Bacterial Challenge Research Articles

Rhodobacter sphaeroides supplementation improves defense ability of Chinese mitten crab Eriocheir sinensis against Shewanella putrefaciens infection via intestinal flora and metabolism regulation

Shewanella putrefaciens is a vital bacterial pathogen implicated in serious diseases in Chinese mitten crab Eriocheir sinensis. Yet the use of probiotics to improve the defense ability of E. sinensis against S. putrefaciens infection remains poorly understood. In the present study, the protective effect of dietary R. sphaeroides against S. putrefaciens infection in E. sinensis was evaluated through antioxidant capability, immune response, and survival under bacterial challenge assays, and its protective mechanism was further explored using a combination of intestinal flora and metabolome assays. Our results indicated that dietary R. sphaeroides could significantly improve immunity and antioxidant ability of Chinese mitten crabs, thereby strengthening their disease resistance with the relative percentage survival of 81.09% against S. putrefaciens. In addition, dietary R. sphaeroides could significantly alter the intestinal microbial composition and intestinal metabolism of crabs, causing not only the reduction of potential threatening pathogen load but also the increase of differential metabolites in tryptophan metabolism, pyrimidine metabolism, and glycerophospholipid metabolism. Furthermore, the regulation of differential metabolites such as N-Acetylserotonin positively correlated with beneficial Rhodobacter could be a potential protection strategy for Shewanella infection. To the best of our knowledge, this is the first study to illustrate the protective effect and mechanism of R. sphaeroides supplementation to protect E. sinensis against S. putrefaciens infection.

Identification and florfenicol-treatment of pseudomonas putida infection in gilthead seabream (Sparus aurata) fed on tilapia-trash-feed

The present study aimed to determine the major cause of the high mortality affecting farmed gilthead seabream (Sparus aurata) and controlling this disease condition. Fifteen diseased S. aurata were sampled from a private fish farm located at Eldeba Triangle, Damietta, fish showed external skin hemorrhages, and ulceration. Bacterial isolates retrieved from the diseased fish were identified biochemically as Pseudomonas putida and then confirmed by phylogenetic analysis of the 16 S rRNA gene sequence. P. putida was also isolated from three batches of tilapia-trash feed given to S. aurata. Biofilm and hemolytic assay indicated that all P. putida isolates produced biofilm, but 61.11% can haemolyse red blood cells. Based on the antibiotic susceptibility test results, P. putida was sensitive to florfenicol with minimum inhibitory concentrations ranging between 0.25 and 1.0 µg mL− 1, but all isolates were resistant to ampicillin and sulfamethoxazole-trimethoprim. Pathogenicity test revealed that P. putida isolate (recovered from the tilapia-trash feed) was virulent for S. aurata with LD50 equal to 4.67 × 107 colony forming unit (CFU) fish− 1. After intraperitoneal (IP) challenge, fish treated with 10 mg kg− 1 of florfenicol showed 16.7% mortality, while no mortality was recorded for the fish group that received 20 mg kg− 1. The non-treated fish group showed 46.7% mortality after bacterial challenge. HPLC analysis of serum florfenicol levels reached 1.07 and 2.52 µg mL− 1 at the 5th -day post-drug administration in the fish groups received 10 and 20 mg kg− 1, respectively. In conclusion, P. putida was responsible for the high mortality affecting cultured S. aurata, in-feed administration of florfenicol (20 mg kg− 1) effectively protected the challenged fish.

Activation of immune pathways in common bed bugs, Cimex lectularius, in response to bacterial immune challenges - a transcriptomics analysis.

The common bed bug, Cimex lectularius, is an urban pest of global health significance, severely affecting the physical and mental health of humans. In contrast to most other blood-feeding arthropods, bed bugs are not major vectors of pathogens, but the underlying mechanisms for this phenomenon are largely unexplored. Here, we present the first transcriptomics study of bed bugs in response to immune challenges. To study transcriptional variations in bed bugs following ingestion of bacteria, we extracted and processed mRNA from body tissues of adult male bed bugs after ingestion of sterile blood or blood containing the Gram-positive (Gr+) bacterium Bacillus subtilis or the Gram-negative (Gr-) bacterium Escherichia coli. We analyzed mRNA from the bed bugs' midgut (the primary tissue involved in blood ingestion) and from the rest of their bodies (RoB; body minus head and midgut tissues). We show that the midgut exhibits a stronger immune response to ingestion of bacteria than the RoB, as indicated by the expression of genes encoding antimicrobial peptides (AMPs). Both the Toll and Imd signaling pathways, associated with immune responses, were highly activated by the ingestion of bacteria. Bacterial infection in bed bugs further provides evidence for metabolic reconfiguration and resource allocation in the bed bugs' midgut and RoB to promote production of AMPs. Our data suggest that infection with particular pathogens in bed bugs may be associated with altered metabolic pathways within the midgut and RoB that favors immune responses. We further show that multiple established cellular immune responses are preserved and are activated by the presence of specific pathogens. Our study provides a greater understanding of nuances in the immune responses of bed bugs towards pathogens that ultimately might contribute to novel bed bug control tactics.

