Chicken Intestinal Tract Research Articles

Penalization and Color Code Technical Approaches for Method Greenness and Whiteness Appraisal in Veterinary Medication: Assay of Toltrazuril Suspension.

Intestinal coccidiosis is a debilitating disease in poultry and livestock, leading to economic impact worldwide. Coccidiosis is prevented and treated in broilers by the inclusion of anticoccidials in feed. Toltrazuril is administered in potable water to treat coccidiosis. Three robust analytical methods for the quantitation of toltrazuril in pure and pharmaceutical formulations are developed. Furthermore, ecological metrics, either penalization- or color-code-based techniques, are applied for the appraisal of assays. First, second-derivative (Δλ; 5 nm) spectrophotometric method is used. Toltrazuril is measured from peak to peak at 244-260 nm within a linearity range of 5-25 μg/mL. The second method is an HPTLC analysis performed on an aluminum sheet of silica gel using ethyl acetate-methanol-ammonium chloride buffer-water (8:1:0.5:0.5, by volume respectively) as the elution phase. Toltrazuril, at a retardation factor of 0.66 ± 0.01, is linearly determined in the range of 1-9 μg/spot at 243 nm. The third method is reversed-phase HPLC with diode array detection, using an Agilent C18 column (5 μm, 4.6 × 150 mm) in isocratic elution mode at 1 mL/min flow rate with a mobile phase of acetonitrile and water in a ratio of 80:20 (v/v). Toltrazuril elutes at a retention time of 2.58 ± 0.1 min and is linearly determined at 243 nm in the range of 0.25-25 μg/mL. Calculated 2D-values and peak areas are highly correlated to their corresponding drug concentrations at coefficients: r > 0.999. All methods were International Council of Harmonization (ICH) validated and applied to the dosage form with satisfactory % recoveries (97-103%). Statistical comparisons versus reported one using t-test and F-test disclose insignificant variation. In examining greenness and whiteness norms, the proposed methods were evaluated and ranked alongside four different reported methods. The proposed methods are green, accurate, and can be applied in routine QC for the determination of toltrazuril in pharmaceutical formulations. Intestinal coccidiosis substantially affects the chicken intestinal tract leading to reduced growth. Toltrazuril is used for the treatment and prevention of intestinal coccidiosis. Three robust, accurate, and precise analytical methods are developed for toltrazuril determination in pure and pharmaceutical formulations. All proposed methods were ecologically assessed and compared with published ones.

Distribution and spatiotemporal development of organised lymphoid tissues in the chicken intestinal tract

Chickens exhibit a distinct immune architecture characterised by the absence of draining lymph nodes and the presence of a well-developed mucosa-associated lymphoid tissue. The structure and spatiotemporal development of chicken lymphoid tissues in the intestine are poorly documented. The macroscopically indistinct structure of chicken Peyer's patches has impeded studies into their development. The generation of CSF1R-eGFP reporter transgenic chickens enables visualisation of the development, organisation and extent of chicken lymphoid tissues by unique macroscopic views. CSF1R-eGFP reporter transgenic chickens were used to investigate the distribution and spatiotemporal development of PP and caecal tonsils in embryonic day 18 to 8-week-old chickens. Peyer's patch anlagen are present at ED18 with a similar frequency and distribution pattern observed in 2- and 8-week-old chickens. These findings can support in ovo and post-hatch mucosal vaccination strategies and the development of vaccine delivery systems targeted to the specialized epithelium overlying the Peyer's patches.

Colonization of chickens with competitive exclusion products results in extensive differences in metabolite composition in cecal digesta

The concept of competitive exclusion is well established in poultry and different products are used to suppress the multiplication of enteric pathogens in the chicken intestinal tract. While the effect has been repeatedly confirmed, the specific principles of competitive exclusion are less clear. The aim of the study was to compare metabolites in the cecal digesta of differently colonized chickens. Metabolites in the cecal contents of chickens treated with a commercial competitive exclusion product or with an experimental product consisting of 23 gut anaerobes or in control untreated chickens were determined by mass spectrometry. Extensive differences in metabolite composition among the digesta of all 3 groups of chickens were recorded. Out of 1,706 detected compounds, 495 and 279 were differently abundant in the chicks treated with a commercial or experimental competitive exclusion product in comparison to the control group, respectively. Soyasaponins, betaine, carnitine, glutamate, tyramine, phenylacetaldehyde, or 3-methyladenine were more abundant in the digesta of control chicks while 4-oxododecanedioic acid, nucleotides, dipeptides, amino acids (except for glutamate), and vitamins were enriched in the digesta of chickens colonized by competitive exclusion products. Metabolites enriched in the digesta of control chicks can be classified as of plant feed origin released in the digesta by degradative activities of the chicken. Some of these molecules disappeared from the digesta of chicks colonized by complex microbiota due to them being metabolized. Instead, nucleotides, amino acids, and vitamins increased in the digesta of colonized chicks as a consequence of the additional digestive potential brought to the cecum by microbiota from competitive exclusion products. It is therefore possible to affect metabolite profiles in the chicken cecum by its colonization with selected bacterial species.

