In vitro susceptibility of Brachyspira hyodysenteriae strains isolated in pigs in northern Italy between 2005 and 2022.

Pleuromutilins (tiamulin and valnemulin) are often used to treat swine dysentery due to recurrent resistance to macrolides and lincosamides. Recently, reduced susceptibility of B. hyodysenteriae to pleuromutilin has been reported. 536 strains of B. hyodysenteriae were isolated from symptomatic pigs weighing 30-150 kg in northern Italy between 2005 and 2022. B. hyodysenteriae was isolated by standard methods and confirmed by PCR. The minimum inhibitory concentration (MIC) to doxycycline, lincomycin, tiamulin, tylosin, tylvalosine and valnemulin was evaluated according to CLSI procedures and MIC data were reported as MIC 50 and MIC 90. The temporal trend of the MIC values was evaluated by dividing the data into two groups (2005-2013 and 2014-2022). Comparison of the distribution in frequency classes in the two periods was performed using Pearson's chi-squared test (p < 0.01). MIC 50 was close to the highest values tested for lincomycin and tylosin, while MIC 90 was close to the highest values tested for all antibiotics. 71.7% of the strains were susceptible to tylvalosin, while 75%-80.4% had reduced susceptibility to valnemulin and tiamulin, respectively. The difference in the distribution of MIC classes was statistically significant in the two periods for doxycycline, tiamulin, tylvalosin and valnemulin, and more MIC classes above the epidemiological cut-off were observed in 2014-2022 compared with 2005-2013. The evaluation of the trends during the period considered shows a decreasing rate of wild-type strains with MIC values below the epidemiological cut-off over time and confirms the presence of resistant strains in northern Italy.

Antimicrobial susceptibility of U.S. porcine Brachyspira isolates and genetic diversity of B. hyodysenteriae by multilocus sequence typing.

Swine dysentery, caused by Brachyspira hyodysenteriae and the newly recognized Brachyspira hampsonii in grower-finisher pigs, is a substantial economic burden in many swine-rearing countries. Antimicrobial therapy is the only commercially available measure to control and prevent Brachyspira-related colitis. However, data on antimicrobial susceptibility trends and genetic diversity of Brachyspira species from North America is limited. We evaluated the antimicrobial susceptibility profiles of U.S. Brachyspira isolates recovered between 2013 and 2022 to tiamulin, tylvalosin, lincomycin, doxycycline, bacitracin, and tylosin. In addition, we performed multilocus sequence typing (MLST) on 64 B. hyodysenteriae isolates. Overall, no distinct alterations in the susceptibility patterns over time were observed among Brachyspira species. However, resistance to the commonly used antimicrobials was seen sporadically with a higher resistance frequency to tylosin compared to other tested drugs. B. hampsonii was more susceptible to the tested drugs than B. hyodysenteriae and B. pilosicoli. MLST revealed 16 different sequence types (STs) among the 64 B. hyodysenteriae isolates tested, of which 5 STs were previously known, whereas 11 were novel. Most isolates belonged to the known STs: ST93 (n = 32) and ST107 (n = 13). Our findings indicate an overall low prevalence of resistance to clinically important antimicrobials other than tylosin and bacitracin, and high genetic diversity among the clinical Brachyspira isolates from pigs in the United States during the past decade. Further molecular, epidemiologic, and surveillance studies are needed to better understand the infection dynamics of Brachyspira on swine farms and to help develop effective control measures.

Improving the consistency of experimental swine dysentery inoculation strategies.