Functional characterization of NOD1 from golden pompano Trachinotus ovatus

Fish rely on innate immune system for immunity, and nucleotide-binding oligomerization domain-like receptors (NLRs) are a vital group of receptor for recognition. In the present study, NOD1 gene was cloned and characterized from golden pompano Trachinotus ovatus, a commercially important aquaculture fish species. The ORF of T. ovatus NOD1 was 2820 bp long, encoding 939 amino acid residues with a highly conserved domains containing CARD-NACHT-LRRs. Phylogenetic analysis revealed that the T. ovatus NOD1 clustered with those of fish and separated from those of birds and mammals. T. ovatus NOD1 has wide tissue distribution with the highest expression in gills. Bacterial challenges (Streptococcus agalactiae and Vibrio alginolyticus) significantly up-regulated the expression of NOD1 with different response time. The results of T. ovatus NOD1 ligand recognition and signaling pathway analysis revealed that T. ovatus NOD1 could recognize iE-DAP at the concentration of ≧ 100 ng/mL and able to activate NF-κB signaling pathway. This study confirmed that NOD1 play a crucial role in the innate immunity of T. ovatus. The findings of this study improve our understanding on the immune function of NOD1 in teleost, especially T. ovatus.

Dietary supplementation of methionine, lysine, and tryptophan as possible modulators of growth, immune response, and disease resistance in striped catfish (Pangasius hypophthalmus).

The present study investigated the potential role of different essential amino acids (AA) in striped catfish (Pangasius hypophthalmus). Fish (initial weight = 17.91±0.27 g, n = 260) were fed with eight isonitrogenous (30%), and isolipidic diets (6%) formulated to include different combinations of tryptophan (Trp), methionine (Met), and lysine (Lys) (T0: Zero AA, T1: Trp, T2: Lys, T3: Met, T4: Trp+Met, T5: Lys+Trp, T6: Met+Lys, T7: Lys+Trp+Met) for eight weeks. The dose of amino acid supplementation, whether individually or in combination, was 5g of each amino acid per kg of diet. The trial comprised eight treatments, with each treatment consisted of three replicates (n = 10/replicate). At the end of the growth experiment, the highest total body weight, crude protein, digestive enzymatic activity, immune response, and amino acids level were observed in treatments supplemented with amino acids compared to T0. After the growth experiment, fish in all treatments were exposed to Staphylococcus aureus (5×105 CFU/ml). For bacterial challenge trial, the T0 treatment was designated as positive (+ve T0) and negative control (-ve T0). Following the S. aureus challenge, fish fed with amino acids showed a better response to reactive oxygen species and lipid peroxidation, as indicated by the increased levels of catalase and superoxide dismutase. Conversely, the concentration of malondialdehyde gradually decreased in all treatments compared to the +ve T0 treatment. It is concluded that supplementation of amino acids improved the growth, protein content, and immunocompetency against S. aureus in striped catfish. The most favorable outcomes in striped catfish were shown by fish supplemented with T7 diet. These essential amino acids hold potential as efficient supplements for use in the intensive aquaculture for striped catfish.

Global warming-related response after bacterial challenge in Astroides calycularis, a Mediterranean thermophilic coral

A worldwide increase in the prevalence of coral diseases and mortality has been linked to ocean warming due to changes in coral-associated bacterial communities, pathogen virulence, and immune system function. In the Mediterranean basin, the worrying upward temperature trend has already caused recurrent mass mortality events in recent decades. To evaluate how elevated seawater temperatures affect the immune response of a thermophilic coral species, colonies of Astroides calycularis were exposed to environmental (23 °C) or elevated (28 °C) temperatures, and subsequently challenged with bacterial lipopolysaccharides (LPS). Using immunolabeling with specific antibodies, we detected the production of Toll-like receptor 4 (TLR4) and nuclear factor kappa B (NF-kB), molecules involved in coral immune responses, and heat shock protein 70 (HSP70) activity, involved in general responses to thermal stress. A histological approach allowed us to characterize the tissue sites of activation (epithelium and/or gastroderm) under different experimental conditions. The activity patterns of the examined markers after 6 h of LPS stimulation revealed an up-modulation at environmental temperature. Under warmer conditions plus LPS-challenge, TLR4-NF-kB activation was almost completely suppressed, while constituent elevated values were recorded under thermal stress only. An HSP70 up-regulation appeared in both treatments at elevated temperature, with a significantly higher activation in LPS-challenge colonies. Such an approach is useful for further understanding the molecular pathogen-defense mechanisms in corals in order to disentangle the complex interactive effects on the health of these ecologically relevant organisms related to global climate change.

