate nitrosonium cation (NO+), such as S-nitrosoglutathione, are potent inducers of the antioxidant protein heme oxygenase 1 (HO-1) (Foresti, R., Clark, J. E., Green, C. J., and Motterlini, R. (1997)J. Biol. Chem. 272, 18411–18417; Motterlini, R., Foresti, R., Bassi, R., Calabrese, V., Clark, J. E., and Green, C. J. (2000) J. Biol. Chem. 275, 13613–13620). Here, we report that Angeli's salt (AS) (0.25–2 mm), a compound that releases nitroxyl anion (NO−) at physiological pH, induces HO-1 mRNA and protein expression in a concentration- and time-dependent manner, resulting in increased heme oxygenase activity in rat H9c2 cells. A time course analysis revealed that NO−-mediated HO-1 expression is transient and gradually disappears within 24 h, in accordance with the short half-life of AS at 37 °C (t = 2.3 min). Interestingly, multiple additions of AS at lower concentrations (50 or 100 μm) over a period of time still promoted a significant increase in heme oxygenase activity. Experiments performed using a NO scavenger and the NO electrode confirmed that NO−, not NO, is the species involved in HO-1 induction by AS; however, the effect on heme oxygenase activity can be amplified by accelerating the rate of NO− oxidation.N-Acetylcysteine almost completely abolished AS-mediated induction of HO-1, whereas a glutathione synthesis inhibitor (buthionine sulfoximine) significantly decreased heme oxygenase activation by AS, indicating that sulfydryl groups are crucial targets in the regulation of HO-1 expression by NO−. We conclude that NO−, in analogy with other reactive nitrogen species, is a potent inducer of heme oxygenase activity and HO-1 protein expression. These findings indicate that heme oxygenase can act both as a sensor to and target of redox-based mechanisms involving NO and extend our knowledge on the biological function of HO-1 in response to nitrosative stress.

During the past two decades, the identification of new scientific developments to improve outcomes in gastrointestinal disorders has been attractive to many researchers. Pharmaceutical industries are now more motivated to introduce novel therapeutic remedies in the treatment of gastrointestinal disorders. Such disorders have increased at an exponential rate in various patient communities and both natural and synthetic compounds have been investigated for their potential biological activity in the treatment of these gastrointestinal disorders. Several interesting works were assessed in this special issue. Amongst them five articles were chosen based on their critical findings in gastrointestinal disorders. The effects of rikkunshito on the decrease in food intake were assessed after induction of stress in mice and showed improvement in the decrease of food intake probably via serotonin 2B receptor antagonism of isoliquiritigenin. In another work, the preventive effect of inositol hexaphosphate extract of rice bran on colon cancer was assessed. The results showed significant reduction in the expression of β-catenin and COX-2 in colon tumors. The Schiff base metal derivatives also may enhance the expression of HSP70 and suppress the expression of BAX proteins in an acute hemorrhagic gastric ulcer model. Another animal study assessed the hepatoprotective activity of the ethanolic extract of rhizomes of Z. officinale against thioacetamide-induced hepatotoxicity in rats. The floating dosage form of an anticancer drug was prepared in another study entitled “Preparation and Characterization of a Gastric Floating Dosage Form of Capecitabine.” The work characterized the sustained release tablet in terms of total floating time, dissolution, friability, hardness, drug content, and weight uniformity to compare the prepared formulation with the commercial tablet in terms of drug release, and to evaluate the stability of the formulation. By presenting these articles, we hope that this issue incorporates new scientific evidence and emerging developments as the basis of rational treatment in medicinal practice using novel therapeutics (natural or synthetic compounds) in the treatment of gastrointestinal disorders. Mahmood Ameen Abdulla Ibrahim Banat Patrick Naughton

Rats were dosed for 6 days with purified SEF 21 fimbriae of Salmonella enterica serovar Enteritidis 10360. The levels of fimbriae in gut contents associated with tissues and in the faeces were quantified by direct non-competitive ELISA. SEF 21 was distributed throughout the gut. The majority was found in the large intestine where it was primarily in the luminal contents. In contrast, a high proportion of SEF 21 detected in the ileum, the main site of salmonella colonisation and invasion, was tissue-bound. Thus, purified SEF 21 survived intestinal passage and associated with the stomach and gastrointestinal tract in a pattern similar to that found with whole Salmonella cells.

