Isolation and characterization of Faecalibacterium prausnitzii from calves and piglets.

The goal of our study was to isolate and characterize Faecalibacterium prausnitzii from fecal samples of healthy calves and piglets, in order to develop a novel probiotic for livestock animals. We identified 203 isolates of Faecalibacterium sp., which were clustered in 40 genetically distinct groups. One representative isolate from each cluster was selected for further characterization. The concentrations of the short chain fatty acids (SCFA) acetate, butyrate, propionate and isobutyrate in the culture media were measured by gas chromatography. We observed reduction in the concentration of acetate followed by concomitant increase in the concentration of butyrate, suggesting that the isolates were consuming acetate present in the media and producing butyrate. Butyrate production correlated positively with bacterial growth. Since butyrate has many benefits to the colonic epithelial cells, the selection of strains that produce higher amounts of butyrate is extremely important for the development of this potential probiotic. The effect of pH and concentration of bile salts on bacterial growth was also evaluated in order to mimic the conditions encountered by F. prausnitzii in vivo. The optimal pH for growth ranged between 5.5 and 6.7, while most isolates were inhibited by of the lowest concentration of bile salts tested (0.1%). Antimicrobial resistance profile showed that most isolates of Faecalibacterium sp. were resistant against ciprofloxacin and sulfamethoxazole-trimethoprim. More than 50% of the isolates were resistant to tetracycline, amikacin, cefepime and cefoxitin. A total of 19 different combinations of multidrug resistance were observed among the isolates. Our results provide new insights into the cultural and physiological characteristics of Faecalibacterium prausnitzii illustrating large variability in short chain fatty acid production, in vitro growth, sensitivity to bile salts, and antibiotic resistance and suggesting that future probiotic candidates should be carefully studied before elected for in vivo studies

Faecalibacterium prausnitzii : from microbiology to diagnostics and prognostics

We thank Dr Xavier Aldeguer and MD David Busquets from the Hospital Dr Josep Trueta (Girona, Spain) and M.D Míriam Sabat Mir from the Hospital Santa Caterina (Salt, Spain) for their help and critical discussion concerning clinical aspects. This work was partially funded by the Spanish Ministry of Education and Science through the projects SAF2010-15896 and SAF2013-43284-P, which has been co-financed with FEDER funds. Dr Sylvia H Duncan acknowledges support from the Scottish Government Food, Land and People program.Peer reviewedPostprin

Genotypic antimicrobial resistance characterization of E. coli from dairy calves at high risk of respiratory disease administered enrofloxacin or tulathromycin.

The objective of this study was to evaluate the longitudinal effect of enrofloxacin or tulathromycin use in calves at high risk of bovine respiratory disease (BRD) on antimicrobial resistance genes and mutation in quinolone resistance-determining regions (QRDR) in fecal E. coli. Calves at high risk of developing BRD were randomly enrolled in one of three groups receiving: (1) enrofloxacin (ENR; n = 22); (2) tulathromycin (TUL; n = 24); or (3) no treatment (CTL; n = 21). Fecal samples were collected at enrollment and at 7, 28, and 56 days after beginning treatment, cultured for Escherichia coli (EC) and DNA extracted. Isolates were screened for cephalosporin, quinolone and tetracycline resistance genes using PCR. QRDR screening was conducted using Sanger sequencing. The only resistance genes detected were aac(6')Ib-cr (n = 13), bla-CTX-M (n = 51), bla-TEM (n = 117), tetA (n = 142) and tetB (n = 101). A significantly higher detection of gyrA mutated at position 248 at time points 7 (OR = 11.5; P value = 0.03) and 28 (OR = 9.0; P value = 0.05) was observed in the ENR group when compared to calves in the control group. Our findings support a better understanding of the potential impacts from the use of enrofloxacin in calves on the selection and persistence of resistance

Isolation and characterization of Faecalibacterium prausnitzii from calves and piglets.

The goal of our study was to isolate and characterize Faecalibacterium prausnitzii from fecal samples of healthy calves and piglets, in order to develop a novel probiotic for livestock animals. We identified 203 isolates of Faecalibacterium sp., which were clustered in 40 genetically distinct groups. One representative isolate from each cluster was selected for further characterization. The concentrations of the short chain fatty acids (SCFA) acetate, butyrate, propionate and isobutyrate in the culture media were measured by gas chromatography. We observed reduction in the concentration of acetate followed by concomitant increase in the concentration of butyrate, suggesting that the isolates were consuming acetate present in the media and producing butyrate. Butyrate production correlated positively with bacterial growth. Since butyrate has many benefits to the colonic epithelial cells, the selection of strains that produce higher amounts of butyrate is extremely important for the development of this potential probiotic. The effect of pH and concentration of bile salts on bacterial growth was also evaluated in order to mimic the conditions encountered by F. prausnitzii in vivo. The optimal pH for growth ranged between 5.5 and 6.7, while most isolates were inhibited by of the lowest concentration of bile salts tested (0.1%). Antimicrobial resistance profile showed that most isolates of Faecalibacterium sp. were resistant against ciprofloxacin and sulfamethoxazole-trimethoprim. More than 50% of the isolates were resistant to tetracycline, amikacin, cefepime and cefoxitin. A total of 19 different combinations of multidrug resistance were observed among the isolates. Our results provide new insights into the cultural and physiological characteristics of Faecalibacterium prausnitzii illustrating large variability in short chain fatty acid production, in vitro growth, sensitivity to bile salts, and antibiotic resistance and suggesting that future probiotic candidates should be carefully studied before elected for in vivo studies