Effects of supplementation with different zinc-based products on the growth and health of Nile tilapia

Zinc is one of the essential microelements for the metabolism of animals. Zinc nanoparticles may have higher bioavailability due to their low specific surface area, facilitating absorption by fish. The present study aimed to evaluate the effects of supplementation with different zinc-based products on the growth and health of Nile tilapia Oreochromis niloticus. Zinc, in different sizes (nanoparticles or bulk) and forms (inorganic or organic), were used as a supplement in the tilapia diet at a dose of 15 mg kg feed−1 for 60 days. At the end of the feeding trial, production performance, hemato-immunological parameters, activity of antioxidant system enzymes, exposure to Streptococcus agalactiae and zinc concentration in the muscle were examined. After the bacterial challenge, the mean corpuscular hemoglobin concentration (MCHC) significantly increased in the fish treated with organic zinc, inorganic nano zinc, and organic nano zinc, while in the control group (inorganic zinc), MCHC remained unchanged. Regarding defense cells, dietary inorganic nano zinc increased the number of basophils (1.50 ± 1.10) compared to organic zinc (0.80 ± 0.90). Lymphocyte count increased after the challenge only in the organic zinc treatments (bulk and nanoparticles). Neutrophils decreased in the control (inorganic zinc) (2.20 ± 1.70) and inorganic nano zinc (2.60 ± 2.70) treatments after the challenge. When compared before and after the bacterial challenge, the plasma antimicrobial titer significantly increased after the bacterial challenge in all treatments. No significant differences were observed for total proteins, enzymes (SOD and CAT), cumulative survival and zinc deposition on fillet. In conclusion, organic zinc in nanoparticles or bulk size increased Nile tilapia innate defense during bacterial infection. However, the other parameters evaluated were not affected by zinc particle size or form (organic or inorganic), indicating that further evaluations should be conducted with organic zinc in nanoparticles or bulk size in the tilapia diet.

Tarragon (Artemisia dracunculus) Essential Oil at Optimized Dietary Levels Prompted Growth, Immunity, and Resistance to Enteric Red-Mouth Disease in the Rainbow Trout (Oncorhynchus mykiss)

Fingerlings of the rainbow trout, Oncorhynchus mykiss (n = 300, 10.63 ± 0.6 g), were fed tarragon (Artemisia dracunculus) essential oil (TGO) for 2 months to examine its effects on growth properties, immunity, and resistance to Yersinia ruckeri infection. The treatments were control or TG1, TG2 (fed 0.5% TGO), TG3 (1% TGO), and TG4 (2% TGO). According to the results, an improvement was observed in growth parameters in all TGO-treated groups compared to the control (P<0.05). The digestive enzyme activities (protease and lipase) were significantly elevated in response to dietary TGO (P<0.05). The immune system of the fish was enhanced by TGO, as it stimulated the immune parameters in serum (lysozyme, myeloperoxidase (MPO), alternative complement (ACH50), Ig) and mucus (lysozyme, protease, ACH50, Ig) (P<0.05). The treatments, TG3 and TG4, showed more immune performance in response to TGO (P<0.05). The fish in TG2 treatment had a higher levels of serum total protein than other groups (P<0.05). The concentration of triglycerides (TRIG) and cholesterol (CHOL) in serum significantly decreased (P<0.05) in response to TGO, as the lowest levels were observed in the treatment, TG3. The antioxidant enzymes (superoxide dismutase (SOD) and catalase (CAT)) of serum elevated in TGO-treated fish, with the maximum values for the TG4 group (P<0.05). TGO reduced (P<0.05) alanine aminotransferase (ALT) and alkaline phosphatase (ALP) levels in serum. After bacterial challenge, the TGO-treated fish showed lower mortality compared to the control, where the lowest mortality was observed in TG4 (P<0.05). In conclusion, TGO improved growth, immunity, and survival after bacterial challenge in the rainbow trout, with more performance in fish fed 1%–2% TGO.