Dynamic distribution of gut microbiota in posthatching chicks and its relationship with average daily gain

The colonization and development of gut microbiota are essential for the health and growth of chicks after hatching. However, the colonization and prevalence of gut microbiota have not been well characterized, and knowledge of which microbes and their relationship with average daily gain in chicks is still limited. This study characterized the dynamic succession of microbiota in the intestinal tract of chicks and investigated its relationship with daily weight gain. A total of 121 fecal samples across 7 time points from d 0 to 10 posthatching were collected from 19 chicks randomly selected from 1,950 chicks. Using 16S rRNA gene sequencing examined microbial composition of fecal samples. The observed species index of alpha diversity increased with age, gradually achieving stability at 3 d of age. The microbiota of chicks after hatching was primarily Clostridium_sensu_stricto_1 (34.49%), and its relative abundance diminishes with age. In contrast, Lactobacillus had a low relative abundance in the first 2 d after hatching and gradually increased with age. Predicted functional capacities found that the microbiota of early-stage posthatching (d 0 and 1 after hatching) was involved in metabolism, including amino acid metabolism, metabolism of cofactors and vitamins, and nitrogen metabolism. However, at the later stage posthatching (from d 3-10 after hatching), the intestinal microbial function was involved in carbohydrate metabolism, amino acid metabolism, cell growth and death, and methane metabolism. It was identified that 47 operational taxonomic units were associated with average daily gain of chicks, 12 of which were annotated with Lactobacillus and significantly positive associated with average daily gain. In addition, Clostridium_sensu_stricto_1 was significantly negatively associated with average daily gain. Taken together, we characterized the dynamic successions of intestinal microbiota in hatching chicks. The intestinal microbiota of chicks has an impact on the host average daily gain. Our findings should be instrumental in improving local chick production.

Cecal Microbiota Development and Physiological Responses of Broilers Following Early Life Microbial Inoculation Using Different Delivery Methods and Microbial Sources.

Broilers in intensive systems may lack commensal microbes that have coevolved with chickens in nature. This study evaluated the effects of microbial inocula and delivery methods applied to day-old chicks on the development of the cecal microbiota. Specifically, chicks were inoculated with cecal contents or microbial cultures, and the efficacies of three delivery methods (oral gavage, spraying inoculum into the bedding, and cohousing) were evaluated. Also, a competitive study evaluated the colonization ability of bacteria sourced from extensive or intensive poultry production systems. The microbiota of inoculated birds presented higher phylogenetic diversity values (PD) and higher relative abundance values of Bacteroidetes, compared with a control. Additionally, a reduction in the ileal villus height/crypt depth ratio and increased cecal IL-6, IL-10, propionate, and valerate concentrations were observed in birds that were inoculated with cecal contents. Across the experiments, the chicks in the control groups presented higher relative abundance values of Escherichia/Shigella than did the inoculated birds. Specific microbes from intensively or extensively raised chickens were able to colonize the ceca, and inocula from intensive production systems promoted higher relative abundance values of Escherichia/Shigella. We concluded that Alistipes, Bacteroides, Barnesiella, Mediterranea, Parabacteroides, Megamonas, and Phascolarctobacterium are effective colonizers of the broiler ceca. In addition, oral gavage, spray, and cohousing can be used as delivery methods for microbial transplantation, as indicated by their effects on the cecal microbiota, intestinal morphology, short-chain fatty acids concentration, and cytokine/chemokine levels. These findings will guide future research on the development of next-generation probiotics that are able to colonize and persist in the chicken intestinal tract after a single exposure. IMPORTANCE The strict biosecurity procedures employed in the poultry industry may inadvertently hinder the transmission of beneficial commensal bacteria that chickens would encounter in natural environments. This research aims at identifying bacteria that can colonize and persist in the chicken gut after a single exposure. We evaluated different microbial inocula that were obtained from healthy adult chicken donors as well as three delivery methods for their effects on microbiota composition and bird physiology. In addition, we conducted a competitive assay to test the colonization abilities of bacteria sourced from intensively versus extensively raised chickens. Our results indicated that some bacteria are consistently increased in birds that are exposed to microbial inoculations. These bacteria can be isolated and employed in future research on the development of next-generation probiotics that contain species that are highly adapted to the chicken gut.