An in vitro intestinal tissue model was developed for the investigation of bacterial association in the pig small intestine under different dietary regimes. In preliminary experiments, jejunal and ileal tissue was taken from Danish Landrace pigs fed standard diet and inoculated with either Salmonella or nonpathogenic Escherichia coli strains. Higher numbers of salmonellae associated with the ileal tissues, but the numbers did not reach significance. Hence, jejunal sections were inoculated with nonpathogenic E. coli and ileal sections were inoculated with salmonellae in the presence of mannose or commercial nondigestible oligosaccharides (NDO) at 2.5%. There was a significant decrease in E. coli associated with the jejunum in the presence of mannose ( P < 0.05). Furthermore, in pigs fed a diet supplemented with commercial NDO at 4% there was a significant reduction in the numbers of E. coli in jejunal organ cultures of pigs fed the FOS diet ( P < 0.05). There was a reduction, though not a significant one, in the association of Salmonella sp. to the ileal sections of pigs fed the commercial FOS diet. The feeding of commercial GOS or its addition to organ cultures did not affect E. coli or Salmonella numbers.

The use of antibiotics in birds and animals intended for human consumption within the European Union (EU) and elsewhere has been subject to regulation prohibiting the use of antimicrobials as growth promoters and the use of last resort antibiotics in an attempt to reduce the spread of multi-resistant Gram negative bacteria. Given the inexorable spread of antibiotic resistance there is an increasing need for improved monitoring of our food. Using selective media, Gram negative bacteria were isolated from retail chicken of UK-Intensively reared (n=27), Irish-Intensively reared (n=19) and UK-Free range (n=30) origin and subjected to an oligonucleotide based array system for the detection of 47 clinically relevant antibiotic resistance genes (ARGs) and two integrase genes. High incidences of β-lactamase genes were noted in all sample types, acc (67%), cmy (80%), fox (55%) and tem (40%) while chloramphenicol resistant determinants were detected in bacteria from the UK poultry portions and were absent in bacteria from the Irish samples. Denaturing Gradient Gel Electrophoresis (DGGE) was used to qualitatively analyse the Gram negative population in the samples and showed the expected diversity based on band stabbing and DNA sequencing. The array system proved to be a quick method for the detection of antibiotic resistance gene (ARG) burden within a mixed Gram negative bacterial population.

The relationship between duodenal myoelectric activity and liquid food intake was investigated in eight conscious piglets with bipolar electrodes implanted on the wall of the duodenum. Piglets were fed with commercial milk formula and the effects of volume and feed dry matter intake on duodenal myoelectric activity were measured by means of an analog-digital recording system. The cyclic pattern of duodenal myoelectric activity, i.e. the migrating myoelectric complex (MMC), was preserved when piglets received small meals at regular intervals. The duration of the whole MMC cycle, as well as the duration of phase II of the MMC, were only weakly correlated (r = 0.2, P - 1 of live body mass (LBM) as well as an increase of volume of a meal from 13 to 26 ml kg- 1 LBM significantly increased the duration of the MMC cycles (P P The present study shows that in piglets fed with a liquid diet the upper gut motility response to food is similar to that previously observed in adult animals. Udgivelsesdato: March The relationship between duodenal myoelectric activity and liquid food intake was investigated in eight conscious piglets with bipolar electrodes implanted on the wall of the duodenum. Piglets were fed with commercial milk formula and the effects of volume and feed dry matter intake on duodenal myoelectric activity were measured by means of an analog-digital recording system.The cyclic pattern of duodenal myoelectric activity, i.e. the migrating myoelectric complex (MMC), was preserved when piglets received small meals at regular intervals. The duration of the whole MMC cycle, as well as the duration of phase II of the MMC, were only weakly correlated (r = 0.2, P - 1 of live body mass (LBM) as well as an increase of volume of a meal from 13 to 26 ml kg- 1 LBM significantly increased the duration of the MMC cycles (P P The present study shows that in piglets fed with a liquid diet the upper gut motility response to food is similar to that previously observed in adult animals

Publisher Summary This chapter discusses the models used to study the pathogenesis of salmonellosis. Salmonella spp. can cause two types of disease in humans: an acute gastroenteritis or systemic typhoid disease. Many animal models have been adopted to study salmonellosis. Few appear to effectively model the overall infection in humans. There is a great variation in the levels of colonization, invasion, and systemic spread; and in the incidence of bacteremia and the effects of the pathogen on long-term health. Factors such as susceptibility to the disease, the infective dose required, and the ease of non-invasive monitoring or ready availability of species-specific reagents often influence the choice of model. The mouse, rat, and pig models are widely used. The chapter evaluates their strengths and weaknesses as models of salmonellosis. It is suggested that malnutrition severely impairs intestinal cell metabolism and reduces the efficacy of the local and systemic immune systems. This may greatly increase the permeability of the gut to the pathogen or its products and predispose the malnourished animal to salmonellosis.