A novel ABC family protein participates in the regulation of fitness cost caused by tet(X4)-bearing plasmids in Escherichia coli

The worldwide prevalence of the plasmid-mediated tigecycline resistance gene tet(X4) in diverse bacterial species challenges the clinical application of tigecycline and poses an increasing threat to global health. However, the underlying mechanisms by which tet(X4)-positive plasmids adapt to different host bacteria and maintain resistance genes remain elusive. Herein, we performed a systematic investigation of the fitness costs of tet(X4)-bearing plasmids in different host strains. We found that the combination of tet(X4) gene and Escherichia coli (E. coli) showed a minimal fitness cost. Meanwhile, E. coli harboring tet(X4)-bearing IncFII plasmid pC41 displayed high lethality in a Galleria mellonella (G. mellonella) infection model, superior conjugation frequency and great competitiveness in murine models. Furthermore, we investigated the regulatory mechanisms for controlling the fitness cost of tet(X4)-bearing pC41 in E. coli. A putative ABC transport family protein, designated as AtlR, balanced the tet(X4) expression and bacterial fitness cost by suppressing the plasmid copy number. It inhibited the plasmid replication by downregulating the expression of repA. Overall, these findings uncover a novel adaptative mechanism accounting for the persistence of tet(X4)-bearing plasmids.

Biological control of Pseudomonas aeruginosa (ATCC-27853) by isolated lytic bacteriophage

Background Pseudomonas aeruginosa is a common bacterial species; it can grow in all fields, even in distilled water. One of the obvious characteristics of P. aeruginosa species is its low antibiotic susceptibility (antibiotic-multi-resistant species); this reflects the urgent need for alternative strategies. Unconventionally, we try to use an alternative destructive antibacterial agent, phage. phage application in our environmental fields (engineering relevance) is a promising safe avenue for replacing chemical biocides. Objective To isolate a lytic phage as a bio-control agent for Pseudomonas aeruginosa (ATCC-27853), evaluate the ability of the isolated phage to inhibit or suppress the growth of the target bacteria. We study phage stability while performing biological and physical characterization, optimizing phage action against bacteria. Materials and methods We use an activated, purified young culture of Pseudomonas aeruginosa (ATCC-27853) as the target bacteria to isolate a specific lytic bacteriophage from a mixture of Enriched sewage and marine water through a sequence of experiments: a Broth clearing assay using nutrient broth and a spot assay test by using nutrient agar medium, both for the detection of phage presence and its lytic action. For the purification of phage, the plaque assay test was performed in many successive subcultures by using the serial dilution method on a semisolid medium. The high-titer lysate obtained from the confluent plates was investigated by TEM to determine its morphology and taxonomy. We performed biological characterization of the isolated phage: one-step growth curve, Phage adsorption rate, host range (using nutrient double layer agar), as well as a bacterial challenge test (using nutrient broth, based on the optical densities). Also, we performed physical characterization of the phage to determine phage Stability, pH, thermal stability, and the influence of UV and organic solvents on the lytic action of the phage. Results and conclusion Pyocyanin and pyoverdin pigments (both fluorescent under UV illumination) are secreted by P. aeruginosa (ATCC-27853), also their ability to thrive at 42°C. After spotting P. aeruginosa plates with enriched sewage filtrate, the results indicated lytic area (+ve). Plaques have a pin tip and are translucent. The lytic profile of bacteria after phage infection revealed that the bacteria was reduced to a MOI of 10 in 4 h. However, the multiplicity of infection(MOI) of 1 and 0.01, respectively, was longer (10, 22/h). The electron microscopy revealed that the phage is a member of the podoviridae family, with a noncontractile short tail (18 nm) and a polyhedral (heptagonal) head (45 nm) in diameter and a phage length of 63 nm. The adsorption rate of the phage was greatest at 20 min. Our goal with the phage application is to replace harmful chemical biocides with lytic bacteriophages in various environmental systems (Phage for Biological Control). Phage bio-control treatment thus holds promise for eliminating the requirement for hazardous chemical biocides.

Isolation and characterization of a relatively broad-spectrum phage against Escherichia coli.

Multiple pathogenic types or serotypes restrict treatment for colibacillosis. In addition, rising antibiotic resistance has heightened public awareness to prevent and control pathogenic Escherichia coli. The bacteriophage is a viable technique to treat colibacillosis as an alternative to antibiotics. In this study, PH444, a relatively broad-spectrum obligate lytic phage, was screened from 48 Shiga toxin-producing Escherichia coli (STEC) phages isolated from farm manure samples and sewage samples in order to conduct genome-wide analysis, biological characterization, and a bacterial challenge experiment in milk. The results demonstrated that PH444 was a T7-like phage with a double-stranded DNA of 115,111bp that belongs to the Kuravirus and was stable at temperatures between 4 and 50°C and a pH range of 3 to 11. After adding PH444, the bacterial load in milk could be reduced from 3 × 103 PFU/ mL to zero within 1h. In consideration of the biological properties of phage PH444, it was, therefore, demonstrated that PH444 has the potential to be used in phage biocontrol.