Tryptanthrin Reduces Campylobacter jejuni Colonization in the Chicken Gut by a Bactericidal Mechanism.

Campylobacter jejuni is a leading cause of foodborne bacterial gastroenteritis worldwide, and raw or undercooked chicken meat is considered the major source of human campylobacteriosis. In this study, we identified 36 compounds that showed inhibitory effects on C. jejuni growth at low concentrations by screening a chemical compound library. Three of the 36 compounds were herbal compounds, including tryptanthrin (TRP), an indoloquinazoline alkaloid. TRP has been reported to have a variety of biological properties, such as antimicrobial, anti-inflammatory, and antitumor activities, but there was previously no information about its anti-C. jejuni activity. We further conducted in vitro and in vivo experiments to evaluate the potential of TRP for the control of C. jejuni in chicken farms. The MIC of TRP for C. jejuni was much lower than that of 13 other herbal compounds that were previously reported to have anti-C. jejuni activities. Time-kill assays under growing and nongrowing conditions demonstrated that TRP has bactericidal activity against C. jejuni. In addition, TRP showed a narrow-spectrum antimicrobial effect against C. jejuni, and there was little potential for the development of TRP-resistant C. jejuni during serially passaged culture. In chick infection experiments, the administration of TRP in drinking water significantly reduced the cecal colonization of C. jejuni when TRP was used either before or after C. jejuni infection. These data suggest that TRP is effective for the control of C. jejuni in chicken farms. IMPORTANCE Campylobacter is a widespread pathogen in the food chain of chickens. Once chickens become infected, large numbers of Campylobacter cells are excreted in their feces. The development of an effective material for reducing the amount of Campylobacter in the chicken intestinal tract will make it possible to reduce the contamination of the food chain with Campylobacter and to produce safe and secure chicken meat. In the present study, in vivo experiments revealed that the use of an herbal compound, tryptanthrin, significantly reduced the number of Campylobacter cells in the chicken gut by a bactericidal mechanism. Furthermore, our in vitro experiments demonstrated that, compared with the other herbal compounds, tryptanthrin achieved antimicrobial activity against C. jejuni at the lowest concentration. The use of tryptanthrin may lead to the development of a novel control measure for reducing the colonization of C. jejuni in the food chain.

Microencapsulated phage composites with increased gastrointestinal stability for the oral treatment of Salmonella colonization in chicken.

Salmonella infection, one of the common epidemics in the livestock and poultry breeding industry, causes great economic losses worldwide. At present, antibiotics are the most commonly used treatment for Salmonella infection, but the widespread use of antibiotics has increased drug resistance to Salmonella. Phage therapy has gradually become an alternative method to control Salmonella infection. However, phage, a specific virus that can infect bacteria, has poor stability and is prone to inactivation during treatment. Microencapsulated phage microspheres can effectively solve this problem. Accordingly, in this study, Salmonella phages were microencapsulated, using the xanthan gum/sodium alginate/CaCl2/chitooligosaccharides method, to improve their gastrointestinal stability. Furthermore, microencapsulated phages were evaluated for in vitro temperature and storage stability and in vivo therapeutic effect. Phage microspheres prepared with 1 g/100 mL xanthan gum, 2 g/100 mL sodium alginate, 2 g/100 mL CaCl2, and 0.6 g/100 mL chitooligosaccharides were regular in shape and stable in the temperature range of 10-30°C. Also, microencapsulated phages showed significantly improved stability in the simulated gastric juice environment than the free phages (p < 0.05). In the simulated intestinal fluid, microencapsulated phages were completely released after 4 h. Moreover, microencapsulated phages showed good storage stability at 4°C. In the in vivo experiments detecting Salmonella colonization in the intestinal tract of chicks, microencapsulated phages showed a better therapeutic effect than the free phages. In conclusion, microencapsulated phages exhibited significantly improved stability, gastric acid resistance, and thereby efficacy than the free phages. Microencapsulated phages can be potentially used as biological control agents against bacterial infections.

