Assessment of the Impact of Humic Acids on Intestinal Microbiota, Gut Integrity, Ileum Morphometry, and Cellular Immunity of Turkey Poults Fed an Aflatoxin B1-Contaminated Diet.

A recent study published data on the growth performance, relative weights of the organs of the gastrointestinal tract, liver histology, serum biochemistry, and hematological parameters for turkey poults fed an experimental diet contaminated with aflatoxin B1 (AFB1) and humic acids (HA) extracted from vermicompost. The negative effects of AFB1 (250 ng AFB1/g of feed) were significantly reduced by HA supplementation (0.25% w/w), suggesting that HA might be utilized to ameliorate the negative impact of AFB1 from contaminated diets. The present study shows the results of the remaining variables, as an extension of a previously published work which aimed to evaluate the impact of HA on the intestinal microbiota, gut integrity, ileum morphometry, and cellular immunity of turkey poults fed an AFB1-contaminated diet. For this objective, five equal groups of 1-day-old female Nicholas-700 turkey poults were randomly assigned to the following treatments: negative control (basal diet), positive control (basal diet + 250 ng AFB1/g), HA (basal diet + 0.25% HA), HA + AFB1 (basal diet + 0.25% HA + 250 ng AFB1/g), and Zeolite (basal diet + 0.25% zeolite + 250 ng AFB1/g). In the experiment, seven replicates of ten poults each were used per treatment (n = 70). In general, HA supplementation with or without the presence of AFB1 showed a significant increase (p < 0.05) in the number of beneficial butyric acid producers, ileum villi height, and ileum total area, and a significant reduction in serum levels of fluorescein isothiocyanate-dextran (FITC-d), a marker of intestinal integrity. In contrast, poults fed with AFB1 showed a significant increase in Proteobacteria and lower numbers of beneficial bacteria, clearly suggesting gut dysbacteriosis. Moreover, poults supplemented with AFB1 displayed the lowest morphometric parameters and the highest intestinal permeability. Furthermore, poults in the negative and positive control treatments had the lowest cutaneous basophil hypersensitivity response. These findings suggest that HA supplementation enhanced intestinal integrity (shape and permeability), cellular immune response, and healthier gut microbiota composition, even in the presence of dietary exposure to AFB1. These results complement those of the previously published study, suggesting that HA may be a viable dietary intervention to improve gut health and immunity in turkey poults during aflatoxicosis.

Corrigendum: Evaluation of the efficacy of humic acids to counteract the toxic effects of aflatoxin B1 in turkey poults.

[This corrects the article DOI: 10.3389/fvets.2023.1276754.].

Evaluation of the Antimicrobial Activity of a Formulation Containing Ascorbic Acid and Eudragit FS 30D Microparticles for the Controlled Release of a Curcumin-Boric Acid Solid Dispersion in Turkey Poults Infected with Salmonella enteritidis: A Therapeutic Model.

The selection of components within a formulation or for treatment must stop being arbitrary and must be focused on scientific evidence that supports the inclusion of each one. Therefore, the objective of the present study was to obtain a formulation based on ascorbic acid (AA) and Eudragit FS 30D microparticles containing curcumin-boric acid (CUR-BA) considering interaction studies between the active components carried out via Fourier transform infrared spectrometry (FTIR) and differential scanning calorimetry (DSC) to minimize antagonistic effects, and comprehensively and effectively treat turkey poults infected with Salmonella enteritidis (S. enteritidis). The DSC and FTIR studies clearly demonstrated the interactions between AA, BA, and CUR. Consequently, the combination of AA with CUR and/or BA should be avoided, but not CUR and BA. Furthermore, the Eudragit FS 30D microparticles containing CUR-BA (SD CUR-BA MP) showed a limited release of CUR-BA in an acidic medium, but they were released at a pH 6.8-7.0, which reduced the interactions between CUR-BA and AA. Finally, in the S. enteritidis infection model, turkey poults treated with the combination of AA and SD CUR-BA MP presented lower counts of S. enteritidis in cecal tonsils after 10 days of treatment. These results pointed out that the use of an adequate combination of AA and CUR-BA as an integral treatment of S. enteritidis infections could be a viable option to replace the indiscriminate use of antibiotics.

Removal of Aflatoxin B1 Using Alfalfa Leaves as an Adsorbent Material: A Comparison between Two In Vitro Experimental Models.