Swine dysentery (SD) caused by pathogenic Brachyspira spp. is an economic challenge for the swine industry. In research settings, experimental reproduction of swine dysentery typically relies on intragastric inoculation which has shown variable success. This project aimed to improve the consistency of the experimental inoculation protocol used for swine dysentery in our laboratory. Over six experiments, we evaluated the influence of group housing in inoculated pigs using a frozen-thawed broth culture of strongly hemolytic B. hyodysenteriae strain D19 (Trial A), compared the relative virulence of B. hyodysenteriae strains D19 and G44 (Trial B), compared inoculum volumes (50 mL vs 100 mL) for G44 and B. hampsonii 30446 (Trial C), and performed three independent trials evaluating intragastric inoculation using different oral inoculation methods: oral feed balls (Trial D), and oral syringe bolus of 100 mL (Trial E) or 300 mL (Trial F). Intragastric inoculation with a fresh broth culture of B. hyodysenteriae strain G44 resulted in a shorter incubation period and a higher proportionate duration of mucohemorrhagic diarrhea (MMHD) compared to D19. Intragastric inoculation with either 50 or 100 mL of B. hampsonii 30446 or B. hyodysenteriae (G44) were statistically equivalent. Oral inoculation with 100 mL or 300 mL also yielded similar results to intragastric inoculation but was more expensive due to the additional work and supplies associated with syringe training. Our future research will use intragastric inoculation with 100 mL of a fresh broth culture containing B. hyodysenteriae strain G44 as it yields a high incidence of mucohaemorrhagic diarrhea with a reasonable cost.

Predictive Power of Long-Read Whole-Genome Sequencing for Rapid Diagnostics of Multidrug-Resistant Brachyspira hyodysenteriae Strains.

Infections with Brachyspira hyodysenteriae, the etiological agent of swine dysentery, result in major economic losses in the pig industry worldwide. Even though microbial differentiation of various Brachyspira species can be obtained via PCR, no quick diagnostics for antimicrobial susceptibility testing are in place, which is mainly due to the time-consuming (4 to 7 days) anaerobic growth requirements of these organisms. Veterinarians often rely on a clinical diagnosis for initiating antimicrobial treatment. These treatments are not always effective, which may be due to high levels of acquired resistance in B. hyodysenteriae field isolates. By using long-read-only whole-genome sequencing and a custom-trained Bonito base-calling model, 81 complete B. hyodysenteriae genomes with median Q51 scores and 99% completeness were obtained from 86 field strains. This allowed the assessment of the predictive potential of genetic markers in relation to the observed acquired resistance phenotypes obtained via agar dilution susceptibility testing. Multidrug resistance was observed in 77% and 21% of the tested strains based on epidemiological cutoff and clinical breakpoint values, respectively. The predictive power of genetic hallmarks (genes and/or gene mutations) for antimicrobial susceptibility testing was promising. Sensitivity and specificity for tiamulin [tva(A) and 50SL3N148S, 99% and 67%], valnemulin [tva(A), 97% and 92%), lincomycin (23SA2153T/G and lnuC, 94% and 100%), tylvalosin (23SA2153T/G, 99% and 93%), and doxycycline (16SG1026C, 93% and 87%) were determined. The predictive power of these genetic hallmarks is promising for use in sequencing-based workflows to speed up swine dysentery diagnostics in veterinary medicine and determine proper antimicrobial use. IMPORTANCE Diagnostics for swine dysentery rely on the identification of Brachyspira species using molecular techniques. Nevertheless, no quick diagnostic tools are available for antimicrobial susceptibility testing due to extended growth requirements (7 to 14 days). To enable practitioners to tailor antimicrobial treatment to specific strains, long-read sequencing-based methods are expected to lead to rapid methods in the future. Nevertheless, their potential implementation should be validated extensively. This mainly implies assessing sequencing accuracy and the predictive power of genetic hallmarks in relation to their observed (multi)resistance phenotypes.

Porcine intestinal glycosphingolipids recognized by Brachyspira hyodysenteriae.

Swine dysentery caused by Brachyspira hyodysenteriae is a disease present worldwide with an important economic impact on the farming business, resulting in an increased use of antibiotics. In the present study, we investigated the binding of B. hyodysenteriae to glycosphingolipids from porcine small intestinal epithelium in order to determine the glycosphingolipids involved in B. hyodysenteriae adhesion. Specific interactions between B. hyodysenteriae and two non-acid glycosphingolipids were obtained. These binding-active glycosphingolipids, were characterized by mass spectrometry as lactotetraosylceramide (Galß3GlcNAcß3Galß4Glcß1Cer) and the B5 glycosphingolipid (Galα3Galß4GlcNAcß3Galß4Glcß1Cer). Comparative binding studies using structurally related reference glycosphingolipids showed that B. hyodysenteriae binding to lactotetraosylceramide required an unsubstituted terminal Galß3GlcNAc sequence, while for binding to the B5 pentaosylceramide the terminal Galα3Galß4GlcNAc sequence is the minimum element recognized by the bacteria. Binding of Griffonia simplicifolia IB4 lectin to pig colon tissue sections from healthy control pig and B. hyodysenteriae infected pigs showed that in the healthy pigs the Galα3Gal epitope was mainly present in the lamina propria. In contrast, in four out of five pigs with swine dysentery there was an increased expression of Galα3Gal in the goblet cells and in the colonic crypts, where B. hyodysenteriae also was present. The one pig that had recovered by the time of necropsy had the Galα3Gal epitope only in the lamina propria. These data are consistent with a model where a transient increase in the carbohydrate sequence recognized by the bacteria occur in colonic mucins during B. hyodysenteriae infection, suggesting that the mucins may act as decoys contributing to clearance of the infection. These findings may lead to novel strategies for treatment of B. hyodysenteriae induced swine dysentery.