Symbiotic bacteria stabilize the intestinal environment by producing phenylpropanoids.

Salmonella enterica serovar Enteritidis (S. Enteritidis) can colonize in the intestinal tract of chickens and transmit to humans. In order to decrypt the mechanism of avian resistance to S. Enteritidis, we utilized two China local chicken breeds to generate the reciprocal crosses (the Cross and the Reverse-cross). The two lines of hybrids were orally inoculated with S. Enteritidis at 2-day old and sampled at 3 days post-inoculation. Along the analysis direction of multi-omics, differential metabolites, functional pathways and correlated microbes, we found that 12 species of microbes thrived upon S. Enteritidis challenge and probably contributed to the intestinal stability in the Cross by enhancing the production of phenylpropanoids. Our findings can help to understand the symbiotic and resistant mechanisms derived from the intestinal microbiota.

Effect of Salmonella Typhimurium Colonization on Microbiota Maturation and Blood Leukocyte Populations in Broiler Chickens.

Simple SummaryReducing Salmonella levels in chickens is vital to reducing the number of human Salmonella infections resulting from contact with contaminated chickens and poultry products. The intestinal tract of chickens is home to a diverse population of bacteria that serve as one of the first lines of defense against pathogenic microbes. Therefore, we sought to understand changes in the intestinal bacterial populations following Salmonella infection. Our results showed a clear change in the intestinal bacterial structure after Salmonella colonization. An inverse relationship between Salmonella and Lactobacillus and Escherichia was observed as well as an increase in Bacteroides. Additionally, we provide insight into the timing of the host immune response with monocytes and thrombocytes acting early following colonization followed by heterophils. In total, this data demonstrates the ability of Salmonella to change intestinal microbiota composition and highlights genera that may be useful as probiotics to fight Salmonella colonization.Reducing Salmonella in commercial chickens is vital to decreasing human salmonellosis infections resulting from contact with contaminated poultry and poultry products. As the intestinal microbiota plays an important role in preventing pathogen colonization, we sought to understand the relationship between Salmonella infection and the cecal microbiota and the host immune system. Day-of-hatch broiler chicks were assigned to three treatments: control, artificial (SA), and natural (SN) Salmonella infection. At seven days of age, control and SA birds were inoculated with PBS or Salmonella Typhimurium, respectively. Five SA birds were transferred to SN cages to facilitate natural infection. Cecal content and blood samples were collected at 0, 8, 14, and 21 days of age for microbiota and leukocyte analysis, respectively. A significant change in microbiota composition was observed in both groups as noted by a decrease in Lactobacillus and Escherichia and an increase in Bacteroides. Leukocyte analysis revealed a decrease in the percentage of circulating monocytes at 7 days post-infection while a decrease in thrombocyte and an increase in heterophil percentages were seen at 14 days post-infection. Taken together, these results demonstrate the ability of Salmonella to modulate the intestinal microbiota to facilitate colonization. Additionally, results indicated an early role of monocytes and thrombocytes during colonization, followed by heterophils.

Functional Characterization of a HAD Phosphatase Involved in Capsular Polysaccharide Biosynthesis in Campylobacter jejuni.

Campylobacter jejuni is a Gram-negative, pathogenic bacterium found in the intestinal tracts of chickens and many other farm animals. C. jejuni infection results in campylobacteriosis, which can cause nausea, diarrhea, fever, cramps, and death. The surface of the bacterium is coated with a thick layer of sugar known as the capsular polysaccharide. This highly modified polysaccharide contains an unusual d-glucuronamide moiety in serotypes HS:2 and HS:19. Previously, we have demonstrated that a phosphorylated glucuronamide intermediate is synthesized in C. jejuni NCTC 11168 (serotype HS:2) by cumulative reactions of three enzymes: Cj1441, Cj1436/Cj1437, and Cj1438. Cj1441 functions as a UDP-d-glucose dehydrogenase to make UDP-d-glucuronate; then Cj1436 or Cj1437 catalyzes the formation of ethanolamine phosphate or S-serinol phosphate, respectively, and finally Cj1438 catalyzes amide bond formation using d-glucuronate and either ethanolamine phosphate or S-serinol phosphate. Here, we investigated the final d-glucuronamide-modifying enzyme, Cj1435. Cj1435 was shown to catalyze the hydrolysis of the phosphate esters from either the d-glucuronamide of ethanolamine phosphate or S-serinol phosphate. Kinetic constants for a range of substrates were determined, and the stereoselectivity of the enzyme for the hydrolysis of glucuronamide of S-serinol phosphate was established using 31P nuclear magnetic resonance spectroscopy. A bioinformatic analysis of Cj1435 reveals it to be a member of the HAD phosphatase superfamily with a unique DXXE catalytic motif.

