Supplementation of lactobacillus-fermented rapeseed meal in broiler diet reduces Campylobacter jejuni cecal colonization and limits the l-tryptophan and l-histidine biosynthesis pathways.

BACKGROUND: Campylobacter jejuni (C. jejuni), a widely distributed global foodborne pathogen, primarily linked with contaminated chicken meat, poses a significant health risk. Reducing the abundance of this pathogen in poultry meat is challenging but essential. This study assessed the impact of Lactobacillus-fermented rapeseed meal (LFRM) on broilers exposed to C. jejuni-contaminated litter, evaluating growth performance, Campylobacter levels, and metagenomic profile. RESULTS: By day 35, the litter contamination successfully colonized broilers with Campylobacter spp., particularly C. jejuni. In the grower phase, LFRM improved (P < 0.05) body weight and daily weight gain, resulting in a 9.2% better feed conversion ratio during the pre-challenge period (the period before artificial infection; days 13-20). The LFRM also reduced the C. jejuni concentration in the ceca (P < 0.05), without altering alpha and beta diversity. However, metagenomic data analysis revealed LFRM targeted a reduction in the abundance of C. jejuni biosynthetic pathways of l-tryptophan and l-histidine and gene families associated with transcription and virulence factors while also possibly leading to selected stress-induced resistance mechanisms. CONCLUSION: The study demonstrated that LFRM inclusion improved growth and decreased cecal Campylobacter spp. concentration and the relative abundance of pivotal C. jejuni genes. Performance benefits likely resulted from LFRM metabolites. At the molecular level, LFRM may have reduced C. jejuni colonization, likely by decreasing the abundance of energy transduction and l-histidine and l-tryptophan biosynthesis genes otherwise required for bacterial survival and increased virulence. © 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Impact of precursor-derived peracetic acid on post-weaning diarrhea, intestinal microbiota, and predicted microbial functional genes in weaned pigs.

Post-weaning diarrhea affects piglets in the nursery phase of production, leading to a substantial impact both at the farm and financial levels. The multifactorial etiology of this disease includes housing conditions, pig genetics, microbial composition, and metagenomic assets. Among the common therapeutic approaches, the widely used zinc oxide underwent a European Union ban in 2022 due to its negative environmental impact and correlation to increased antimicrobial resistance. During this study, we have tested two levels of inclusion of the potential antimicrobial alternative peracetic acid, delivered in water via the hydrolysis of the precursors sodium percarbonate and tetraacetylethylenediamine, in comparison to zinc oxide and an untreated control during a 2-week animal study. We assessed the microbial composition and predicted the metagenome, together with performance and physiological parameters, in order to describe the microbial functional role in etiopathology. Both zinc oxide and peracetic acid resulted in amelioration of the diarrheal status by the end of the trial period, with noticeable zinc oxide effects visible from the first week. This was accompanied by improved performance when compared to the first-week figures and a decreased stomach pH in both peracetic acid levels. A significant reduction in both stomach and caecal Proteobacteria was recorded in the zinc oxide group, and a significant reduction of Campylobacter in the stomach was reported for both zinc oxide and one of the peracetic acid concentrations. Among other functional differences, we found that the predicted ortholog for the zonula occludens toxin, a virulence factor present in pathogens like Escherichia coli and Campylobacter jejuni, was less abundant in the stomach of treated pigs compared to the control group. In water, peracetic acid delivered via precursor hydrolysis has the potential to be a valid intervention, an alternative to antimicrobial, to assist the weaning of piglets. Our findings support the view that post-weaning diarrhea is a complex multifactorial disease with an important metagenomic component characterized by the differential abundance of specific predicted orthologs and microbial genera in the stomach and caecum of pigs.

Estimating the contribution of the porcine fecal core microbiota to metabolite production via mathematical modeling and in vitro fermentation.