Carbon monoxide, which is generated in mammals during the degradation of heme by the enzyme heme oxygenase, is an important signaling mediator. Transition metal carbonyls have been recently shown to function as carbon monoxide–releasing molecules (CO-RMs) and to elicit distinct pharmacological activities in biological systems. In the present study, we report that a water-soluble form of CO-RM promotes cardioprotection in vitro and in vivo. Specifically, we found that tricarbonylchloro(glycinato)ruthenium(II) (CORM-3) is stable in water at acidic pH but in physiological buffers rapidly liberates CO in solution. Cardiac cells pretreated with CORM-3 (10 to 50 μmol/L) become more resistant to the damage caused by hypoxia-reoxygenation and oxidative stress. In addition, isolated hearts reperfused in the presence of CORM-3 (10 μmol/L) after an ischemic event displayed a significant recovery in myocardial performance and a marked and significant reduction in cardiac muscle damage and infarct size. The cardioprotective effects mediated by CORM-3 in cardiac cells and isolated hearts were totally abolished by 5-hydroxydecanoic acid, an inhibitor of mitochondrial ATP-dependent potassium channels. Predictably, cardioprotection is lost when CORM-3 is replaced by an inactive form (iCORM-3) that is incapable of liberating CO. Using a model of cardiac allograft rejection in mice, we also found that treatment of recipients with CORM-3 but not iCORM-3 considerably prolonged the survival rate of transplanted hearts. These data corroborate the notion that transition metal carbonyls could be used as carriers to deliver CO and highlight the bioactivity and potential therapeutic features of CO-RMs in the mitigation of cardiac dysfunction. The full text of this article is available online at http://www.circresaha.org.

Bilirubin (BR) and biliverdin (BV), two metabolites produced during haem degradation by haem oxygenase, possess strong antioxidant activities toward peroxyl radical, hydroxyl radical and hydrogen peroxide. Considering the importance attributed to nitric oxide (NO) and its congeners in the control of physiological and pathophysiological processes, we examined the interaction of BR and BV with NO and NO-related species in vitro. Exposure of BR and BV to agents that release NO or nitroxyl resulted in a concentration- and time-dependent loss of BR and BV, as assessed by high performance liquid chromatography. Peroxynitrite, a strong oxidant derived from the reaction of NO with superoxide anion, also showed high reactivity toward BR and BV. The extent of BR and BV consumption largely depended on the NO species being analysed and on the half-lives of the pharmacological compounds considered. Of major importance, BR and BV decomposition occurred also in the presence of pure NO under anaerobic conditions, confirming the ability of bile pigments to scavenge the gaseous free radical. Increasing concentrations of thiols prevented BR consumption by nitroxyl, indicating that bile pigments and thiol groups can compete and/or synergise the cellular defence against NO-related species. In view of the high inducibility of haem oxygenase-1 by NO-releasing agents in different cell types, the present findings highlight novel anti-nitrosative characteristics of BR and BV suggesting a potential function for bile pigments against the damaging effects of uncontrolled NO production.

Nitric oxide and S-nitrosothiols modulate a variety of important physiological activities. In vascular cells, agents that release NO and donate nitrosonium cation (NO(+)), such as S-nitrosoglutathione, are potent inducers of the antioxidant protein heme oxygenase 1 (HO-1) (Foresti, R., Clark, J. E., Green, C. J., and Motterlini, R. (1997) J. Biol. Chem. 272, 18411-18417; Motterlini, R., Foresti, R., Bassi, R., Calabrese, V., Clark, J. E., and Green, C. J. (2000) J. Biol. Chem. 275, 13613-13620). Here, we report that Angeli's salt (AS) (0.25-2 mm), a compound that releases nitroxyl anion (NO(-)) at physiological pH, induces HO-1 mRNA and protein expression in a concentration- and time-dependent manner, resulting in increased heme oxygenase activity in rat H9c2 cells. A time course analysis revealed that NO(-)-mediated HO-1 expression is transient and gradually disappears within 24 h, in accordance with the short half-life of AS at 37 degrees C (t(12) = 2.3 min). Interestingly, multiple additions of AS at lower concentrations (50 or 100 microm) over a period of time still promoted a significant increase in heme oxygenase activity. Experiments performed using a NO scavenger and the NO electrode confirmed that NO(-), not NO, is the species involved in HO-1 induction by AS; however, the effect on heme oxygenase activity can be amplified by accelerating the rate of NO(-) oxidation. N-Acetylcysteine almost completely abolished AS-mediated induction of HO-1, whereas a glutathione synthesis inhibitor (buthionine sulfoximine) significantly decreased heme oxygenase activation by AS, indicating that sulfydryl groups are crucial targets in the regulation of HO-1 expression by NO(-). We conclude that NO(-), in analogy with other reactive nitrogen species, is a potent inducer of heme oxygenase activity and HO-1 protein expression. These findings indicate that heme oxygenase can act both as a sensor to and target of redox-based mechanisms involving NO and extend our knowledge on the biological function of HO-1 in response to nitrosative stress.