High-Throughput Identification of Epigenetic Compounds to Enhance Chicken Host Defense Peptide Gene Expression

Enhancing the synthesis of endogenous host defense peptides (HDPs) has emerged as a novel antibiotic-free approach to infectious disease control and prevention. A number of epigenetic compounds have been identified as HDP inducers and several have proved beneficial in antimicrobial therapy. However, species-specific regulation of HDP synthesis is evident. In attempt to identify epigenetic compounds with potent HDP-inducing activity for poultry-specific application, we developed a stable luciferase reporter cell line, known as HTC/AvBD10-luc, following our earlier construction of HTC/AvBD9-luc. HTC/AvBD10-luc was developed through permanent integration of a chicken macrophage cell line, HTC, with a lentiviral luciferase reporter vector driven by a 4-Kb AvBD10 gene promoter. Using a high throughput screening assay based on the two stable cell lines, we identified 33 hits, mostly being histone deacetylase (HDAC) inhibitors, from a library of 148 epigenetic compounds. Among them, entinostat and its structural analog, tucidinostat, were particularly effective in promoting multiple HDP gene expression in chicken macrophages and jejunal explants. Desirably, neither compounds triggered an inflammatory response. Moreover, oral gavage of entinostat significantly enhanced HDP gene expression in the chicken intestinal tract. Collectively, the high throughput assay proves to be effective in identifying HDP inducers, and both entinostat and tucidinostat could be potentially useful as alternatives to antibiotics to enhance intestinal immunity and disease resistance.

Enrofloxacin Promotes Plasmid-Mediated Conjugation Transfer of Fluoroquinolone-Resistance Gene qnrS

This study aimed to determine the effect of enrofloxacin (ENR) on the transfer of the plasmid-mediated quinolone resistance (PMQR) gene qnrS from opportunistic pathogen Escherichia coli (E2) to Salmonella Enteritidis (SE211) and to analyze the resistance characteristics of SE211-qnrS isolates. The plasmid carrying qnrS gene of E2 was sequenced by Oxford Nanopore technology. The plasmid carrying qnrS gene belonged to incompatibility group IncY. In vitro, the transfer experiment of IncY plasmid was performed by the liquid medium conjugation method. The conjugation transfer frequency of the IncY plasmid was 0.008 ± 0.0006 in the absence of ENR, 0.012 ± 0.003 in 1/32 MICENR, 0.01 ± 0.008 in 1/8 MICENR, and 0.03 ± 0.015 (Mean±SD) in 1/2 MICENR, respectively. After inoculation of E. coli E2 and SE211, chickens were treated with different doses of ENR (3.03, 10, and 50 mg/kg b.w.) for 7 days consecutively. To screen the SE211-qnrS strains from intestinal tract of chickens, the resistance genes and susceptibility of isolates were identified. The amount of E. coli E2 and the copy number of qnrS gene in the chicken intestinal tract were determined by colony counting and qPCR, respectively. In vivo, more SE211-qnrS strains were isolated from the treated group compared with the untreated group. SE211-qnrS strains not only obtained IncY plasmid, but also showed similar resistance phenotype as E2. In conclusion, ENR treatment can promote the spread of a IncY-resistance plasmid carrying the qnrS fluoroquinolone-resistance gene in Escherichia coli and the development of drug-resistant bacteria.

Probiotic Lactobacilli Do Not Protect Chickens against Salmonella Enteritidis Infection by Competitive Exclusion in the Intestinal Tract but in Feed, Outside the Chicken Host.