Family Disadvantage, Education, and Health Outcomes Among Black Youths Over a 20-Year Period

Upward mobility (via educational attainment) is highly valued, but longitudinal associations with mental and physical health among Black youths are less understood. To examine associations of childhood family disadvantage and college graduation with adult mental and physical health in Black youths followed up into adulthood. This longitudinal, prospective cohort study of Black youths from the state of Georgia who were studied for 20 years (ages 11 to 31 years) was conducted between 2001 and 2022. Participants for this study were drawn from the Strong African American Healthy Adults Program. Data analysis was conducted from April 2023 to January 2024. Family economic disadvantage (measured during the adolescent years) and college graduation (indicating upward mobility). Primary outcomes included mental health, substance use, and physical health. Mental health included a composite of internalizing and disruptive problems (anxiety, depression, anger, aggressive behaviors, and emotional reactivity). Substance use included a composite of smoking, drinking, and drug use. Physical health included metabolic syndrome (MetS) and proinflammatory phenotypes (immune cells mounting exaggerated cytokine responses to bacterial challenge and being insensitive to inhibitory signals from glucocorticoids). Mental and physical health measures were taken at age 31 and during the adolescent years. Linear and logistic regression analyses, as well as mediated moderation analyses, were conducted. The study population consisted of 329 Black youths (212 women [64%]; 117 men [36%]; mean [SD] age at follow-up, 31 [1] years). Compared with those who did not graduate college, those who graduated from college had 0.14 SD fewer mental health problems (b = -1.377; 95% CI, -2.529 to -0.226; β = -0.137; P = .02) and 0.13 SD lower levels of substance use (b = -0.114; 95% CI, -0.210 to -0.018; β = -0.131; P = .02). Residualized change scores revealed that college graduates showed greater decreases from age 16 to 31 years in mental health problems (b = -1.267; 95% CI, -2.360 to -0.174; β = -0.133; P = .02) and substance use problems (b = -0.116; 95% CI, -0.211 to -0.021; β = -0.136; P = .02). For physical health, significant interactions between childhood family disadvantage and college completion emerged in association with MetS (OR, 1.495; 95% CI, 1.111-2.012; P = .008) and proinflammatory phenotype (b = 0.051; 95% CI, 0.003 to 0.099; β = 0.131; P = .04). Among youths growing up in disadvantaged households, college completion was associated with a 32.6% greater likelihood of MetS (OR, 3.947; 95% CI, 1.003-15.502; P = .049) and 0.59 SD more proinflammatory phenotype (mean difference, 0.249, 95% CI, 0.001 to 0.497; P = .049). Conversely, among those from economically advantaged backgrounds, college completion was correlated with lower MetS and less proinflammatory phenotype. Findings held after controlling for body mass index at age 19 years. In this longitudinal cohort study of Black youths, graduating from college was associated with an adult profile of better mental health but poorer physical health among those from economic disadvantage. These findings suggest that developing interventions that foster healthy outcomes across multiple life domains may be important for ensuring that striving for upward mobility is not accompanied by unintended cardiometabolic risk.

Natural history of periodontal disease: The original Sri Lanka and Oslo studies.

Susceptibility to periodontal disease depends on individual factors within the host response to the bacterial challenge. The study of these factors requires longitudinal studies of an undisturbed development of the disease process. On the basis of the original longitudinal studies on the natural histology of periodontal disease staged in Sri Lanka and Oslo/Norway, several analyses of periodontal parameters and tooth status have been performed. The main findings were that in the first 20 years of complete absence of oral hygiene practices or preventive services attachment was lost at various rates. Three groups of subjects could be identified: rapidly progressing (RP) (8%), moderately progressing (MP) (81%), and subjects with no disease progression (NP) (11%). In the second two decades, the RP subjects have lost most of their teeth and no NP patients were identified anymore. The progression rate in these two decades was much slower, and the tooth mortality decreased. It could be predicted that subjects who had lost more than 2 mm at age 30 would not maintain a functional dentition at age 60. The corresponding control population in Oslo was used to study the influence of gingival inflammation on the initiation and progression of periodontal disease. The pattern and rates of attachment loss were identified in a population that was exposed to optimal and regular preventive services from age 3 onward. In the observation period of 26 years, it could be demonstrated that gingival inflammation varied little throughout adult life and always bleeding sites occurred consistently in 10% to 20%. The role of ongoing gingivitis in the pathogenesis of attachment loss was identified and also reflected stability whenever it was absent. Tooth mortality was only found in subjects with ongoing gingivitis. After 50 years of tooth age, 63% of the teeth were still maintained, while 99.8% were maintained after 50 years when gingival inflammation had been absent. Consequently, continuous gingivitis represented a risk factor for tooth loss.