Factors driving pig owners' motivation and satisfaction to perform eradications from Swine dysentery.

Brachyspira hyodysenteriae is one of the agents of swine dysentery (SD) and its eradication is an effective, but costly control measure. Being a voluntary measure, knowledge about drivers of motivation and satisfaction regarding the eradication of SD would help to convince farmers to eradicate. We aimed to describe eradications performed in Switzerland and to analyse factors influencing the pig owners' perception (motivation and satisfaction) of SD eradications to provide a basis to formulate recommendations and guidelines. Pig farmers (n = 68) having conducted an SD eradication and being interested in the study were interviewed using a standardised digital questionnaire. We assessed their motivation as moderately or highly motivated. Based on the farmers' evaluation of nine aspects of the eradication, satisfaction was considered to be moderate (<7/9 aspects positively evaluated) or high (≥7/9). Farms with fattening pigs and farms with breeding stock were analysed separately in subsets. First, multivariable factor analysis for mixed data (FAMD) were performed to describe the main patterns of variation. Then, risk factors for motivation and satisfaction were quantified by means of logistic regression models. Mainly total depopulations (73.5%) had been performed. Of the 36 farmers with breeding pigs, 24 were highly motivated, and 20 highly satisfied. Of the 61 farmers with fattening pigs, 45 were highly motivated and 42 highly satisfied. The FAMD revealed that the two main components explained only 17.0% and 11.0% (breeding stock) and 13.0% and 11.0% (fattening pigs) of the total variation, respectively. For farmers with breeding stock no significant factors for motivation were detected, but they were more satisfied (OR 25.0) when they had a batch farrowing of 3 weeks. Farmers with fattening pigs were more likely to be more motivated when providing access to outdoor areas (OR 3.3) and when it was their own initiative (OR 5.5). Farmers were more likely to be satisfied when they had only fattening pigs (OR 5.7), when the eradication was their own initiative (OR 5.5) and when they did not disinfect the barns during the eradication (OR 15.6). Farmers deciding themselves to eradicate are presumably more likely convinced of the benefits of the eradication. Satisfaction associated with a 3-weeks batch farrowing might be related to an easier to organise eradication and no disinfection to reduced labour and costs. In summary, the majority of the farmers were satisfied with the eradication. Education could promote self-motivation of farmers, and subsidies might support the implementation of SD eradications.

Carriage of Brachyspira hyodysenteriae on common insect vectors.

The interactions of likely insect and murine vectors of the causative agent of swine dysentery, Brachyspira hyodysenteriae, were investigated. Insects were collected and analysed from 3 pig farms positive for B hyodysenteriae. Within these farms, several Musca domestica and Orphyra adult fly, Blatta sp. cockroach digestive tracts and hover fly (Eristalis sp) pupal form contents were positive in a standard PCR assay for B hyodysenteriae, whereas all other insect samples on these and case control farms were negative. In challenge exposure studies, B hyodysenteriae DNA was detected in the digestive tract of cockroaches and M domestica flies from day 1 post-inoculation with cultured B hyodysenteriae, for up to 5 days or 10 days respectively, while control non-inoculated insects remained negative. Isolates consistent with B hyodysenteriae were only cultured from frass samples of these inoculated cockroach and flies on days 1-3 post-inoculation. Isolates consistent with B hyodysenteriae were detected by analysis of agar plates exposed to live B hyodysenteriae-inoculated adult flies wandering and feeding on these plates for 20 min per day. In generational challenge inoculation studies, B hyodysenteriae was detected in the adult emergent flies, and internal components of fly pupae on days 1-7 of the pupation period, after being inoculated with B hyodysenteriae as larvae. Five-week-old conventional mice (C3H) that consumed 2 meals of B hyodysenteriae-infected flies remained negative for B hyodysenteriae throughout the next 10 days. The results indicated that pathogenic Brachyspira sp have a limited ability to internally colonise likely insect vectors and do not readily transmit infection to mice. However, the insect vectors analysed were demonstrably capable of mechanical carriage and likely on-farm involvement in consequence.