Lactobacilli are commonly used as probiotics in poultry to improve production parameters and to increase chicken resistance to enteric infections. However, lactobacilli do not efficiently colonise the chicken intestinal tract, and also, their anti-infection effect in vivo is sometimes questionable. In this study, we therefore evaluated the potential of a mixture of four Lactobacillus species (L. salivarius, L. reuteri, L. ingluviei and L. alvi) for the protection of chickens against Salmonella Enteritidis infection. Whenever the chickens were inoculated by lactobacilli and S. Enteritidis separately, there was no protective effect of lactobacilli. This means that when lactobacilli and S. Enteritidis are exposed to each other as late as in the crop of chickens, lactobacilli did not influence chicken resistance to S. Enteritidis at all. The only positive effect was recorded when the mixture of lactobacilli and S. Enteritidis was used for the inoculation of feed and the feed was anaerobically fermented for 1 to 5 days. In this case, chickens fed such a diet remained S. Enteritidis negative. In vitro experiments showed that the protective effect was caused by acidification of feed down to pH 4.6 due to lactobacilli fermentation and was associated with S. Enteritidis inactivation. The probiotic effect of lactobacilli was thus expressed in the feed, outside the chicken host.

A neurochemical biogeography of the broiler chicken intestinal tract

The study of neurochemical-based interkingdom signaling and its impact on host-microbe interaction is called microbial endocrinology. Neurochemicals play a recognized role in determining bacterial colonization and interaction with the gut epithelium. While much attention has been devoted to the determination of neurochemical concentrations in the mammalian gut to better understand tissue and region-specific microbial endocrinology-based mechanisms of host-microbe interaction, little is known regarding the biogeography of neurochemicals in the avian gut. Greater resolution of avian gut neurochemical concentrations is needed especially as recent microbial endocrinology-based investigations into bacterial foodborne pathogen colonization of the chicken gut have demonstrated neurochemicals to affect Campylobacter jejuni and Salmonella spp. in vivo and in vitro. The aim of the present study was to determine the concentrations of stress-related neurochemicals in the tissue and luminal content of the duodenum, jejunum, ileum, cecum, and colon of the broiler intestinal tract, and to investigate if this biogeography changes with age of the bird. While all neurochemicals measured were detected in the intestinal tract, many displayed differences in regional concentrations. Whereas the catecholamine norepinephrine was detected in each region of the intestinal tract, epinephrine was present only in the cecum and colon. Likewise, dopamine, and its metabolite 3,4-dihydroxyphenylacetic acid were found in the greatest quantities in the cecum and colon. Serotonin and histamine were identified in each gut region. Region-specific age-related changes were observed (P < 0.05) for serotonin, its metabolite 5-hydroxyindole acetic acid as well as for histamine. Several neurochemicals, including norepinephrine, were found in the contents of each gut region. Epinephrine was not detected in the gut content of any region. Salsolinol, a microbial-produced neuroactive compound was detected in the gut content but not in tissue. Together, our data establish a neurochemical biogeography of the broiler chicken intestinal tract. By providing researchers with a region-by-region map of in vivo gut neurochemical concentrations of a modern broiler chicken breed, this neurochemical map is expected to inform future investigations that seek to utilize avian enteric neurochemistry.

Interdependence and biodiversity of pathogens in intestinal channel parasitocenoses of chickens in the eastern region of Ukraine

Changes in the forms of management, the concentration of poultry in limited territories lead to a violation of homeostasis in biotopes and parasitic systems. The research aimed to find out the peculiarities of distribution, biodiversity, and interdependence of pathogens in parasitocenoses of the intestinal tract of chickens in the Eastern region of Ukraine. According to the results of research, the biodiversity of pathogens in parasitocenoses of the chicken intestinal tract in poultry farms in the Eastern region of Ukraine has been determined. 17 species of pathogens were identified, including 10 species of protists (from Apicomplexa and Zoomastygophora), and 7 species of helminths: 5 species from Nematoda, 2 species from Cestoda. The influence of breeding technologies and systems of keeping chickens on the prevalence, biodiversity of parasitocenoses, species indices of parasitocenosis (SIP, %), and correlations between components of parasitocenoses have been determined. For the industrial technology of raising chickens in cages, the parasitocenosis was formed by 6 species of Eimeria (SIP — 100%, prevalence — 15.72%). When poultry was kept on a deep when kept on a deep unchanging litter in the structure of the parasitocenosis the share of eimeriosis was 87.72%, ascariasis — 12.28%. For the traditional extensive technology of raising chickens using pastures (farms and homestead farms), the prevalence in chickens was 64.07%. In the parasitocenosis of the intestinal tract in chickens there are relationships between pathogens — a high correlation between eimeriosis, histomonosis, trichomoniasis and helminthiasis (0.718–0.944) and between histomonosis, ascariasis, heterococcus and capillary (0.975–0.998), and a low correlation between histomonosis and trichomoniasis (0.449), which indicates the presence of synergetic (high correlation) and competitive (low correlation) relationships between the components of parasitocenoses of the intestinal tract

Protective effect of Lactobacillus reuteri Lb11 from chicken intestinal tract against Salmonella Enteritidis SE05 in vitro.