An adsorbent material derived from alfalfa leaves was prepared and further characterized, and its efficacy for removing aflatoxin B1 (AFB1) was investigated. Characterization consisted of the use of attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR), environmental scanning electron microscopy (ESEM), X-ray fluorescence spectroscopy (XRF), X-ray diffraction (XRD), point of zero charge (pHpzc), zeta potential (ζ-potential), UV-Vis diffuse reflectance spectroscopy, and spectral analysis. To determine the adsorption capacity against AFB1 (250 ng AFB1/mL), pH-dependent and avian intestinal in vitro models were used. The adsorbent inclusion percentage was 0.5% (w/w). In general, the pH-dependent model gave adsorption percentages of 98.2%, 99.9%, and 98.2%, evaluated at pH values of 2, 5, and 7, respectively. However, when the avian intestinal model was used, it was observed that the adsorption percentage of AFB1 significantly decreased (88.8%). Based on the characterization results, it is proposed that electrostatic, non-electrostatic, and the formation of chlorophyll-AFB1 complexes were the main mechanisms for AFB1 adsorption. From these results, it can be concluded that the adsorbent derived from alfalfa leaves could be used as an effective material for removing AFB1 in in vitro digestion models that mimic the physiological reality.

Evaluation of the efficacy of humic acids to counteract the toxic effects of aflatoxin B1 in turkey poults.

This study aims to evaluate the efficacy of humic acid (HA) from worm compost as an adsorbent for aflatoxin B1 (AFB1) in turkey poults. The experiment involved the inclusion of 0.25% (w/w) HA in the diet of turkey poults consuming aflatoxin-contaminated feed (250 ng AFB1/g). A total of 350 1-day-old female Nicholas-700 turkey poults were randomly allocated to five equal groups: negative control (basal diet); positive control (basal diet + 250 ng AFB1/g; HA (basal diet + 0.25% HA); HA + AFB1 (basal diet + HA + 250 ng AFB1/g); and zeolite + AFB1 (basal diet + 0.25% zeolite + 250 ng AFB1/g). Each group had seven replicates of 10 poults (n = 70). The impact of HA addition was evaluated in terms of performance parameters, relative organ weights, liver histological lesions, and serum biochemical and hematological constituents. In general, the addition of HA improved body weight (BW), body weight gain (BWG), and feed conversion rate (FCR). Furthermore, HA effectively mitigated the toxic effects caused by AFB1 in the majority of the analyzed variables. The results indicated that HA effectively counteracted the AFB1-induced toxic effects in turkey poults. Based on these findings, it can be concluded that HA is capable of removing AFB1 from the contaminated diet.

Discriminative Dissolution Method Using the Open-Loop Configuration of the USP IV Apparatus to Compare Dissolution Profiles of Metoprolol Tartrate Immediate-Release Tablets: Use of Kinetic Parameters.

The use of the USP IV apparatus (flow-through cell) has gained acceptance in recent years due to its versatility and ability to discriminate due to its hydrodynamic conditions. Therefore, the objective of the present study was to develop a discriminative dissolution method in the USP IV apparatus using the open-loop configuration, as well as to propose a method to compare non-cumulative dissolution profiles obtained in the open-loop configuration considering kinetic parameters and validate its predictive power through its comparison with independent and dependent methods using five commercial immediate-release tablet drugs (one reference drug and four generic drugs) of metoprolol tartrate as a model drug. The comparison of the non-accumulated dissolution profiles consisted of determining the geometric ratio of Cmax, AUC0∞, AUC0Cmax, and Tmax (kinetic parameters) of the generic/reference drugs, whereby generic drugs "C" and "D" presented the highest probability of similarity since their 90% confidence intervals were included, or they were very close to the acceptance interval (80.00-125.00%). These results were consistent with the f2, bootstrap f2, and dissolution efficiency approaches (independent models). In conclusion, the proposed comparison method can be an important tool to establish similarity in dissolution profiles and to facilitate the development/selection of new formulations and positively ensure bioequivalence in clinical studies.

Humic Acids Preparation, Characterization, and Their Potential Adsorption Capacity for Aflatoxin B1 in an In Vitro Poultry Digestive Model.

