Complete genome sequences of two avian pathogenic Escherichia coli strains isolated from broilers exhibiting colibacillosis in Mississippi.

We report the whole-genome sequences of Escherichia coli strains APEC-O2-MS1266 and APEC-O2-MS1657 isolated from the liver and heart of infected broilers in Mississippi State, US. The genomic information of these two causative strains may provide a valuable reference for comparative studies of avian pathogenic E. coli.

Phenotypic virulence characterization of avian pathogenic Escherichia coli (APEC) isolates from broiler breeders with colibacillosis in Mississippi.

AIMS: In this study, we evaluated the phenotypic virulence characteristics of avian pathogenic Escherichia coli (APEC) isolates from broiler breeders with colibacillosis in Mississippi. Also, the relationship between phenotypic and genotypic virulence patterns was determined. METHODS AND RESULTS: Twenty-eight APEC isolated from lesions of broiler breeders diagnosed with colibacillosis were used for embryo lethality assay and chick challenge study. The percentage of embryo mortality following embryo lethality assay and pathogenicity score following the chick challenge study were used to categorize the isolates based on virulence. Pearson correlation analysis was performed to determine the relationship between embryo mortality, chick pathogenicity, and the presence of virulence-associated genes in the isolates. Overall, 39.3% of the isolates were highly virulent and 3.5% were avirulent, following both assays. There existed a positive correlation between embryo mortality and chick pathogenicity (r = 0.73, P < .01), as well as percentage embryo mortality and pathogenicity score with the presence of some virulence genes. CONCLUSIONS: Even though all the APEC were isolated from lesions of diseased breeders, the virulence potential varied from being avirulent to highly virulent. Further, we identified a positive relationship between phenotypic virulence and the frequency of virulence-associated genes.

Changes in gene expression in the intestinal mucus of broilers with woody breast myopathy.

Previous work has shown that dietary treatments affect woody breast (WB) incidence differently, which indicates that gut conditions such as gut barrier function, inflammation, and oxidative stress are likely related to WB. In this study, dietary supplementation with antibiotics (bacitracin) or probiotics (Bacillus subtilis) was investigated for their effects on the expression of transcripts related to gut barrier function, inflammation, and oxidative stress in the mucus lining of the jejunum from broilers with or without WB. A split-plot experimental design was used in this study. The dietary treatments served as the main plot factor and the breast muscle condition was the subplot factor. On d 41, jejunum mucus was collected from 1 bird from each of 3 replicate pens in each 3 dietary treatment groups that exhibited WB and an additional bird that contained a normal breast (3 biological replicates/treatment/phenotype; 3 × 3 × 2, total N = 18). Total RNA was extracted using a commercial RNA extraction kit. The expression levels of CLDN1, MUC6, TLR2A, TLR2B, TLR4, IFN-γ, IL-1ß, IL-8L1, IL-10, NOS2, and SOD were determined using 2-step RT-qPCR analysis. The gene expression difference in ΔCt values was determined after normalizing with the chicken 18S rRNA gene. When the significant differences occurred between treatments, the relative fold change was calculated using the ΔΔCt method and the significance level was calculated. The PROC GLM procedure of SAS 9.4 was used, and the level of significance was set at P ≤ 0.05. There were no significant interactive effects between diet and the breast muscle condition on the expression of any of the genes tested. However, birds with WB exhibited higher MUC6 (P < 0.0001) gene expression levels than birds with normal breast muscles. In addition, the expression of SOD decreased in birds that were fed the antibiotic diet when compared to birds that were fed the probiotic diet (P = 0.014). In conclusion, WB identified in broilers tested in the current study is attributed to increased expression of mucin, indicating a correlation between WB incidence and gel-forming mucin secretion and pathogen signaling.

Application of Engineered Dendritic Cell Vaccines in Cancer Immunotherapy: Challenges and Opportunities.