The involvement of type 1 fimbriae in colonisation of the rat gastrointestinal tract in vivo was investigated with Salmonella enterica serotype Enteritidis LA5 and a mutant of LA5 denoted EAV3 unable to elaborate type 1 fimbriae (SEF 21). Rats were given a single dose of LA5 or EAV3 or a 1:1 mixture of both. LA5 was found in higher numbers in the stomach and small intestine than EAV3 at 6 h after infection with a single strain, but not after 6 days. LA5 did not out-compete EAV3 when the strains were administered together. Indeed, after 6 and 21 days, EAV3 was found in the distal small intestine and large intestine in far higher numbers than LA5. These findings suggest that SEF 21 have an important role(s) in the early stages of infection in vivo. However, SEF 21 expression may disadvantage the pathogen in the longer term as indicated by EAV3 out- competing LA5 in the gut at 21 days.

The batch culture system included six bioreactors. Three bioreactors containing stomach slurry were maintained at pH 4.5 and 6 respectively. Bioreactors containing small intestine slurry were maintained at pH 5.6 and 7 respectively. The bioreactors were inoculated with 10 ml of viable Salmonella. The bioreactors were maintained for 6 hours. Samples of 10 ml were taken at 0 time and at 1, 2, 4 and 6 hours. The samples were analysed for the presence of Salmonella and SCFA. In the stomach samples Salmonella numbers increased at pH 6 but fell at pH 4. In the small intestine sample Salmonella numbers increased at pH 6 and 7. In terms of SCFA production, in the stomach, with samples at pH 6 there was little change in the amounts of lactate, succinate and formate to that detected at 0 time, however levels of acetate did increase slightly. In the small intestine samples levels of succinate and formate increased slightly up to 4 hours, levels of acetate increased significantly from 0 to 6 hours. In terms of the specific growth rates of the individual strains, both strains grew at pH 6 in the stomach content and to a greater extent in the small intestinal content. A bactericidal effect was observed at pH 4 in the stomach content while neither killing nor growth occurred at pH 5 either in the stomach or the small intestine content. Both strains grew well in the small intestine content at pH 7, showing generation times of up to 24 min.

In a series of experiments rats were dosed with purified type 1 fimbriae from Salmonella enterica var Enteritidis or with fimbriated cultures of either S. enterica var Typhimurium or S. enterica var Enteritidis. Paraffin-wax embedded histological sections of jejunal and ileal tissue were taken and stained by the streptavidin biotin complex (sABC) staining technique for the detection of salmonella and type 1 fimbriae. On oral infection with Enteritidis and Typhimurium both bacteria were shown to be closely associated with the rat ileal epithelium and expressed type 1 fimbriae, thus clearly demonstrating that type 1 fimbriae are expressed by salmonellae in vivo. Moreover, association with the ileum was also shown to occur when purified type 1 fimbriae were orally administered to rats. Our results suggest that type 1 fimbriae alone or in combination with other fimbriae may play an important role in the early stages of infection with these pathogenic bacteria.

Salmonellosis in the rat has many similarities with the disease in humans, with the ileum thought to be the main site of colonization/invasion in both species. Thus, the rat may be a useful way to study the mechanism of infection by these pathogenic bacteria. A series of infection trials carried out with Hooded Lister rats showed that a salmonella infection persisted for an extended period of time and that salmonellae bind to the small intestinal epithelium as early as 4 h after intragastric intubation. Reinfection from the large intestine may not therefore initially play a significant role in the salmonella infection process. The rat model may therefore provide a means to test in vivo interventionist strategies, designed to block binding of the pathogens in the gastrointestinal tract.