Vermicompost was used for humic acid (HA) preparation, and the adsorption of aflatoxin B1 (AFB1) was investigated. Two forms of HA were evaluated, natural HA and sodium-free HA (SFHA). As a reference, a non-commercial zeolitic material was employed. The adsorbents were characterized by attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR), energy-dispersive X-ray spectroscopy (EDS), zeta potential (ζ-potential), scanning electron microscopy (SEM), and point of zero charge (pHpzc). The adsorbent capacity of the materials when added to an AFB1-contaminated diet (100 µg AFB1/kg) was evaluated using an in vitro model that simulates the digestive tract of chickens. Characterization results revealed the primary functional groups in HA and SFHA were carboxyl and phenol. Furthermore, adsorbents have a highly negative ζ-potential at the three simulated pH values. Therefore, it appears the main influencing factors for AFB1 adsorption are electrostatic interactions and hydrogen bonding. Moreover, the bioavailability of AFB1 in the intestinal section was dramatically decreased when sorbents were added to the diet (0.2%, w/w). The highest AFB1 adsorption percentages using HA and SFHA were 97.6% and 99.7%, respectively. The zeolitic material had a considerable adsorption (81.5%). From these results, it can be concluded that HA and SFHA from vermicompost could be used as potential adsorbents to remove AFB1 from contaminated feeds.

Whole-Genome Sequence and Interaction Analysis in the Production of Six Enzymes From the Three Bacillus Strains Present in a Commercial Direct-Fed Microbial (Norum™) Using a Bliss Independence Test.

The three Bacillus strains present in Norum™ were initially selected by their excellent to good relative enzyme activity (REA) production score for amylase, protease, lipase, phytase, cellulase, ß-glucanase, and xylanase. Further studies confirmed that the three isolates also showed an antibacterial activity, Gram-positive and Gram-negative poultry pathogens. Norum™ (Eco-Bio/Euxxis Bioscience LLC) is a Bacillus spore direct-fed microbial (DFM). The Bacillus isolates were screened and selected based on in vitro enzyme production profiles. Moreover, in chickens fed high non-starch polysaccharides, this DFM demonstrated to reduce digesta viscosity, bacterial translocation, increase performance, bone mineralization, and balance the intestinal microbiota. In the present study, we present the whole-genome sequence of each of the three isolates in Norum™, as well as the synergistic, additive, or antagonistic effects on the enzyme production behavior of the three Bacillus strains and their combinations when grown together vs. when grown individually. The whole-genome sequence identified isolate AM1002 as Bacillus subtilis (isolate 1), isolate AM0938 as Bacillus amyloliquefaciens (isolate 2), and isolate JD17 as Bacillus licheniformis (isolate 3). The three Bacillus isolates used in the present study produce different enzymes (xylanase, cellulase, phytase, lipase, protease, and ß-glucanase). However, this production was modified when two or more Bacillus strains were combined, suggesting possible synergistic, antagonistic, or additive interactions. The Bliss analysis suggested (p < 0.05) that the combination of Bacillus strains 1-2 and 1-2-3 had intermediate effects and predicted that the combination of Bacillus strains 2-3 could have better effects than the combination of all the three Bacillus strains. In summary, the current study demonstrated the need of selecting Bacillus strains based on quantitative enzyme determination and data analysis to assess the impacts of combinations to avoid antagonistic interactions that could limit treatment efficacy. These results suggest that using Bacillus strains 2-3 together could lead to a new generation of DFMs with effects superior to those already examined in Bacillus strains 1-2-3 and, therefore, a potential alternative to growth-promoting antibiotics. More research utilizing poultry models is being considered to confirm and expand the existing findings.

Effects of humic acids on the recovery of different bacterial strains in an in vitro chicken digestive model.

Humic acids (HA) have been evaluated as growth promoters in poultry, but their effects on the gut microbiota remains controversial using in vitro and in vivo models. The objective of this study was to determine the effect of HA extracted from a wormcompost on the recovery of bacteria: Salmonella Enteritidis (S. Enteritidis), Escherichia coli (E. coli), Clostridium perfringens (C. perfringens), Bacillus subtilis (B. subtilis) and Lactobacillus salivarius (L. salivarius) using an in vitro chicken digestive system. Independent in vitro trials were run for each bacteria using six treatments: 1) Negative control with no bacteria added (Control-), 2) Positive control added with bacteria (Control+), 3) 0.1% HA + bacteria, 4) 0.2% HA + bacteria, 5) 0.5% HA + bacteria and 6) 1% HA + bacteria. Data was subjected to analysis of variance and linear regression. In the crop, S. Enteritidis was lower, C. perfringes and B. subtilis were not affected by HA, while E. coli and L. salivarius were higher at 0.5 and 1% HA inclusion (P ≤ 0.0001). In the proventriculus, S. Enteritidis, E. coli and B. subtilis were higher at 0.5 and 1% HA inclusion (P ≤ 0.0001); C. perfringens and L. salivarius were not affected by HA. In intestine, significant increases of all bacteria strains were observed (P ≤ 0.0001). In conclusion, the results suggests that HA can be used as prebiotic, but their mechanisms of action to stimulate the growth of gut bacteria remains to be elucidated.