OPINION STATEMENT: The primary objective of this study is to evaluate the effectiveness of cancer vaccines containing genetically modified dendritic cells (DCs) in inducing transformational immune responses. This paper sheds considerable light on DCs' function in advancing treatment techniques. This objective is achieved by thoroughly analyzing the many facets of DCs and their strategic integration into cancer treatment. Due to their role as immune response regulators, DCs can potentially enhance cancer treatment strategies. DCs have the potential to revolutionize immunotherapy, as shown by a comprehensive analysis of their numerous characteristics. The review deftly transitions from examining the fundamentals of preclinical research to delving into the complexities of clinical implementation while acknowledging the inherent challenges in translating DC vaccine concepts into tangible progress. The analysis also emphasizes the potential synergistic outcomes that can be achieved by combining DC vaccines with established pharmaceuticals, thereby emphasizing the importance of employing a holistic approach to enhance treatment efficacy. Despite the existence of transformative opportunities, advancement is hindered by several obstacles. The exhaustive analysis of technical complexities, regulatory dynamics, and upcoming challenges provides valuable insights for overcoming obstacles requiring strategic navigation to incorporate DC vaccines successfully. This document provides a comprehensive analysis of the developments in DC-based immunotherapy, concentrating on its potential to transform cancer therapy radically.

In silico prediction and expression analysis of vaccine candidate genes of Campylobacter jejuni.

Campylobacter jejuni (C. jejuni) is the most common food-borne pathogen that causes human gastroenteritis in the United States. Consumption of contaminated poultry products is considered as the major source of human Campylobacter infection. An effective vaccine would be a promising alternative to antibiotic supplements to curb C. jejuni colonization in poultry gastrointestinal (GI) tract. However, the genetic diversity among the C. jejuni isolates makes vaccine production more challenging. Despite many attempts, an effective Campylobacter vaccine is not yet available. This study aimed to identify suitable candidates to develop a subunit vaccine against C. jejuni, which could reduce colonization in the GI tract of the poultry. In the current study, 4 C. jejuni strains were isolated from retail chicken meat and poultry litter samples and their genomes were sequenced utilizing next-generation sequencing technology. The genomic sequences of C. jejuni strains were screened to identify potential antigens utilizing the reverse vaccinology approach. In silico genome analysis predicted 3 conserved potential vaccine candidates (phospholipase A [PldA], TonB dependent vitamin B12 transporter [BtuB], and cytolethal distending toxin subunit B [CdtB]) suitable for the development of a vaccine. Furthermore, the expression of predicted genes during host-pathogen interaction was analyzed by an infection study using an avian macrophage-like immortalized cell line (HD11). The HD11 was infected with C. jejuni strains, and the RT-qPCR assay was performed to determine the expression of the predicted genes. The expression difference was analyzed using ΔΔCt methods. The results indicate that all 3 predicted genes, PldA, BtuB, and CdtB, were upregulated in 4 tested C. jejuni strains irrespective of their sources of isolation. In conclusion, in silico prediction and gene expression analysis during host-pathogen interactions identified 3 potential vaccine candidates for C. jejuni.

A compilation of virulence-associated genes that are frequently reported in avian pathogenic Escherichia coli (APEC) compared to other E. coli.

Escherichia coli survive in various hosts and environments due to their highly diversified genome. These bacteria have coevolved with humans, colonized a broad range of hosts, and survive as a commensal organism or pathogen. Escherichia coli that adopted a pathogenic lifecycle in avian hosts typically belong to phylogroups B2 and D. Phylogenic investigations discovered these E. coli are noticeably overlapped with the phylogroup of E. coli infecting humans. This overlapping is possibly due to a parallel evolution in both hosts from a common ancestor, which indicates a high zoonotic potential of avian pathogenic E. coli (APEC). However, some contrasting evidence of other phylogroups infecting the avian host has also been reported in recent studies indicating phylogroups of E. coli are not definitive, only suggestive to their virulence in chickens. Furthermore, virulence-associated genes that contribute to bacterial features necessary to establish APEC infection, are predominantly located in plasmids. Therefore, phylogenetic classification based on chromosomal markers is often inadequate to identify APEC. Moreover, E. coli can obtain virulent plasmids from other bacteria, which further complicates the link between phylogenetic classification and pathotype. Previous research has reported an array of virulence-associated genes highly prevalent only in APEC isolates. Function of these genes are possibly a prerequisite to establishing APEC infections in chickens. Consequently, these genes can be used to distinguish APEC from environmental, commensal, intestinal, and other extraintestinal E. coli. Therefore, we have extensively reviewed previous literature to compile the virulence-associated genes that are highly prevalent in APEC compared to other E. coli. From this review, we have identified 10 key virulence-associated genes (iss,tsh,iroN, episomal/chromosomal ompT,iutA,cvaC,hlyF,iucD,papG allel(II/III), and papC) that are frequently reported in APEC isolates than nonpathogenic E. coli. A compilation of these research findings can be crucial to the molecular identification of APEC. Furthermore, it can serve as a guideline for future investigation and aid in formulation of intervention strategies.