Influence of infection with Brachyspira hyodysenteriae on clinical expression, growth performance, and digestibility in growing pigs fed diets varying in type and level of fiber.

Research on the effects of different fiber types and levels on infection with Brachyspira hyodysenteriae on growth performance and nutrients digestibility in pigs is scarce. The objective of the current study was to investigate the effects of infection with B. hyodysenteriae when feeding diets varying in soluble and insoluble dietary fiber (DF) on the expression of swine dysentery, growth performance, and digestibility of organic matter (OM) nutrients. A total of 96 growing pigs (26.9 ± 2.5 kg) were used for the experiment and divided into six blocks. The growing pigs were fed one of four diets for 12 wk: low fiber (LF), high fiber (HF), high soluble fiber (HS), and high insoluble fiber (HI). After 2 wk, half of the pigs were inoculated with B. hyodysenteriae. Half of the pigs in each group were euthanized at week 6 for the measurement of the apparent digestibility at the ileum, cecum, colon, and total tract. The remaining pigs were maintained to observe and analyze the clinical expression of fecal score and excretion of B. hyodysenteriae, growth performance, and total tract digestibility up to 12 wk. In the current study, the experimental diets did not influence the expression of infection in the pigs. The body weight and average daily gain (ADG) were in line with the results of clinical expression from week 4 to 6. However, the ADG of the infected pigs started to recover from week 6 (P < 0.05) and then recovered from week 8 to 12 (P < 0.05). The infection with B. hyodysenteriae did not impair apparent ileal digestibility (AID; P > 0.05), whereas the apparent digestibility of OM, total non-starch polysaccharide, non-cellulosic polysaccharide, and cellulose in the cecum of the infected pigs was higher than non-infected pigs (P < 0.05). The apparent colonic digestibility of ash and nitrogen was higher in non-infected pigs than in infected pigs (P < 0.05). The pigs fed the LF diet had a higher digestibility in all segments of the intestinal tract, whereas the HS diet had the lowest AID but higher or similar to the LF diet in the cecum, colon, and the total tract (P < 0.05). The pigs fed the HF and HI diets, with a high proportion of insoluble fiber, had a lower digestibility in the hindgut than the other two diets (P < 0.05). In conclusion, the infection with B. hyodysenteriae negatively influenced clinical signs of swine dysentery and growth performance but did not impair AID, and neither soluble nor insoluble DF influenced the expression of the infection.

Swine dysentery is a severe disease that can cause increased mortality and poor feed efficiency with bloody diarrhea. This disease can be treated with antibiotics, but there is a limitation of using antibiotics due to governmental policy, thereby the incidence of swine dysentery has been increased. We, therefore, try to find alternatives with diverse fiber sources and understand the mechanism of swine dysentery in growing pigs. In this study, infection of Brachyspira hyodysenteriae showed a negative influence on growth performance, but compensatory growth and recovery were observed in pigs after 6 wk of the infection. The apparent ileal digestibility was not affected by the infection, and the digestibility of non-starch polysaccharides in the cecum was rather increased than decreased probably because of interaction between B. hyodysenteriae and specific bacteria, which can stimulate fiber degradation in the cecum. However, fiber type and level did not influence the prevention and alleviation of the infection.

Diversity and potential genetic relationships amongst Brazilian Brachyspira hyodysenteriae isolates from cases of swine dysentery.