Potential of Kale and Lettuce Residues as Natural Adsorbents of the Carcinogen Aflatoxin B1 in a Dynamic Gastrointestinal Tract-Simulated Model.

Adsorption of the carcinogen aflatoxin B1 (AFB1) onto agro-waste-based materials is a promising alternative over conventional inorganic binders. In the current study, two unmodified adsorbents were eco-friendly prepared from kale and lettuce agro-wastes. A dynamic gastrointestinal tract-simulated model was utilized to evaluate the removal efficiency of the sorptive materials (0.5%, w/w) when added to an AFB1-contaminated diet (100 µg AFB1/kg). Different characterization methodologies were employed to understand the interaction mechanisms between the AFB1 molecule and the biosorbents. Based on adsorption results, the biosorbent prepared from kale was the best; its maximum adsorption capacity was 93.6%, which was significantly higher than that of the lettuce biosorbent (83.7%). Characterization results indicate that different mechanisms may act simultaneously during adsorption. Non-electrostatic (hydrophobic interactions, dipole-dipole interactions, and hydrogen bonding) and electrostatic interactions (ionic attractions) together with the formation of AFB1-chlorophyll complexes appear to be the major influencing factors driving AFB1 biosorption.

Curcumin reduces enteric isoprostane 8-iso-PGF2α and prostaglandin GF2α in specific pathogen-free Leghorn chickens challenged with Eimeria maxima.

The purpose of this pilot study was to evaluate and determine the concentration of prostaglandin GF2α (PGF2α) and isoprostane 8-iso-PGF2α in plasma and intestine of specific pathogen-free (SPF) Leghorn chickens challenged with Eimeria maxima, with or without dietary supplementation of curcumin using solid-phase microextraction and ultra-performance liquid chromatography/tandem mass spectrometry. Eighty 1-day-old male SPF chickens were randomly allocated to one of four groups with four replicates (n = 5 chickens/replicate). Groups consisted of: (1) Control (no challenge), (2) Curcumin (no challenge), (3) Eimeria maxima (challenge), and (4) Eimeria maxima (challenge) + curcumin. At day 28 of age, all chickens in the challenge groups were orally gavaged with 40,000 sporulated E. maxima oocysts. No significant differences (P > 0.05) were observed in the groups regardless of the treatment or challenge with E. maxima. Enteric levels of both isoprostane 8-iso-PGF2α and PGF2α at 7 days and 9 days post-challenge were significantly increased (P < 0.01) compared to the non-challenge control chickens. Interestingly, the enteric levels of both isoprostane 8-iso-PGF2α and PGF2α at 7 days post-challenge were significantly reduced in chickens fed curcumin, compared to control chickens challenge with E. maxima. At 9 days post-challenge, only levels of isoprostane 8-iso-PGF2α in the enteric samples were significantly reduced in chickens challenged with E. maxima supplemented with curcumin, compared with E. maxima challenge chickens. No differences of isoprostane 8-iso-PGF2α or PGF2α were observed in plasma at both days of evaluation. Similarly, no significant differences were observed between the challenge control or chickens challenge with E. maxima and supplemented with curcumin at both times of evaluation. The results of this pilot study suggests that the antioxidant anti-inflammatory properties of curcumin reduced the oxidative damage and subsequent intestinal mucosal over-production of lipid oxidation products. Further studies to confirm and extend these results in broiler chickens are required.

Evaluation of curcumin and copper acetate against Salmonella Typhimurium infection, intestinal permeability, and cecal microbiota composition in broiler chickens.