The swine gut microbiota is a complex ecosystem found throughout the gastrointestinal tract, with multiple exchanges with the host and whose composition is linked to both external and internal factors, such as diet or breed. Diet, probiotic, or prebiotic interventions have been designed to boost beneficial host-microbiota interactions, such as the production of anti-inflammatory molecules, or the fermentation of otherwise undigested resources. In parallel, a smaller microbial population, shared among the same host species, independent of external or internal factors, has been described and defined as the "core microbiota." Therapies targeting the core microbiota could possibly lead to more precise and long-lasting effects. However, the metabolic role of the porcine core microbiota, especially in relation to the rest of the microbial community, is currently missing. We present here the first dynamic model of the porcine core microbiota, which we used to estimate the core-microbiota metabolite production and to forecast the effect of a synbiotic intervention targeting the core genera of the core microbiota. We developed a community model in which a total of 17 microbial groups were established based on culture-based information of representative species. First, the model parameters were estimated, and the resulting model simulations were compared favorably with in vitro experimentation. The model was then used to predict the microbial dynamics of the core and non-core members under different experimental conditions. Therefore, it was able to theorize the main-metabolite core microbiota contribution, hypothesizing that it could be mainly responsible for acetate and propionate, but not for butyrate production.IMPORTANCECurrently, little information is present in the literature to describe the generic metabolic role of the porcine core microbiota or to inform on the effect of interventions targeting the core genera. Moreover, both in vitro and in vivo experimentations aiming to explore the core microbiota dynamics are technically demanding, expensive, or restricted by ethical considerations. Modeling approaches can be used as an initial exploratory tool to develop hypotheses for targeted experimentation. Our mathematical model provides initial information on the microbial and metabolite dynamics of the core microbiota in relation to diet and therapeutic intervention.

Encapsulated peracetic acid as a valid broad-spectrum antimicrobial alternative, leading to beneficial microbiota compositional changes and enhanced performance in broiler chickens.

BACKGROUND: Antimicrobial alternatives are urgently needed, including for poultry production systems. In this study, we tested the potential broad-range antimicrobial alternative peracetic acid, delivered in feed via the hydrolysis of encapsulated precursors through a 28-day study using 375 Ross 308 broiler chickens. We tested two peracetic acid concentrations, 30 and 80 mg/kg on birds housed on re-used litter, and we evaluated the impact of both levels on gut microbial communities, bacterial concentration, antimicrobial resistance genes relative abundance and growth performance when compared to control birds housed on either clean or re-used litter. RESULTS: Body weight gain and feed conversion ratio improved in peracetic acid fed birds. At d 28, birds given 30 mg/kg of peracetic acid had a decreased Firmicutes and an increased Proteobacteria abundance in the jejunum, accompanied by an increase in Bacillus, Flavonifractor and Rombustia in the caeca, and a decreased abundance of tetracycline resistance genes. Chicken given 80 mg/kg of peracetic acid had greater caecal abundance of macrolides lincosamides and streptogramins resistance genes. Growth performance on clean litter was reduced compared to re-used litter, which concurred with increased caecal abundance of Blautia, decreased caecal abundance of Escherichia/Shigella, Anaerostipes and Jeotgalicoccus, and greater gene abundance of vancomycin, tetracycline, and macrolides resistance genes. CONCLUSIONS: Peracetic acid could be used as a safe broad-spectrum antimicrobial alternative in broilers. Encapsulated precursors were able to reduce the bacterial concentration in the jejunum whilst promoting the proliferation of probiotic genera in the caeca, especially at the low peracetic acid concentrations tested, and improve growth performance. Moreover, our findings offer further insights on potential benefits of rearing birds on re-used litter, suggesting that the latter could be associated with better performance and reduced antimicrobial resistance risk compared to clean litter rearing.

Precursor-derived in-water peracetic acid impacts on broiler performance, gut microbiota, and antimicrobial resistance genes.

Past antimicrobial misuse has led to the spread of antimicrobial resistance amongst pathogens, reportedly a major public health threat. Attempts to reduce the spread of antimicrobial resistant (AMR) bacteria are in place worldwide, among which finding alternatives to antimicrobials have a pivotal role. Such molecules could be used as "green alternatives" to reduce the bacterial load either by targeting specific bacterial groups or more generically, functioning as biocides when delivered in vivo. In this study, the effect of in-water peracetic acid as a broad-spectrum antibiotic alternative for broilers was assessed via hydrolysis of precursors sodium percarbonate and tetraacetylethylenediamine. Six equidistant peracetic acid levels were tested from 0 to 50 ppm using four pens per treatment and 4 birds per pen (i.e., 16 birds per treatment and 96 in total). Peracetic acid was administered daily from d 7 to 14 of age whilst measuring performance parameters and end-point bacterial concentration (qPCR) in crop, jejunum, and ceca, as well as crop 16S sequencing. PAA treatment, especially at 20, 30, and 40 ppm, increased body weight at d 14, and feed intake during PAA exposure compared to control (P < 0.05). PAA decreased bacterial concentration in the crop only (P < 0.05), which was correlated to better performance (P < 0.05). Although no differences in alpha- and beta-diversity were found, it was observed a reduction of Lactobacillus (P < 0.05) and Flectobacillus (P < 0.05) in most treatments compared to control, together with an increased abundance of predicted 4-aminobutanoate degradation (V) pathway. The analysis of the AMR genes did not point towards any systematic differences in gene abundance due to treatment administration. This, together with the rest of our observations could indicate that proximal gut microbiota modulation could result in performance amelioration. Thus, peracetic acid may be a valid antimicrobial alternative that could also positively affect performance.