The aim of this study was to evaluate genetic diversity, distribution, evolution and population structure of Brazilian Brachyspira hyodysenteriae strains isolated from swine. Multilocus Sequence Typing (MLST) analysis using seven housekeeping genes was applied to 46 isolates obtained from outbreaks of swine dysentery that occurred between 2011 and 2015 in the states of Minas Gerais, São Paulo, Mato Grosso, Rio Grande do Sul and Santa Catarina. Historical isolates from Rio Grande do Sul obtained in 1998 were also included in the study. An independent international profile of the global sequences deposited in the B. hyodysenteriae database was used for comparisons with the Brazilian strains. All isolates from 2011 to 2015 were classified into nine sequence type (STs) and divided into four clonal complexes. These findings indicated genetic relationships among the B. hyodysenteriae from different Brazilian states, among historical strains isolated in 1998 and from recent outbreaks, and relatedness with global isolates. Seven STs were unique and, to date, only reported in Brazil.

Whole-genome analyses reveal a novel prophage and cgSNPs-derived sublineages of Brachyspira hyodysenteriae ST196.

BACKGROUND: Brachyspira (B.) hyodysenteriae is a fastidious anaerobe spirochete that can cause swine dysentery, a severe mucohaemorragic colitis that affects pig production and animal welfare worldwide. In Switzerland, the population of B. hyodysenteriae is characterized by the predominance of macrolide-lincosamide-resistant B. hyodysenteriae isolates of sequence type (ST) ST196, prompting us to obtain deeper insights into the genomic structure and variability of ST196 using pangenome and whole genome variant analyses. RESULTS: The draft genome of 14 B. hyodysenteriae isolates of ST196, sampled during a 7-year period from geographically distant pig herds, was obtained by whole-genome sequencing (WGS) and compared to the complete genome of the B. hyodysenteriae isolate Bh743-7 of ST196 used as reference. Variability results revealed the existence of 30 to 52 single nucleotide polymorphisms (SNPs), resulting in eight sublineages of ST196. The pangenome analysis led to the identification of a novel prophage, pphBhCH20, of the Siphoviridae family in a single isolate of ST196, which suggests that horizontal gene transfer events may drive changes in genomic structure. CONCLUSIONS: This study contributes to the catalogue of publicly available genomes and provides relevant bioinformatic tools and information for further comparative genomic analyses for B. hyodysenteriae. It reveals that Swiss B. hyodysenteriae isolates of the same ST may have evolved independently over time by point mutations and acquisition of larger genetic elements. In line with this, the third type of mobile genetic element described so far in B. hyodysenteriae, the novel prophage pphBhCH20, has been identified in a single isolate of B. hyodysenteriae of ST196.

Acute infection with Brachyspira hyodysenteriae affects mucin expression, glycosylation, and fecal MUC5AC.

Introduction: Infection with strongly ß-hemolytic strains of Brachyspira hyodysenteriae leads to swine dysentery (SD), a production-limiting disease that causes mucohemorrhagic diarrhea and typhlocolitis in pigs. This pathogen has strong chemotactic activity toward mucin, and infected pigs often have a disorganized mucus layer and marked de novo expression of MUC5AC, which is not constitutively expressed in the colon. It has been shown that fucose is chemoattractant for B. hyodysenteriae, and a highly fermentable fiber diet can mitigate and delay the onset of SD. Methods: We used lectins targeting sialic acids in α-2,6 or α-2,3 linkages, N-acetylglucosamine (GlcNAc), α-linked L-fucose, and an immunohistochemical stain targeting N-glycolylneuraminic acid (NeuGc) to investigate the local expression of these mucin glycans in colonic tissues of pigs with acute SD. We used a commercial enzyme-linked immunosorbent assay (ELISA) to quantify fecal MUC5AC in infected pigs and assess its potential as a diagnostic monitoring tool and RNA in situ hybridization to detect IL-17A in the colonic mucosa. Results: Colonic mucin glycosylation during SD has an overall increase in fucose, a spatially different distribution of GlcNAc with more expression within the crypt lumens of the upper colonic mucosa, and decreased expression or a decreased trend of sialic acids in α-2,6 or α-2,3 linkages, and NeuGc compared to the controls. The degree of increased fucosylation was less in the colonic mucosa of pigs with SD and fed the highly fermentable fiber diet. There was a significant increase in MUC5AC in fecal and colonic samples of pigs with SD at the endpoint compared to the controls, but the predictive value for disease progression was limited. Discussion: Fucosylation and the impact of dietary fiber may play important roles in the pathogenesis of SD. The lack of predictive value for fecal MUC5AC quantification by ELISA is possibly due to the presence of other non-colonic sources of MUC5AC in the feces. The moderate correlation between IL-17A, neutrophils and MUC5AC confirms its immunoregulatory and mucin stimulatory role. Our study characterizes local alteration of mucin glycosylation in the colonic mucosa of pigs with SD after B. hyodysenteriae infection and may provide insight into host-pathogen interaction.