BACKGROUND: Interest in the use of natural feed additives as an alternative to antimicrobials in the poultry industry has increased in recent years because of the risk of bacterial resistance. One of the most studied groups are polyphenolic compounds, given their advantages over other types of additives and their easy potentiation of effects when complexes are formed with metal ions. Therefore, the objective of the present study was to evaluate the impact of dietary supplementation of copper acetate (CA), curcumin (CR), and their combination (CA-CR) against Salmonella Typhimurium colonization, intestinal permeability, and cecal microbiota composition in broiler chickens through a laboratory Salmonella infection model. S. Typhimurium recovery was determined on day 10 post-challenge by isolating Salmonella in homogenates of the right cecal tonsil (12 chickens per group) on Xylose Lysine Tergitol-4 (XLT-4) with novobiocin and nalidixic acid. Intestinal integrity was indirectly determined by the fluorometric measurement of fluorescein isothiocyanate dextran (FITC-d) in serum samples from blood obtained on d 10 post-S. Typhimurium challenge. Finally, microbiota analysis was performed using the content of the left caecal tonsil of 5 chickens per group by sequencing V4 region of 16S rRNA gene. RESULTS: The results showed that in two independent studies, all experimental treatments were able to significantly reduce the S. Typhimurium colonization in cecal tonsils (CT, P < 0.0001) compared to the positive control (PC) group. However, only CA-CR was the most effective treatment in reducing S. Typhimurium counts in both independent studies. Furthermore, the serum fluorescein isothiocyanate dextran (FITC-d) concentration in chickens treated with CR was significantly lower when compared to PC (P = 0.0084), which is related to a decrease in intestinal permeability and therefore intestinal integrity. The effect of dietary treatments in reducing Salmonella was further supported by the analysis of 16S rRNA gene sequences using Linear discriminant analysis effect size (LEfSe) since Salmonella was significantly enriched in PC group (LDA score > 2.0 and P < 0.05) compared to other groups. In addition, Coprobacillus, Eubacterium, and Clostridium were significantly higher in the PC group compared to other treatment groups. On the contrary, Fecalibacterium and Enterococcus in CR, unknown genus of Erysipelotrichaceae at CA-CR, and unknown genus of Lachnospiraceae at CA were significantly more abundant respectively. CONCLUSIONS: CR treatment was the most effective treatment to reduce S. Typhimurium intestinal colonization and maintain better intestinal homeostasis which might be achieved through modulation of cecal microbiota.

Assessment of Fermented Soybean Meal on Salmonella typhimurium Infection in Neonatal Turkey Poults.

This study's objective was to evaluate the effect of the fermented soybean meal (FSBM) on Salmonella typhimurium (ST) to turkey poults using two models of infection. In the prophylactic model, one-day-old turkeys were randomly allocated to one of four different groups (n = 30 turkeys/group): (1) Control group, (2) FSBM group, (3) Control group challenged with ST (Control + ST), and (4) FSBM group challenged with ST (FSBM + ST). On day 9 of age, all poults were orally challenged with 106 colony forming units (CFU) ST and 24 h post-inoculation, intestinal samples were collected to determine ST recovery and morphometric analysis. Blood samples were collected to evaluate serum fluorescein isothiocyanate-dextran (FITC-d). In the therapeutic model, a similar experimental design was used, but turkeys were orally gavaged 104 CFU ST on day 1, and samples were collected at day 7. FSBM improved performance and reduced leaky gut in both experimental infective models. In the prophylactic model, FSBB induced morphology changes in the mucosa. Although the strains (Lactobacillus salivarius and Bacillus licheniformis) used for the fermentation process showed in vitro activity against ST, no significant effect was observed in vivo. The fermentation with different beneficial bacteria and different inclusion rates of FSBM requires further investigation.

In ovo Administration of Defined Lactic Acid Bacteria Previously Isolated From Adult Hens Induced Variations in the Cecae Microbiota Structure and Enterobacteriaceae Colonization on a Virulent Escherichia coli Horizontal Infection Model in Broiler Chickens.

