RESUMO
Salmonellosis is a poultry industry and public health concern worldwide. Recently, Salmonella enterica serovar Heidelberg (SH) has been reported in broilers in Brazil. The effect of feeding a blend of three strains of Bacillus subtilis (PRO) was studied in broilers orally challenged (107 CFU/chick) or not with a SH isolated in south of Brazil (UFPR1 strain). Twelve male Cobb 500 broilers per pen were randomly assigned to six treatments in a 3 × 2 factorial experiment where PRO was added at 0, 250, or 500 g/ton of broiler feed and fed to either SH-challenged (SH Control, SH + PRO 250, and SH + PRO 500) or non-challenged birds (Control, PRO 250, and PRO 500). Broiler performance, histologic alterations in intestinal morphology, Salmonella quantification and immune cells counts in liver (macrophages, T CD4+ and T CD8+) were analyzed. Changes in the intestinal microbiota of broilers were also studied by metagenomics for Control, SH Control, SH + PRO 250, and SH + PRO 500 only. Feeding PRO at 250 or 500 g/ton reduced SH counts and incidence in liver and cecum at 21 days of age. It was observed that PRO groups increased the macrophage mobilization to the liver in SH-challenged birds (P < 0.05) but reduced these cells in the liver of non-challenged birds, showing an interesting immune cell dynamics effect. PRO at 250 g/ton did not affect gut histology, but improved animal performance (P < 0.05) while PRO at 500/ton did not affect animal performance but increased histologic alteration related to activation of the defense response in the ileum in SH challenged birds compared to control birds (P < 0.05). SH + PRO 500 group presented a more diverse cecal microbiota (Shannon-Wiener index; P < 0.05) compared to Control and SH Control groups; while SH + PRO 250 had greater ileal richness (JackkNife index) compared to Control (P < 0.05). PRO was effective in reducing Salmonella colonization in liver and cecum when fed at 250 or 500 g/ton to broilers inoculated with SH strain UFPR1. PRO promotes positive alterations in performance (at 250 g/ton), immune modulatory effect in the gastrointestinal tract, SH reduction, and intestinal microbiota modulation.
RESUMO
Salmonella enterica serovar Heidelberg is a human pathogen also found in broilers. A strain (UFPR1) has been associated with field reports of resistance to short-chain organic acids (SCOA) in broilers in the South of Brazil, but was susceptible to a Bacillus subtilis-based probiotic added in feed in a related study. This work aimed to (i) report clinical symptoms caused by SH UFPR1 in broilers, (ii) study its susceptibility to some antibiotics in vitro, and (iii) SCOA in vivo; and (iv) relate these phenotypic observations with its genome characteristics. Two in vivo trials used 1-day-old chicks housed for 21 days in 8 sterilized isolated negative pressure rooms with 4 battery cages of 12 birds each. Birds were challenged or not with 107 CFU/bird of SH UFPR1 orally and exposed or not to SCOA in a 2 × 2 factorial design. Zootechnical parameters were unaffected (P > 0.05), no clinical signs were observed, and few cecal and hepatic histologic and immune-related alterations were seen, in birds challenged with SH. Formic and propionic acids added together in drinking water, fumaric and benzoic acid in feed (Trial 1), and coated calcium butyrate in feed (Trial 2) did not reduce the SH isolation frequencies seen in cecum and liver in broilers after SH challenge (P > 0.05). SH UFPR1 was susceptible to amikacin, amoxicillin + clavulanate, ceftiofur, cephalexin, doxycycline and oxytetracycline; and mildly susceptible to ampicillin + sulbactam, cephalothin, ciprofloxacin, enrofloxacin, and gentamycin in an in vitro minimum inhibitory concentration model using Mueller-Hinton agar. The whole genome of SH UFPR1 was sequenced and consisted of a circular chromosome, spanning 4,760,321 bp with 52.18% of GC-content encoding 84 tRNA, 22 rRNA, and 4,427 protein-coding genes. The comparison between SH UFPR1 genome and a multidrug-resistant SL476 strain revealed 11 missing genomic fragments and 5 insertions related to bgt, bgr, and rpoS genes. The deleted genes codify proteins associated with cell cycle regulation, virulence, drug resistance, cellular adhesion, and salt efflux which collectively reveal key aspects of the evolution and adaptation of SH strains such as organic acids resistance and antibiotic sensitivity and provide information relevant to the control of SH in poultry.