Salmonella infections in eggs with increasing morbidity and mortality exhibit worldwide prevalence. The present study intends to evaluate the efficacy of Lactobacillus reuteri Lb11 (L. reuteri Lb11, isolated from chicken intestinal tract) in inhibiting the growth of multi-drug resistant (MDR) Salmonella Enteritidis SE05 (obtained from egg content). The cell-free cell lysates (CFCL) of L. reuteri Lb11 obtained by the agar spot test performed well on inhibition of the MDR (Multi-DrugResistant) Salmonella Enteritidis SE05, The heat-inactivated (HI) fraction of L. reuteri Lb11 showed no inhibition activity. By co-culturing with L. reuteri Lb11 in vitro, the growth of S. Enteritidis SE05 decreased along with time, while, the pH value decreased significantly. Furthermore, In order to evaluate the mechanism of action of CFCL of L.reuteri Lb11, the genes related to the transcription level of AcrAB-TolC efflux pump, outer membrane protein OMPs genes and drug resistance genes have been quantified by real-time PCR, when the S. Enteritidis was SE05 exposed to the CFCL of L. reuteri Lb11 (1 × 1012CFU/mL). Almost all of the AcrAB-TolC efflux pump genes, outer membrane protein genes and antibiotic resistance genes were down-regulated. Especially, the level of ramA, tetA and tetB genes were down-regulated -20.77, -15.85 and -12.42 folds, respectively. L. reuteri Lb11 can effectively prevent the formation of efflux pump to inhibit the production of multidrug-resistant Salmonella Enteritidis in eggs.

The Spatial and Temporal Characterization of Gut Microbiota in Broilers.

The gut microbiota of chickens plays an important role in host physiology. However, the colonization and prevalence of gut microbiota have not been well-characterized. Here, we performed 16S rRNA gene sequencing on the duodenal, cecal and fecal microbiota of broilers at 1, 7, 21, and 35 days of age and characterized the dynamic succession of microbiota across the intestinal tract. Our results showed that Firmicutes was the most abundant phylum detected in each gut site at various ages, while the microbial diversity and composition varied among the duodenum, cecum, and feces at different ages. The microbial diversity and complexity of the cecal microbiota increased with age, gradually achieving stability at 21 days of age. As a specific genus in the cecum, Clostridium_sensu_stricto_1 accounted for 83.50% of the total abundance at 1 day of age, but its relative abundance diminished with age. Regarding the feces, the highest alpha diversity was observed at 1 day of age, significantly separated from the alpha diversity of other ages. In addition, no significant differences were observed in the alpha diversity of duodenal samples among 7, 21, and 35 days of age. The predominant bacterium, Lactobacillus, was relatively low (0.68–6.04%) in the intestinal tract of 1-day-old chicks, whereas its abundance increased substantially at 7 days of age and was higher in the duodenum and feces. Escherichia-Shigella, another predominant bacterium in the chicken intestinal tract, was also found to be highly abundant in fecal samples, and the age-associated dynamic trend coincided with that of Lactobacillus. In addition, several genera, including Blautia, Ruminiclostridium_5, Ruminococcaceae_UCG-014, and [Ruminococcus]_torques_group, which are related to the production of short-chain fatty acids, were identified as biomarker bacteria of the cecum after 21 days of age. These findings shed direct light on the temporal and spatial dynamics of intestinal microbiota and provide new opportunities for the improvement of poultry health and production.

Dual Inhibition of Salmonella enterica and Clostridium perfringens by New Probiotic Candidates Isolated from Chicken Intestinal Mucosa.