The effects of in ovo administration of a defined lactic acid microbiota (LAM), previously isolated from adult hens, in the cecae microbiota structure and Enterobacteriaceae colonization after exposure to virulent Escherichia coli during the hatching phase of broiler chickens were evaluated. Embryos inoculated with LAM showed a significant (P < 0.05) reduction of Enterobacteriaceae colonization at day-of-hatch (DOH) and day (d) 7. Furthermore, there was a significant increase in total lactic acid bacteria on DOH, body weight (BW) DOH, BW d7, and d0-d7 BW gain and reduced mortality d0-d7 was observed in the LAM group compared with that in phosphate-buffered saline (PBS) control. The bacterial composition at the family level revealed that the Enterobacteriaceae was numerically reduced, whereas the Ruminococcaceae was significantly increased in the LAM group when compared with that in the PBS control. Moreover, the bacterial genera Proteus and Butyricicoccus and unidentified bacterial genera of family Lachnospiraceae and Erysipelotrichaceae were significantly enriched in the LAM group. In contrast, the Clostridium of the family Peptostreptococcaceae and unidentified genus of family Enterobacteriaceae were significantly abundant in the PBS control group. In summary, in ovo administration of a defined LAM isolated from adult hens did not affect hatchability, improved body weight gain and reduced mortality at d7, induced variations in the cecae microbiota structure and reduced Enterobacteriaceae colonization on a virulent E. coli horizontal infection model in broiler chickens.

Evaluation of a Bacillus -Based Direct-Fed Microbial on Aflatoxin B1 Toxic Effects, Performance, Immunologic Status, and Serum Biochemical Parameters in Broiler Chickens.

The aim of the present study was to evaluate the effect of a commercial Bacillus direct-fed microbial (DFM) on aflatoxin B1 toxic effects, performance, and biochemical and immunologic parameters in broiler chickens. Ninety 1-day-old Cobb 500 male broiler chicks were raised in floor pens for a period of 21 days. Chicks were neck-tagged, individually weighed, and randomly allocated to one of three groups: Negative control (basal feed), aflatoxin B1 (basal feed + 2 ppm AFB1), and DFM (basal feed + 2 ppm AFB1 + Bacillus direct-fed microbial). Each group had three replicates of 10 chickens (n = 30/group). Body weight and body weight gain were calculated weekly, while feed intake and feed conversion ratio were determined when broilers were 21 days old. On day 21, all chickens were bled, gastrointestinal samples were collected, and spleen and bursa of Fabricius were weighed. This study confirmed that 2 ppm of AFB1 causes severe detrimental effects on performance, biochemical parameters, and immunologic parameters, generating hepatic lesions in broiler chickens (P < 0.05). However, it was also observed that DFM supplementation provided beneficial effects that might help to improve gut barrier function, anti-inflammatory and antioxidant activities, as well as humoral and cellular immunomodulation. The results of the present study suggest that this Bacillus-DFM added at a concentration of 106 spores/gram of feed can be used to counteract the negative effects that occur when birds consume diets contaminated with AFB1, showing beneficial effects on performance parameters, relative organ weights, hepatic lesions, immune response, and serum biochemical variables. The addition of this Bacillus-DFM might mitigate and decrease aflatoxicosis problems in the poultry industry, improving food security, alleviating public health problems, and providing economic benefits. Future studies are needed to fully elucidate the specific mechanisms by which this Bacillus-DFM counteracts the toxic effects of aflatoxin B1.

Evaluación de un producto comercial adicionado en el alimento elaborado con Bacillus sobre los efectos tóxicos de la aflatoxina B1, el rendimiento productivo, el estado inmunológico y los parámetros bioquímicos en suero de pollos de engorde. El objetivo del presente estudio fue evaluar el efecto de un producto comercial de Bacillus adicionado al alimento (DFM) sobre los efectos tóxicos de la aflatoxina B1, el rendimiento productivo, así como en los parámetros bioquímicos e inmunológicos en pollos de engorde. Noventa pollitos de engorde machos Cobb 500 de un día de edad fueron criados en corrales en piso por un período de 21 días. Los pollos se etiquetaron en el cuello, se pesaron individualmente y se asignaron al azar en uno de tres grupos: control negativo (alimentación basal); aflatoxina B1 (alimentación basal + 2 ppm de AFB1) y DFM (alimentación basal + 2 ppm de AFB1 + producto comercial de Bacillus). Cada grupo tenía tres réplicas de 10 pollos (n = 30/grupo). El peso corporal (BW) y la ganancia de peso corporal (BWG) se calcularon semanalmente, mientras que la ingesta de alimento (FI) y la conversión alimentaria (FCR) se determinaron cuando los pollos tenían 21 días de edad. Al día 21 de edad, todos los pollos se sangraron, se recolectaron muestras gastrointestinales y se pesaron el bazo y la bolsa de Fabricio. Este estudio confirmó que 2 ppm de aflatoxina B1 causan efectos detrimentales graves sobre los parámetros productivos, bioquímicos e inmunológicos, generando lesiones hepáticas en pollos de engorde (P < 0.05). Sin embargo, también se observó que la suplementación con el producto comercial de Bacillus proporcionó efectos benéficos que podrían ayudar a mejorar la función de la barrera intestinal, las actividades antiinflamatorias y antioxidantes, así como la inmunomodulación humoral y celular. Los resultados del presente estudio sugieren que este producto comercial de Bacillus agregado a una concentración de 106 esporas/gramo de alimento puede usarse para contrarrestar los efectos negativos que se producen cuando las aves consumen dietas contaminadas con aflatoxina B1, mostrando efectos beneficiosos en los parámetros productivos, peso relativo de órganos, lesiones hepáticas, respuesta inmune y variables bioquímicas séricas. La adición de este Bacillus podría mitigar y disminuir los problemas de aflatoxicosis en la industria avícola, mejorando la seguridad alimentaria, los problemas de salud pública y los beneficios económicos. Se requieren estudios futuros para dilucidar completamente los mecanismos específicos por los cuales este producto comercial con Bacillus contrarresta los efectos tóxicos de la aflatoxina B1.