Brachyspira Species Avidity to Colonic Mucins from Pigs with and without Brachyspira hyodysenteriae Infection Is Species Specific and Varies between Strains.

Brachyspira hyodysenteriae is commonly associated with swine dysentery (SD), a disease that has an economic impact on the swine industry. B. hyodysenteriae infection results in changes to the colonic mucus niche with massive mucus induction, which substantially increases the number of B. hyodysenteriae binding sites in the mucus. We previously determined that a B. hyodysenteriae strain binds to colon mucins in a manner that differs between pigs and mucin types. Here, we investigated if adhesion to mucins is a trait observed across a broad set of B. hyodysenteriae strains and isolates and furthermore at a genus level (B. innocens, B. pilosicoli, B. murdochii, B. hampsonii, and B. intermedia strains). Our results show that binding to mucins appears to be specific to B. hyodysenteriae, and within this species, the binding ability to mucins varies between strains/isolates, increases for mucins from pigs with SD, and is associated with sialic acid epitopes on mucins. Infection with B. hyodysenteriae strain 8dII results in mucin glycosylation changes in the colon, including a shift in sialic acid-containing structures. Thus, we demonstrate through hierarchical cluster analysis and orthogonal projections to latent structures discriminant analysis (OPLS-DA) models of the relative abundances of sialic acid-containing glycans that sialic acid-containing structures in the mucin O-glycome are good predictors of B. hyodysenteriae strain 8dII infection in pigs. The results emphasize the role of sialic acids in governing B. hyodysenteriae interactions with its host, which may open perspectives for therapeutic strategies.

Brachyspira hyodysenteriae isolate multilocus sequence type dashboard.

This focus article has been prepared by Susanna Williamson, Emma Stubberfield and Anna Brzozowska of the APHA and Jill Thomson of SRUC - Veterinary Consulting Services, Edinburgh.

Validation of an antimicrobial susceptibility testing protocol for Brachyspira hyodysenteriae and Brachyspira pilosicoli in an international ring trial.

Brachyspira hyodysenteriae and Brachyspira pilosicoli cause economically important enteric disease in pigs. Treatment of these infections often includes antimicrobial administration, which can be most effective when therapeutic options are informed by antimicrobial susceptibility testing data. Here we describe a method for broth dilution antimicrobial susceptibility testing of these bacteria, both of which are difficult to culture in vitro. The protocol was evaluated for its fitness for use in an inter-laboratory ring trial involving eight laboratories from seven countries, and employing eleven test strains (5 Brachyspira hyodysenteriae including the type strain B78T and 6 Brachyspira pilosicoli) and six antibiotics. Overall intra- and inter-laboratory reproducibility of this method was very good (>90 % MICs at mode +/- 1 log2). Whole genome sequencing revealed good correspondence between reduced susceptibility and the presence of previously defined antimicrobial resistance determinants. Interestingly, lnu(C) was identified in B. pilosicoli isolates with elevated MICs of lincomycin, whilst tva(B) was associated with elevated MICs of pleuromutilins in this species. We designated two new control strains with MICs lying within currently tested ranges, including for the pleuromutilins, in contrast to the control strain B. hyodysenteriae B78T. These were deposited at the DSMZ-German Collection of Microorganisms and Cell Cultures GmbH. The validation of a standard protocol and identification of new control strains facilitates comparisons between studies, establishment of robust interpretative criteria, and ultimately contributes to rational antimicrobial use when treating infected livestock.

Differential expression of hemolysin genes in weakly and strongly hemolytic Brachyspira hyodysenteriae strains.