Recent Progress in Phage Therapy to Modulate Multidrug-Resistant Acinetobacter baumannii, including in Human and Poultry.

Acinetobacter baumannii is a multidrug-resistant and invasive pathogen associated with the etiopathology of both an increasing number of nosocomial infections and is of relevance to poultry production systems. Multidrug-resistant Acinetobacter baumannii has been reported in connection to severe challenges to clinical treatment, mostly due to an increased rate of resistance to carbapenems. Amid the possible strategies aiming to reduce the insurgence of antimicrobial resistance, phage therapy has gained particular importance for the treatment of bacterial infections. This review summarizes the different phage-therapy approaches currently in use for multiple-drug resistant Acinetobacter baumannii, including single phage therapy, phage cocktails, phage-antibiotic combination therapy, phage-derived enzymes active on Acinetobacter baumannii and some novel technologies based on phage interventions. Although phage therapy represents a potential treatment solution for multidrug-resistant Acinetobacter baumannii, further research is needed to unravel some unanswered questions, especially in regard to its in vivo applications, before possible routine clinical use.

Bacterial concentration and Campylobacter spp. quantification differ when fresh or ultra-frozen samples are analysed over time using molecular biology and culture-based methods.

The study aimed to delineate the robustness of the culture-based and molecular biology methods to assess the total bacterial concentration and Campylobacter jejuni (C. jejuni) quantification in caecal content, analysed as fresh or after being stored immediately at ultra-low (-80°C) temperature at different time points (for 3, 7, 14, 28 and 62 days post collection). The caecal content was collected from birds that were artificially colonised with C. jejuni (in-vivo), and quantification was performed using both colony-forming unit (CFU) and qPCR. The results showed that storage time affected the output of culture-based analyses but mostly did not alter concentration retrieved via qPCR. After an initial ~4.5 log10 reduction in CFU observed from fresh (day 0) to frozen samples, bacterial concentration retrieved with culture-based methods seemed to be constant in samples frozen for 3 to 62 days, indicating a possible threshold for C. jejuni loss of viability due to effect of storage temperature. Ranking order analyses, revealed that the molecular biology technique was able to attribute somewhat the same relative C. jejuni concentrations to the samples analysed via qPCR. However, day 0 measurements from culture-based methods were associated with the absence of or negatively weak correlations with the rest of the time points, but ranking order was maintained from day 3 onwards. On the other hand, ranking order correlations were less constant when measuring total bacterial concentration through qPCR. The study suggests that if biological samples can't be analysed as fresh (immediately after collection) and have to be stored prior to analysis, then storage at -80°C samples be recommended to avoid the temporal-dependent effects on C. jejuni concentrations. In addition, irrespective of the method of analysis, an initial loss of CFU must be factored in when interpreting the results obtained from frozen samples.

Lim mineralization protein is involved in the premature calvarial ossification in sporadic craniosynostoses.

Sporadic mono-sutural craniosynostosis represents a highly prevalent regional bone disorder, where a single cranial suture undergoes premature ossification due to a generally unclear etiopathogenesis. The LIM mineralization protein (LMP) was recently described as an efficient osteogenic molecule involved in osteoblast differentiation, expressed in calvarial tissues upon corticosteroid-osteogenic induction and used as a potent inducer of bone formation in several animal models. In this study, calvarial cells isolated from both prematurely fused and physiologically patent sutures of children with sporadic craniosynostosis, were used as an in vitro paradigmatic model for the study of the molecular events involved in calvarial osteogenesis, focusing on the possible role of the LMP-related osteogenic signaling. Calvarial cells isolated from both patent and fused sutures expressed a mesenchymal-like immunophenotype. Cells isolated from fused sutures displayed an increased osteogenic potential, being able to undergo spontaneous mineralization and premature response to osteogenic induction, leading to in vitro bone nodule formation. The expression of LMP and its target genes (bone morphogenetic protein-2, osteocalcin and Runt-related transcription factor 2) was significantly up-regulated in cells derived from the fused sutures. Upon silencing the expression of LMP in fused suture-derived cells, the osteogenic potential along with the expression of osteo-specific transcription factors decreased, restoring the "physiologic" cell behavior. These results suggested that: 1. mesenchymal cells residing in fused sutures display a constitutionally active osteogenic disposition leading to the premature suture ossification; 2. the molecular basis of the overactive osteogenic process may at least in part involve a deregulation of the LMP-related pathway in calvarial cells.