Evaluation of Ascorbic Acid or Curcumin Formulated in a Solid Dispersion on Salmonella Enteritidis Infection and Intestinal Integrity in Broiler Chickens.

Two experimental models were conducted to evaluate and compare the effect of ascorbic acid (AA) or curcumin formulated in a solid dispersion (SD-CUR) as prophylactic or therapeutic alternatives to prevent or control S. Enteritidis (SE) infection in broiler chickens. In the prophylactic model, dietary administration of AA showed a significant reduction in SE counts in crop compared to the positive control (PC) group (p < 0.05), whereas in cecal tonsils (CT), SD-CUR significantly reduced SE recovery. Superoxide dismutase (SOD) activity was significantly higher in chickens supplemented with AA or SD-CUR, and total intestinal IgA levels were significantly lower in both treatments when compared to the PC group. Serum fluorescein isothiocyanate-dextran (FITC-d) levels were reduced by SD-CUR compared to PC, while AA presented significantly lower total aerobic bacteria. In the therapeutic model, only the dietary administration of AA significantly decreased SE in crop and CT on days 3 and 10 post-challenge. FITC-d levels were significantly lower in both treated groups in comparison to PC, but IgA levels were significantly reduced only by AA. The results suggest that dietary AA and SD-CUR have different modes of action to reduce SE intestinal colonization in two different challenge models in broiler chickens.

Evaluation of the Antimicrobial and Anti-inflammatory Properties of Bacillus-DFM (Norum™) in Broiler Chickens Infected With Salmonella Enteritidis.

Restrictions of in-feed antibiotics use in poultry has pushed research toward finding appropriate alternatives such as Direct-Fed Microbials (DFM). In this study, previously tested Bacillus isolates (B. subtilis and B. amyloliquefaciens) were used to evaluate their therapeutic and prophylactic effects against Salmonella enterica serovar Enteritidis (S. Enteritidis) in broiler chickens. For this purpose, initial antibacterial activity of Bacillus-DFM (104 spores/g or 106 spores/g) against S. Enteritidis colonization in crop, proventriculus and intestine was investigated using an in vitro digestive model. Furthermore, to evaluate therapeutic and prophylactic effects of Bacillus-DFM (104 spores/g) against S. Enteritidis colonization, altogether 60 (n = 30/group) and 30 (n = 15/group) 1-day-old broiler chickens were randomly allocated to either DFM or control group (without Bacillus-DFM), respectively. Chickens were orally gavaged with 104 cfu of S. Enteritidis per chicken at 1-day old, and cecal tonsils (CT) and crop were collected 3 and 10 days later during the therapeutic study, whereas they were orally gavaged with 107 cfu of S. Enteritidis per chicken at 6-day-old, and CT and crop were collected 24 h later from two independent trials during the prophylactic study. Serum superoxide dismutase (SOD), FITC-d and intestinal IgA levels were reported for both chicken studies, in addition cecal microbiota analysis was performed during the therapeutic study. DFM significantly reduced S. Enteritidis concentration in the intestine compartment, and in both proventriculus and intestine compartments as compared to the control when used at 104 spores/g and 106 spores/g, respectively (p < 0.05). DFM significantly reduced FITC-d and IgA as well as SOD and IgA levels (p < 0.05) compared to the control in therapeutic and prophylactic studies, respectively. Interestingly, in the therapeutic study, there were significant differences in bacterial community structure and predicted metabolic pathways between DFM and control. Likewise, phylum Actinobacteria and the genera Bifidobacterium, Roseburia, Proteus, and cc_115 were decreased, while the genus Streptococcus was enriched significantly in the DFM group as compared to the control (MetagenomeSeq, p < 0.05). Thus, the overall results suggest that the Bacillus-DFM can reduce S. Enteritidis colonization and improve the intestinal health in chickens through mechanism(s) that might involve the modulation of gut microbiota and their metabolic pathways.