The poultry industry is the fastest-growing agricultural sector globally. With poultry meat being economical and in high demand, the end product’s safety is of importance. Globally, governments are coming together to ban the use of antibiotics as prophylaxis and for growth promotion in poultry. Salmonella and Clostridium perfringens are two leading pathogens that cause foodborne illnesses and are linked explicitly to poultry products. Furthermore, numerous outbreaks occur every year. A substitute for antibiotics is required by the industry to maintain the same productivity level and, hence, profits. We aimed to isolate and identify potential probiotic strains from the ceca mucosa of the chicken intestinal tract with bacteriocinogenic properties. We were able to isolate multiple and diverse strains, including a new uncultured bacterium, with inhibitory activity against Salmonella Typhimurium ATCC 14028, Salmonella Abony NCTC 6017, Salmonella Choleraesuis ATCC 10708, Clostridium perfringens ATCC 13124, and Escherichia coli ATCC 25922. The five most potent strains were further characterized for their probiotic potential (i.e., sensitivity to antibiotics and tolerance to gastrointestinal physicochemical conditions). Our analyzed lactobacilli strains exhibited some interesting probiotic features while being inhibitory against targeted pathogens.

Immune response and protective efficacy of recombinant Enterococcus faecalis displaying dendritic cell–targeting peptide fused with Eimeria tenella 3-1E protein

Avian coccidiosis causes significant economic losses on the global poultry breeding industry. Exploration of new-concept vaccines against coccidiosis has gradually become a research hotspot. In this study, an Enterococcus faecalis strain (MDXEF-1) showing excellent performance isolated from chicken intestinal tract was used as a vector to deliver Eimeria target protein. The plasmid pTX8048-SP-DCpep-NAΔ3-1E-CWA harboring dendritic cell–targeting peptide (DCpep) fusion with Eimeria tenella NAΔ3-1E gene (3-1E protein–coding gene without start codon ATG and terminator codon TAA) was electrotransformed into MDXEF-1 to generate the recombinant bacteria MDXEF-1/pTX8048-SP-DCpep-NAΔ3-1E-CWA in which NAΔ3-1E protein was covalently anchored to the surface of bacteria cells by cell wall anchor (CWA) sequence. The expression of target fusion protein DCpep-NAΔ3-1E-CWA was detected by Western blot. Each chicken was immunized 3 times at 2-wk intervals with live E. faecalis expressing DCpep-NAΔ3-1E fusion protein (DCpep-NAΔ3-1E group), live E. faecalis expressing NAΔ3-1E protein (NAΔ3-1E group), and live E. faecalis containing empty vector only. The 3 immunized groups were then challenged with homologous E. tenella sporulated oocyst after immunizations, and the immune response and protective efficacy in each group were evaluated. The results showed that serum IgG levels, secretory IgA levels in cecal lavage, proportion of CD4+ and CD8α+ cells in peripheral blood, and mRNA expression levels of IL-2 and IFN-γ in the spleen were significantly higher in chickens in the DCpep-NAΔ3-1E group than in chickens of the NAΔ3-1E group (P < 0.05). Oral immunization to chickens with live E. faecalis expressing DCpep-NAΔ3-1E offered more protective efficacy against homologous challenge including significant improved body weight gain, increased oocyst decrease ratio, and reduced average lesion scores in cecum compared with chickens with live E. faecalis expressing NAΔ3-1E protein. These results suggest that recombinant E. faecalis expressing dendritic cell–targeting peptide fusion with E. tenella 3-1E protein could be a potential approach for prevention of Eimeria infection.

Research Note: Lyophilization of hyperimmune egg yolk: effect on antibody titer and protection of broilers against Campylobacter colonization

Oral administration of antibodies is a promising strategy against various infectious diseases. Previously, it was demonstrated that passive immunization by providing hyperimmune egg yolk through the feed reduces Campylobacter jejuni colonization in broilers. Campylobacteriosis is the most commonly reported bacterial foodborne zoonosis worldwide, and poultry products are the number one origin of these bacteria for human infection. To date, no effective control measures exist to limit Campylobacter colonization in the chicken's intestinal tract. Here, the effect of lyophilization of hyperimmune egg yolk on protection of broilers against C. jejuni was investigated. During an in vivo trial, broiler chickens were prophylactically given feed with lyophilized hyperimmune or non-immunized egg yolk powder starting from day 1 after hatch. At day 11, broilers were inoculated with C. jejuni according to a seeder model. Five days later, all broilers were euthanized and cecal content was examined for C. jejuni colonization. No decrease in C. jejuni colonization was found. The freeze-drying resulted in a 16-fold decrease of the antibody titer in the yolk powder compared to the fresh yolks, presumably caused by structural changes in the antibodies. In conclusion, applying freeze-dried hyperimmune egg yolk failed to protect broilers against C. jejuni colonization, possibly because lyophilization affected the antibodies' functionality.