BACKGROUND: Swine dysentery (SD) is a diarrheal disease in fattening pigs that is caused by the strongly hemolytic species Brachyspira (B.) hyodysenteriae, B. hampsonii and B. suanatina. As weakly hemolytic Brachyspira spp. are considered less virulent or even non-pathogenic, the hemolysin is regarded as an important factor in the pathogenesis of SD. Four hemolysin genes (tlyA, tlyB, tlyC, and hlyA) and four putative hemolysin genes (hemolysin, hemolysin activation protein, hemolysin III, and hemolysin channel protein) have been reported, but their role in strong hemolysis is not entirely clear. Our study aimed to assess the transcriptional activity of eight (putative) hemolysin genes in a strongly hemolytic (B204) and a weakly hemolytic (G423) B. hyodysenteriae strain during non-hemolytic and hemolytic growth stages. RESULTS: Strongly and weakly hemolytic B. hyodysenteriae strains caused hemolysis on blood agar at different growth stages, namely during log phase (B204) and stationary/death phase (G423). During the lag, early log, late log (stationary phase in G423) and death phase (time points 1-4) strains differed in their hemolysin gene transcription patterns. At time point 1, transcription of the putative hemolysin gene was higher in B204 than in G423. At time point 2, tlyA and tlyC were upregulated in B204 during hemolysis. TlyB and hlyA were upregulated in both strains at all time points, but higher transcription rates were observed in the weakly hemolytic strain G423. The transcription activity of the hemolysin channel protein gene was quite similar in both strains, whereas the hemolysin activation protein gene was upregulated in the non-hemolytic stage of B204 at time point 4. Sequence analysis revealed deletions, insertions and single nucleotide polymorphisms in the G423 hlyA promoter, although without altering the transcription activity of this gene. CONCLUSION: Our data indicate a combined activity of TlyA and TlyC as the most probable underlying mechanism of strong hemolysis in B. hyodysenteriae. Further studies should verify if the expression of tlyA is upregulated by the putative hemolysin gene. Depending on their immunogenic potential TlyA and TlyC may serve as possible vaccine candidates, especially since vaccines for an effective control of swine dysentery are currently not available.

Review: Occurrence and Distribution of Galanin in the Physiological and Inflammatory States in the Mammalian Gastrointestinal Tract.

Galanin (GAL) is a broad-spectrum peptide that was first identified 37 years ago. GAL, which acts through three specific receptor subtypes, is one of the most important molecules on an ever-growing list of neurotransmitters. Recent studies indicate that this peptide is commonly present in the gastrointestinal (GI) tract and GAL distribution can be seen in the enteric nervous system (ENS). The function of the GAL in the gastrointestinal tract is, inter alia, to regulate motility and secretion. It should be noted that the distribution of neuropeptides is largely dependent on the research model, as well as the part of the gastrointestinal tract under study. During the development of digestive disorders, fluctuations in GAL levels were observed. The occurrence of GAL largely depends on the stage of the disease, e.g., in porcine experimental colitis GAL secretion is caused by infection with Brachyspira hyodysenteriae. Many authors have suggested that increased GAL presence is related to the involvement of GAL in organ renewal. Additionally, it is tempting to speculate that GAL may be used in the treatment of gastroenteritis. This review aims to present the function of GAL in the mammalian gastrointestinal tract under physiological conditions. In addition, since GAL is undoubtedly involved in the regulation of inflammatory processes, and the aim of this publication is to provide up-to-date knowledge of the distribution of GAL in experimental models of gastrointestinal inflammation, which may help to accurately determine the role of this peptide in inflammatory diseases and its potential future use in the treatment of gastrointestinal disorders.

Risk factors for the infection with Brachyspira hyodysenteriae in pig herds.