Exploring the nutritional value of corn fermented protein as a replacement for soybean meal in diets for juvenile channel catfish (Ictalurus punctatus): Impacts on production performance, intestinal health, and disease resistance

The use of corn fermented protein (CFP) can be a promising alternative ingredient to replace soybean meal (SBM) in aquafeeds, because of its attractive price and constant supply. In this study, the apparent digestibility of CFP was evaluated with channel catfish (Ictalurus punctatus), followed by a comparative feeding trial in which dehulled, solvent extracted SBM in the control diet was incrementally replaced with graded levels of CFP. A slight but significantly higher apparent protein digestibility coefficient was observed for SBM (91%) compared to CFP (89%). However, phosphorus and lipid were twice as available from CFP than SBM. A 60-day feeding trial was conducted, testing the gradual replacement of SBM by CFP on an equalnitrogen basis at 0, 25, 50, 75, and 100%. Channel catfish juveniles (∼11.6 g) were stocked in 30 aquaria (30 fish/ 110-L tank; n = 6) operating as a recirculating system and offered the experimental diets to apparent satiation twice daily. No differences were observed for hepatosomatic index, survival, or serum superoxide dismutase and lysozyme activities. Also, no treatment effects were observed for whole-body protein, ash, and protein conversion efficiency. However, weight gain, feed efficiency, intraperitoneal fat, and whole-body lipid were significantly affected by the dietary treatments with CFP able to replace up to 40 to 58% of the SBM in the diet while providing similar fish performance and body composition as fish fed the control diet. Digesta samples from the posterior segment of the intestine were subjected to DNA extraction, followed by 16S rRNA gene (V3-V4 region) sequencing to profile the intestinal microbiota. A higher relative abundance of Aeromonas spp. was observed for fish fed the control diet. The remaining fish from the feeding trial were challenged with Edwardsiella ictaluri, and cumulative survival was monitored for 21 days. Survival after the bacterial challenge gradually increased as the inclusion levels of CFP increased in the diets, with the 100% replacement treatment being significantly different from the control diet. In conclusion, CFP is a promising alternative plant protein ingredient with high protein and phosphorus digestibility, and it can replace up to 40 to 58% of the SBM in feed formulations for juvenile channel catfish based on weight gain and feed efficiency while potentially increasing disease resistance against E. ictaluri.

Group 3 innate lymphoid cells: A trained Gutkeeper.

Group 3 innate lymphoid cells (ILC3s) are tissue-resident immune lymphocytes that critically regulate intestinal homeostasis, organogenesis, and immunity. ILC3s possess the capacity to "sense" the inflammatory environment within tissues, especially in the context of pathogen challenges that imprints durable non-antigen-specific changes in ILC3 function. As such, ILC3s become a new actor in the emerging field of trained innate immunity. Here, we summarize recent discoveries regarding ILC3 responses to bacterial challenges and the role these encounters play in triggering trained innate immunity. We further discuss how signaling events throughout ILC3 ontogeny potentially control the development and function of trained ILC3s. Finally, we highlight the open questions surrounding ILC3 "training" the answers to which may reveal new insights into innate immunity. Understanding the fundamental concepts behind trained innate immunity could potentially lead to the development of new strategies for improving immunity-based modulation therapies for inflammation, infectious diseases, and cancer.

The Potential of Fish Protein Hydrolysate Supplementation in Nile Tilapia Diets: Effects on Growth and Health Performance, Disease Resistance, and Farm Economic Analysis.