Isolation and Identification of Lactic Acid Bacteria Probiotic Culture Candidates for the Treatment of Salmonella enterica Serovar Enteritidis in Neonatal Turkey Poults.

The effect of Lactobacillus spp.-based probiotic candidates on Salmonella enterica serovar Enteritidis (SE) colonization was evaluated in two separate experiments. In each experiment, sixty-one day-of-hatch female turkey poults were obtained from a local hatchery. In both experiments, poults were challenged via oral gavage with 104 cfu/poult of SE and randomly allocated to one of two groups (n = 30 poults): (1) the positive control group and (2) the probiotic treated group. Heated brooder batteries were used for housing each group separately and poults were allowed ad libitum access to water and unmedicated turkey starter feed. 1 h following the SE challenge, poults were treated with 106 cfu/poult of probiotic culture via oral gavage or phosphate-buffered saline （PBS）to control groups. A total of 24 h post-treatment, poults were euthanized and the ceca and cecal tonsils from twenty poults were collected aseptically for SE recovery. In both trials, a significant reduction in the incidence and log10 cfu/g of SE were observed in poults treated with the probiotic when compared with control poults (p ≤ 0.05). The results of the present study suggest that the administration of this lactic acid-producing bacteria (LAB)-based probiotic 1 h after an SE challenge can be useful in reducing the cecal colonization of this pathogen in neonatal poults.

Impact of a Bacillus Direct-Fed Microbial on Growth Performance, Intestinal Barrier Integrity, Necrotic Enteritis Lesions, and Ileal Microbiota in Broiler Chickens Using a Laboratory Challenge Model.

Decreases in the use of antibiotics and anticoccidials in the poultry industry have risen the appearance of necrotic enteritis (NE). The purpose of this study was to evaluate the effect of a Bacillus direct-fed microbial (DFM) on growth performance, intestinal integrity, NE lesions and ileal microbiota using a previously established NE-challenged model. At day-of-hatch, chicks were randomly assigned to three different groups: Negative control (NC), Positive control (PC) challenged with Salmonella Typhimurium (day 1), Eimeria maxima (EM, day 13) and Clostridium perfringens (CP, day 18-19), and Bacillus-DFM group (DFM) challenged as the PC. Body weight (BW) and body weight gain (BWG) were measured weekly. Total feed intake (FI) and feed conversion ratio (FCR) were evaluated at day 21. Liver samples were collected to assess bacterial translocation and blood samples were used to measure superoxide dismutase (SOD) and fluorescein isothiocyanate-dextran (FITC-d). Intestinal contents were obtained for determination of total IgA and microbiota analysis. NE lesion scores (LS) were performed at day 21. Chickens consuming the DFM significantly improved BW and had a numerically more efficient FCR compared to PC at day 21. Additionally, there were no significant differences in FCR between the DFM group and NC. Furthermore, the DFM group showed significant reductions in LS, IgA and FITC-d levels compared to the PC. However, there were no significant differences in SOD between the groups. The microbiota analysis indicated that the phylum Proteobacteria was significantly reduced in the DFM group in comparison to PC. At the genus level, Clostridium, Turicibacter, Enterococcus, and Streptococcus were reduced, whereas, Lactobacillus and Bacillus were increased in the DFM group as compared to PC (p < 0.05). Likewise, the DFM significantly reduced CP as compared to PC. In contrary, no significant differences were observed in bacterial composition between NC vs. DFM. In addition, beta diversity showed significant differences in the microbial community structure between NC vs. PC, and PC vs. DFM. These results suggest that the dietary inclusion of a selected DFM could mitigate the complex negative impacts caused by NE possibly through mechanism(s) that might involve modulation of the gut microbiota.