Swine dysentery (SD), caused by infection with Brachyspira hyodysenteriae, is a serious disease in pig production worldwide. Quantitative risk factors triggering the occurrence of infection are unknown. The present case-control study aimed at identifying major risk factors related to presence of B. hyodysenteriae in pig herds. Twenty case herds and 60 randomly selected control herds with a minimum herd size of '10 sows/ 80 fattening pigs' were examined by means of a questionnaire-based interview and a herd examination. Herds with previous eradication of SD were excluded. Logistic regression models revealed that the 'positive/suspicious SD status of source herds', the regular application of treatment, purchasing more than 4 batches/ year, contact to foxes, diagnostics performed during last 12 months, liquid feeding systems, rats on farm, and >250 fatting places were associated with higher chances of a herd to be infected. On the contrary, having different sources of grower pigs within one batch, the presence of raptor birds and the presence of martens in the region were associated with fewer chances of being infected. The final multivariable logistic regression model identified purchasing more than 4 batches/ year (OR = 7.5, 95 % CI 1.8-54.3) and contact to foxes (OR = 5.9; 97.5 % CI 1.2-34.6) as the two main risk factors in our study. 'More than 4 batches/ year' implies continuous herd management supporting persistence of B. hyodysenteriae in an infected herd, but also increased number of purchases each increasing the risk of B. hyodysenteriae introduction by carrier pigs or transport vehicles. Foxes might be infected with B. hyodysenteriae by feeding on positive piglets and rodents. Besides, 'contact to foxes' might represent a lack in biosecurity. In conclusion, the risk factors detected underline the importance of biosecurity in SD prevention and control.

Treatment of clinical Brachyspira hyodysenteriae with zinc chelate in pigs: a blinded, randomised controlled trial.

BACKGROUND: Brachyspira hyodysenteriae infection in pigs ('swine dysentery') leads to decreased feed conversion, growth losses and mortality. Current countermeasures have the downside of antibiotic resistance (antibiotics) and ecotoxicity (zinc oxide). The aim of this study was to evaluate the effect of a novel zinc chelate (Intra Dysovinol (ID)) on clinical signs of swine dysentery and shedding of B hyodysenteriae under field conditions. METHODS: In a randomised, double-blinded, controlled trial under Good Clinical Practice on two commercial farms, 58 B hyodysenteriae positive pigs from 16 pens received drinking water containing ID, or placebo, during six consecutive days. Faecal quality (consistency, colour, additions) was scored and faeces were analysed for presence of B hyodysenteriae by PCR. ID treatment positively affected faecal quality (consistency) and daily growth rates. RESULTS: At the last treatment day, B hyodysenteriae was not detectable in the faeces of any of the ID-treated animals, while all placebo animals remained B hyodysenteriae positive by PCR. All ID-treated animals recovered, while 5 placebo-treated animals died and 12 placebo pigs required additional treatment before the end of the study (up to 14 days after treatment start). CONCLUSION: This non-antibiotic treatment stopped the clinical signs and shedding of B hyodysenteriae in naturally infected pigs.

Testing the efficacy of kitasamycin for use in the control and treatment of swine dysentery in experimentally infected pigs.

BACKGROUND: Swine dysentery (SD) caused by Brachyspira hyodysenteriae is an important disease in Australia. AIM: The aim of this study is to evaluate the macrolide antibiotic kitasamycin for use in SD control. METHODS: The minimum inhibitory concentrations (MICs) of kitasamycin, tylosin and lincomycin for 32 Australian isolates of B. hyodysenteriae were evaluated. Mutations in the 23S rRNA gene were examined. Isolate '13' with a low kitasamycin MIC was used to challenge weaner pigs. Sixty pigs were housed in 20 pens each containing three pigs: pigs in four pens received 2 kg/tonne of a product containing kitasamycin (3.1% active) prophylactically in their food starting 4 days before B. hyodysenteriae challenge (group 1); pigs in four pens were challenged and received the same dose therapeutically once one pig in a pen showed diarrhoea (group 2); four pens were challenged and received 4 kg/tonne of the product therapeutically (group 3); four pens were challenged but not medicated (group 4); two pens were unmedicated and unchallenged (group 5) and two pens received 2 kg/tonne and were unchallenged (group 6). Pigs were monitored for B. hyodysenteriae excretion and disease. RESULTS: Macrolide resistance was widespread, and mutations in the 23S rRNA gene were identified in 23 isolates. Four isolates with kitasamycin MICs < 5 µg/mL were considered susceptible. Following experimental challenge, 10 of 12 unmedicated pigs developed SD. No pigs receiving kitasamycin prophylactical or therapeutically developed SD. Medicated pigs shed low numbers of B. hyodysenteriae in their faeces. CONCLUSIONS: Kitasamycin can help control SD in pigs infected with susceptible isolates of B. hyodysenteriae.