Fish protein hydrolysate (FPH) has shown immense potential as a dietary protein supplement and immunostimulant in aquaculture, especially in Nile tilapia production. Four isoproteic diets (30% crude protein) were prepared by including FPH at varying percentages (0%, 0.5%, 1%, and 2%). Nile tilapia fed with FPH diets for 90 days, and their growth performance, feed utilization, blood biochemistry, liver and gut morphology, and resistance against Streptococcus iniae were investigated. The findings revealed that diets physical attributes such as pellet durability index and water stability were remarkably (p < 0.05) varied between experimental diet groups. Furthermore, the test diets were more palatable when FPH was included at 1% and 2%. Fish that were fed with a 2% FPH-treated diet had significantly (p < 0.05) greater growth indices than other treatments. Additionally, their feed utilization was significantly (p < 0.05) improved. The experimental diets and intestinal total bacteria count (TBC) exhibited a rising trend with FPH levels, where the 2% FPH-treated diet recorded the highest TBC. Neutrophil (109/L), lymphocyte (109/L), eosinophil (109/L), and red blood cell(1012/L) counts were significantly (p < 0.05) higher in the 2% FPH-treated group, while the white blood cell (109/L), and basophil (109/L) counts were not influenced by the FPH inclusion. Moreover, the FPH-treated groups displayed lower creatinine, bilirubin, and urea levels than the control. The histological examination demonstrated that themid-intestine of 2% FPH-fed Nile tilapia had an unbroken epithelial wall, more villi with frequent distribution of goblet cells, wider tunica muscularis, and stronger stratum compactum bonding than other treatments. Additionally, this group exhibited more nuclei and erythrocytes and less vacuolar cytoplasm in liver than their counterparts. Nile tilapia that were given a diet containing 2% FPH had significantly (p < 0.05) higher resistance (83.33%) to S. iniae during the bacterial challenge test. A significant (p < 0.05) enhancement in farm economic efficiency was observed in the higher inclusion of FPH in diets. In summary, 2% FPH supplementation in Nile tilapia diets improved their growth performance, feed utilization, health status, disease resistance, and farm economic efficiency.

Effect of an immune challenge and two feed supplements on broiler chicken individual breast muscle protein synthesis rate

Optimization of broiler chicken breast muscle protein accretion is key for the efficient production of poultry meat, whose demand is steadily increasing. In a context where antimicrobial growth promoters use is being restricted, it is important to find alternatives as well as to characterize the effect of immunological stress on broiler chicken's growth. Despite its importance, research on broiler chicken muscle protein dynamics has mostly been limited to the study of mixed protein turnover. The present study aims to characterize the effect of a bacterial challenge and the feed supplementation of citrus and cucumber extracts on broiler chicken individual breast muscle proteins fractional synthesis rates (FSR) using a recently developed dynamic proteomics pipeline. Twenty-one day-old broiler chickens were administered a single 2H2O dose before being culled at different timepoints. A total of 60 breast muscle protein extracts from five experimental groups (Unchallenged, Challenged, Control Diet, Diet 1 and Diet 2) were analysed using a DDA proteomics approach. Proteomics data was filtered in order to reliably calculate multiple proteins FSR making use of a newly developed bioinformatics pipeline. Broiler breast muscle proteins FSR uniformly decreased following a bacterial challenge, this change was judged significant for 15 individual proteins, the two major functional clusters identified as well as for mixed breast muscle protein. Citrus or cucumber extract feed supplementation did not show any effect on the breast muscle protein FSR of immunologically challenged broilers. The present study has identified potential predictive markers of breast muscle growth and provided new information on broiler chicken breast muscle protein synthesis which could be essential for improving the efficiency of broiler chicken meat production. SignificanceThe present study constitutes the first dynamic proteomics study conducted in a farm animal species which has characterized FSR in a large number of proteins, establishing a precedent for biomarker discovery and assessment of health and growth status. Moreover, it has been evidenced that the decrease in broiler chicken breast muscle protein following an immune challenge is a coordinated event which seems to be the main cause of the decreased growth observed in these animals.

Reproducible isolation of bovine mammary macrophages for analysis of host pathogen interactions