Internal organ and skeletal muscle development in commercial broilers with woody breast myopathy.

Increasing growth rate, body weight, and breast muscle yield have been linked to broiler muscle problems such as woody breast (WB). The aim of this study was to investigate the internal organ and skeletal muscle development of broilers with WB myopathy under dietary and Eimeria challenge treatments. A 3 diet (control, antibiotic, or probiotic) × 2 challenge (control or Eimeria) × 2 sex factorial arrangement of treatments was used in a randomized complete block design. Ross × Ross 708 chicks were randomly assigned to 96 floor pens with 12 treatment combinations (8 replicates per treatment). Internal organs were sampled on d 13 and 41. Skeletal muscles were sampled on d 41. Internal organ and skeletal muscle weights were analyzed using a 3-way analysis of variance (ANOVA). Relationships between WB and internal organ and skeletal muscle weights were analyzed using one-way ANOVA as all treatments were pooled together and regrouped according to WB scores. On d 41, absolute and relative heart weights were greater in males when they were averaged over diet and challenge treatments (P < 0.001 and P = 0.026, respectively). The birds with WB score 3 had greater absolute heart (P = 0.0002) and spleen weights (P = 0.016), but there was no difference in relative spleen weight (P > 0.05). When averaged over diet and challenge treatments, males have greater absolute duodenum, jejunum, and ileum weights (for all P < 0.0001). Compared with birds with normal breasts, the birds with WB scores 1, 2, and 3 had a greater live weight (for all P < 0.0001) and absolute and relative breast weights (for all P < 0.0001). The birds with WB score 1, 2, and 3 had greater (P < 0.0001) absolute but lower (P < 0.0001) relative drumstick, thigh, and wing weights. Results indicated that broilers with WB had lower relative proventriculus and gizzard weights and greater relative breast meat weight with lower relative drumstick, thigh, and wing muscle weights.

Effects of dietary bacitracin or Bacillus subtilis on the woody breast myopathy-associated gut microbiome of Eimeria spp. challenged and unchallenged broilers.

Study suggested that dysbiosis of the gut microbiota may affect the etiology of woody breast (WB). In the current study, the cecal microbiota and WB in chickens fed three different diets were investigated. A total of 504 male chicks were used in a randomized complete block design with a 3 (Diet) × 2 (Challenge) factorial arrangement of treatments with 6 replicates per treatment, 6 treatments per block, and 14 birds per treatment. The experimental diets were a control diet (corn-soybean meal basal diet), an antibiotic diet (basal diet + 6.075 mg bacitracin/kg feed), and a probiotic diet (basal diet + 2.2 × 108 CFU Bacillus subtilis PB6/kg feed). On d 14, birds that were assigned to the challenge treatment received a 20 × live cocci vaccine. On d 41, breast muscle hardness in live birds was palpated and grouped into normal (NB) and WB phenotypes. Cecal contents were collected and their bacterial compositions were analyzed and compared. The genomic DNA of the cecal contents was extracted and the V3 and V4 regions of 16S rRNA gene were amplified and sequenced via an Illumina MiSeq platform. There were no differences (P > 0.05) in Shannon and Chao 1 indexes between the challenges, diets, and phenotypes (NB vs. WB). However, there was a difference (P = 0.001) in the beta diversity of the samples between the challenged and nonchallenged groups. Relative bacterial abundance differed (false discovery rate, FDR < 0.05) between the challenge treatments, but there were no significant differences (FDR > 0.05) among the three diets or two phenotypes. Predicted energy metabolism, nucleotide metabolism, and amino acid and coenzyme biosynthesis activities only differed (q-value < 0.05) between challenged and nonchallenged groups. The cocci challenge altered the gut microbial composition on Butyricicoccus pullicaecorum, Sporobacter termitidis, and Subdoligranulum variabile, but the dietary antibiotic and probiotic treatments did not impact gut microbial composition. No strong association was found between WB myopathy and gut microbial composition in this study.

Effects of ground robot manipulation on hen floor egg reduction, production performance, stress response, bone quality, and behavior.