Understanding the effect of producers' attitudes, perceived norms, and perceived behavioral control on intentions to use antimicrobials prudently on New York dairy farms

Understanding farmers' behavior, motivations, and perceptions toward antimicrobial use can influence how veterinarians translate research into practice and guide effective ways of implementing protocols. A multidisciplinary team investigated behavioral tendencies of New York dairy farmers toward antimicrobial use by administering a survey modeled with the reasoned action approach. This approach is a framework from social psychology containing the constructs attitude, perceived norms, and perceived behavioral control, and is used in combination with structural equation modeling to determine what drives intentions. Multiple indicators and multiple causes (MIMIC) models were then used to determine the effects of beliefs on their underlying constructs. The objective of the study was to provide direct and indirect measures of the constructs using survey data to determine importance of and associations with intention to use antimicrobials prudently. The structural equation model indicated that perceived behavioral control explained intention. Thus, farmers who feel capable of prudent use expressed positive intentions. Attitude and perception of others also had influence to a lesser extent. MIMIC models showed that the most important attributes of instrumental attitude were increasing profitability, decreasing risk of residues, and increasing herd health. Contributing attributes of affective attitude were job satisfaction, decreasing resistance, and increasing milk production. For perceived norms, the attributes were opinions/approval of family and peers, veterinarians, and milk processors. Finally, for perceived behavioral control, attributes focused on saving money on labor and treatment, ability to fit into the daily routine, and effectiveness with veterinary guidance. In conclusion, the best approach for adoption of practices might be presentation of examples of successful strategies by other producers, particularly in peer groups. In addition, veterinarians should provide the tools and guidance needed to produce economic gain, reduction of risks associated with residues and resistance, and positive experiences when using the tactics. </p

Isolation and characterization of Faecalibacterium prausnitzii from calves and piglets.

The goal of our study was to isolate and characterize Faecalibacterium prausnitzii from fecal samples of healthy calves and piglets, in order to develop a novel probiotic for livestock animals. We identified 203 isolates of Faecalibacterium sp., which were clustered in 40 genetically distinct groups. One representative isolate from each cluster was selected for further characterization. The concentrations of the short chain fatty acids (SCFA) acetate, butyrate, propionate and isobutyrate in the culture media were measured by gas chromatography. We observed reduction in the concentration of acetate followed by concomitant increase in the concentration of butyrate, suggesting that the isolates were consuming acetate present in the media and producing butyrate. Butyrate production correlated positively with bacterial growth. Since butyrate has many benefits to the colonic epithelial cells, the selection of strains that produce higher amounts of butyrate is extremely important for the development of this potential probiotic. The effect of pH and concentration of bile salts on bacterial growth was also evaluated in order to mimic the conditions encountered by F. prausnitzii in vivo. The optimal pH for growth ranged between 5.5 and 6.7, while most isolates were inhibited by of the lowest concentration of bile salts tested (0.1%). Antimicrobial resistance profile showed that most isolates of Faecalibacterium sp. were resistant against ciprofloxacin and sulfamethoxazole-trimethoprim. More than 50% of the isolates were resistant to tetracycline, amikacin, cefepime and cefoxitin. A total of 19 different combinations of multidrug resistance were observed among the isolates. Our results provide new insights into the cultural and physiological characteristics of Faecalibacterium prausnitzii illustrating large variability in short chain fatty acid production, in vitro growth, sensitivity to bile salts, and antibiotic resistance and suggesting that future probiotic candidates should be carefully studied before elected for in vivo studies

Effect of pH on bacterial growth.

<p>The increment of optical density after 48 h of incubation for each isolate in media containing different pH. Each box-and-whisker plot constitutes one pH value. The extent of the box encompasses the interquartile range of the optical density increment, whiskers extend to maximum and minimum values, and the line within each box represents the median. Outliers are represented as open black circles.</p

Correlations between the average difference of short chain fatty acids (acetate, butyrate, propionate and isobutyrate) concentrations and the growth performance of the <i>F. prausnitzii</i> isolates, represented by the increment of optical density.

<p>Correlation coefficients and fitted lines with confidence intervals are shown in each box.</p

Average difference of propionate and isobutyrate concentrations in the growth media after 48 h of incubation of each <i>F. prausnitzii</i> isolate.

<p>The bars represent the mean concentration in ppm. Error bars indicate the standard error of the mean.</p

Resistance profile of the <i>Faecalibacterium prausnitzii</i> isolated from feces of dairy calves and piglets, as determined by E-test (BioMérieux, Inc., Durham, NC).

<p>Distribution of the minimal inhibitory concentration (MIC) for the 29 isolates among the different antimicrobial concentrations (0.002–256 mg/µl). Shaded cells highlight the MIC range. Triple vertical bars mark the (approximate) breakpoint between sensitive and resistant. The resistance breakpoints are based on CLSI interpretative criteria for <i>Bacteroides fragilis</i> (ATCC 25285) (CLSI, 2012). No published MIC breakpoint for amikacin and gentamicin was found in the literature, therefore the highest concentration was considered as MIC.</p><p>MIC₅₀, the MIC that inhibited at least 50% of the isolates; MIC₉₀, the MIC that inhibited at least 90% of the isolates.</p><p>Resistance profile of the <i>Faecalibacterium prausnitzii</i> isolated from feces of dairy calves and piglets, as determined by E-test (BioMérieux, Inc., Durham, NC).</p