BackgroundMacrophages residing in milk are vital during intramammary infections. This study sought to develop a method enabling the investigation of macrophage responses to pathogens. Streptococcus uberis is the predominant cause of bovine mastitis UK-wide and its pathogenesis is unusual compared to other intramammary pathogens. Previous studies utilise macrophage cell lines, isolated bovine blood derived monocytes, or macrophages from raw milk through complex or inconsistent strategies such as fluorescence activated cell sorting (FACS), centrifugation and selective adherence, and CD14 antibody-microbeads. The centrifuge steps required in the initial stages often damage cells. Thus, the aim of this study was to develop a reliable, reproducible, and cost-effective method for isolating mammary macrophages from milk in a way that allows their culture, challenge with bacteria, and measurement of their response ex-vivo.ResultsThis method achieves an average yield of 1.27 × 107 cells per litre of milk. Whole milk with somatic cell range of 45–65 cells/µL produced excellent yields, with efficient isolations accomplished with up to 150 cells/µL. This strategy uses milk diluted in PAE buffer to enable low-speed centrifugation steps followed by seeding on tissue-culture-treated plastic. Seeding 1,000,000 milk-extracted cells onto tissue culture plates was sufficient to obtain 50,000 macrophage. Isolated macrophage remained responsive to challenge, with the highest concentration of IL-1β measured by ELISA at 20 h after challenge with S. uberis. In this model, the optimal multiplicity of infection was found to be 50:1 bacteria:macrophage. No difference in IL-1β production was found between macrophages challenged with live or heat-killed S. uberis. Standardisation of the production of IL-1β to that obtained following macrophage stimulation with LPS allowed for comparisons between preparations.ConclusionsA cost-effective method, utilising low-speed centrifugation followed by adherence to plastic, was established to isolate bovine mammary macrophages from raw milk. This method was shown to be appropriate for bacterial challenge, therefore providing a cost-effective, ex-vivo, and non-invasive model of macrophage-pathogen interactions. The optimal multiplicity of infection for S. uberis challenge was demonstrated and a method for standardisation against LPS described which removes sample variation. This robust method enables, reproducible and reliable interrogation of critical pathogen-host interactions which occur in the mammary gland.

Chronic TNF exposure induces glucocorticoid-like immunosuppression in the alveolar macrophages of aged mice that enhances their susceptibility to pneumonia.

Chronic low-grade inflammation, particularly elevated tumor necrosis factor (TNF) levels, occurs due to advanced age and is associated with greater susceptibility to infection. One reason for this is age-dependent macrophage dysfunction (ADMD). Herein, we use the adoptive transfer of alveolar macrophages (AM) from aged mice into the airway of young mice to show that inherent age-related defects in AM were sufficient to increase the susceptibility to Streptococcus pneumoniae, a Gram-positive bacterium and the leading cause of community-acquired pneumonia. MAPK phosphorylation arrays using AM lysates from young and aged wild-type (WT) and TNF knockout (KO) mice revealed multilevel TNF-mediated suppression of kinase activity in aged mice. RNAseq analyses of AM validated the suppression of MAPK signaling as a consequence of TNF during aging. Two regulatory phosphatases that suppress MAPK signaling, Dusp1 and Ptprs, were confirmed to be upregulated with age and as a result of TNF exposure both exvivo and invitro. Dusp1 is known to be responsible for glucocorticoid-mediated immune suppression, and dexamethasone treatment increased Dusp1 and Ptprs expression in cells and recapitulated the ADMD phenotype. In young mice, treatment with dexamethasone increased the levels of Dusp1 and Ptprs and their susceptibility to infection. TNF-neutralizing antibody reduced Dusp1 and Ptprs levels in AM from aged mice and reduced pneumonia severity following bacterial challenge. We conclude that chronic exposure to TNF increases the expression of the glucocorticoid-associated MAPK signaling suppressors, Dusp1 and Ptprs, which inhibits AM activation and increases susceptibility to bacterial pneumonia in older adults.

Bovine Colostrum Supplementation in Rabbit Diet Modulates Gene Expression of Cytokines, Gut-Vascular Barrier, and Red-Ox-Related Molecules in the Gut Wall.

Rabbits, pivotal in the EU as livestock, pets, and experimental animals, face bacterial infection challenges, prompting a quest for alternatives to curb antibiotic resistance. Bovine colostrum (BC), rich in immunoregulatory compounds, antimicrobial peptides, and growth factors, is explored for disease treatment and prevention. This study assesses BC diet supplementation effects on rabbit intestines, examining gene expression. Thirty female New Zealand White rabbits at weaning (35 days) were divided into three experimental groups: control (commercial feed), 2.5% BC, and 5% BC. The diets were administered until slaughtering (81 days). BC-upregulated genes in the jejunum included IL-8, TGF-β, and CTNN-β1 at 5% BC, while PLVAP at 2.5% BC. Antioxidant-related genes (SOD1, GSR) were downregulated in the cecum and colon with 2.5% BC. BC 5% promoted IL-8 in the jejunum, fostering inflammation and immune cell migration. It also induced genes regulating inflammatory responses (TGF-β) and gastrointestinal permeability (CTNN-β1). BC 5% enhanced antioxidant activity in the cecum and colon, but no significant impact on anti-myxo antibody production was observed. These results suggest that BC has significant effects on the rabbit gastrointestinal tract's inflammatory and antioxidant response, but further research is required to fully understand its histological and physiological impact.