Reducing floor eggs in cage-free (CF) housing systems is among primary concerns for egg producers. The objective of this research was to evaluate the effects of ground robot manipulation on reduction of floor eggs. In addition, the effects of ground robot manipulation on production performance, stress response, bone quality, and behavior were also investigated. Two successive flocks of 180 Hy-Line Brown hens at 34 weeks of this age were used. The treatment structure for each flock consisted of six pens with three treatments (without robot running, with one-week robot running, and with two-weeks robot running), resulting in two replicates per treatment per flock and four replicates per treatment with two flocks. Two phases were involved with each flock. Phase 1 (weeks 35-38) mimicked the normal scenario, and phase 2 (weeks 40-43) mimicked a scenario after inadvertent restriction to nest box access. Results indicate that the floor egg reduction rate in the first two weeks of phase 1 was 11.0% without the robot treatment, 18.9% with the one-week robot treatment, and 34.0% with the two-week robot treatment. The effect of robot operation on floor egg production was not significant when the two phases of data were included in the analysis. Other tested parameters were similar among the treatments, including hen-day egg production, feed intake, feed conversion ratio, live body weight, plasma corticosterone concentration, bone breaking force, ash percentage, and time spent in nest boxes. In conclusion, ground robot operation in CF settings may help to reduce floor egg production to a certain degree for a short period right after being introduced. Additionally, robot operation does not seem to negatively affect hen production performance and well-being.

Bacitracin, Bacillus subtilis, and Eimeria spp. challenge exacerbates woody breast incidence and severity in broilers.

Woody breast (WB) is a myopathy that is related to the increasing growth rate. Understanding the influence of management factors on WB formation and development is important to minimize WB. This study was conducted to define how management factors affect broiler growth performance, processing yield, and WB incidence. Ross × Ross 708 chicks were randomly assigned to a 3 (diet) × 2 (cocci challenge) × 2 (sex) factorial arrangement of treatments. The 3 dietary treatments were: control diet (corn-soybean meal basal diet), antibiotic diet (basal diet + 6.075 mg bacitracin /kg feed), and probiotic diet (basal diet + 2.2 × 108 CFU Bacillus subtilis PB6/kg feed). Birds in cocci challenge treatments received 20 × live cocci vaccine on d 14. The hardness of breast muscle in live birds was determined by palpation and grouped into Normal, Slight, Moderate, and Severe categories. Across diet and sex treatments, the cocci challenge resulted in decreases in body weight (BW) on d 29 and 35 (P < 0.0001 and = 0.032) in body weight gain (BWG) from d 14 to 29 (P < 0.0001). However, an increase of BW occurred on d 35 (P = 0.032) and an increase of BWG occurred from d 29 to 35 and d 35 to 43 (P = 0.0001 and 0.002), and the cocci challenge increased WB incidence on d 29 (P = 0.043) and d 43 (P = 0.013). Across challenge and sex treatments, birds fed the antibiotic diet exhibited a higher growth rate (GR) than those fed the control or probiotic diet from d 0 to 14 (P = 0.016), but not after d 14 (P > 0.05). Across sex, the antibiotic and probiotic diets increased WB incidence for those birds that did not receive a cocci challenge on d 43 (P = 0.040). Across challenge and diet treatments, males exhibited a higher BW, BWG, and GR throughout all growth phases, and males showed a higher WB incidence on d 29, 35, and 43 (P = 0.002, P < 0.0001, and P = 0.0002, respectively). In conclusion, bacitracin and Eimeria spp. increased WB incidence, BW, and GR. However, Bacillus subtilis increased WB incidence in male broilers without affecting BW and GR.

Synthesis of Quinazolin-4(3 H)-ones via the Reaction of 2-Halobenzamides with Nitriles.

This paper describes a convenient method to synthesize quinazolin-4(3 H)-ones from simple and readily available 2-halobenzamides and nitriles. The Lewis acid Cu-catalyzed nucleophilic addition of 2-halobenzamide to nitriles followed by SNAr reaction proceeds smoothly in the presence of tBuOK as a base to produce quinazolinone derivatives.

Flocculation kinetics mechanism and floc formation prepared by poly aluminum chloride coupled with polyacrylamide for ship ballast water.

The performance of flocculants prepared by poly aluminum chloride (PAC) and polyacrylamide (PAM) on treating ballast water collected at the Dalian new port area, the evaluation depending on the values of reaction parameters, and kinetics mechanism of flocculation were investigated in this study. Accordingly, the flocculants of 0.1 g·L(-1), prepared by mixing PAC of 10% with PAM of 2.0‰, enabled the removal rate of zooplankton and phytoplankton to reach 91% in ballast water at 20 °C. Based on flocculation kinetics mechanism analysis, the efficient vortex size during stirring should be larger than the floc particles, and gradient of fluctuating velocity provide the impetus for turbulence flocculation. The results of this study could be relevant to understanding particle-floc interactions during developmental flocculation, and during application of ballast water treatment.