Bacterial Challenge Research Articles

Identification and expression analysis of Th2 immune-related gene IL4/13A in turbot (Scophthalmus maximus)

Th2 immunity is a primary host defense against extracellular pathogens, and different IL4/13 paralogues are involved in this immune response in fish. Here, we identified IL4/13A for further Th2 immune response providing information in turbot. The results showed that the full length of the IL4/13A gene is 1,333 bp, containing a 432-bp open reading frame (ORF) that encoded 144 amino acids. Phylogenetic analysis recently showed that turbot IL4/13A has a relationship with Dicentrarchus labrax. Moreover, syntenic analysis revealed similar neighboring genes associated with turbot IL4/13A, compared with other teleosts and mammals. In addition, IL4/13A was widely expressed in all examined tissues with the highest expression level in skin, followed by liver and gill. Finally, IL4/13A showed a general trend of upregulation in immune tissues following bacterial challenge. The significant quick induction of IL4/13A indicated its key roles to prevent pathogens. Characterizations of IL4/13A will probably contribute to understanding of Th2 immunity in fish.

Study on the Effect of Phillyrin on Streptococcus suis In Vivo and In Vitro

As a zoonotic pathogen, S. suis serotype 2 (SS2) can cause severe diseases in both pigs and humans, and develop resistance to antibiotics. Plant natural compounds are regarded as promising alternatives to conventional antibiotics. Phillyrin is the major bioactive components of Chinese herbal medicine Forsythia suspensa. In this study, we explored the activity and action mechanism of phillyrin against SS2. The results showed that phillyrin could disrupt membrane integrity, destroy intracellular structures, and increase the exosmosis of DNA. Results of PCR revealed that phillyrin affected bacterial-virulence-related genes’ expression levels. Meanwhile, phillyrin significantly decreased the adhesion activity, inhibited lactate dehydrogenase (LDH) secretion, and reduced biofilm formation of SS2 in Newborn pig trachea epithelial (NPTr) cells. Furthermore, phillyrin protected tight junction protein of NPTr cells from SS2. We reported that phillyrin (0.1 mg/kg) treatment after bacterial challenge significantly improved the survival rate, ameliorated pulmonary inflammation, and inhibited the accumulation of multiple cytokines (IL-1, IL-6, IL-8, and TNF-α). Molecular docking showed that phillyrin had a good binding activity with the Ala88 and Asp111 of suilysin (SLY), one of the most important virulence factors of SS2. Collectively, phillyrin possesses antibacterial and anti-inflammatory activities, and is a promising candidate for preventing SS2 infection.

Phagostimulant MTSG supplementation benefits growth-promoting and defensive effects of Rhodobacter azotoformans on Chinese mitten crab Eriocheir sinensis

The use of phagostimulant and Rhodobacter has been well proposed in Chinese mitten crab Eriocheir sinensis culture. Yet the influence of phagostimulant supplementation on the growth-promoting and defensive effects of Rhodobacter on Chinese mitten crabs is still unknown. In this study, a 40-day feeding trial was carried out to examine the contribution of dietary phagostimulant MTSG to the effects of R. azotoformans on the growth performance, immunity, antioxidant capability, intestinal flora diversity, and resistance of E. sinensis to bacterial challenge. The results revealed higher weight gain, specific growth rate, as well as serum and hepatopancreatic acid phosphatase, alkaline phosphatase, superoxide dismutase, and catalase activities in the 3.0 to 9.0 g/kg diet MTSG + R. azotoformans-fed crabs than R. azotoformans-fed crabs, indicating that the supplementation of MTSG at 3.0 to 9.0 g/kg diet could improve the effect of R. azotoformans on growth performance, immunity and antioxidant capability of Chinese mitten crabs. Furthermore, the supplementation of MTSG at 3.0 to 9.0 g/kg diet could strengthen the effect of R. azotoformans on intestinal flora diversity of Chinese mitten crabs, and enhance the effect of R. azotoformans on the disease resistance against S. putrefaciens infection. To the best of our knowledge, this is the first study to reveal the contribution of a phagostimulant (MTSG) to the growth-promoting and defensive effects of R. azotoformans on Chinese mitten crabs.

Immunological Responses, Expression of Immune-Related Genes, and Disease Resistance of Rainbow Trout (Oncorhynchus mykiss) Fed Diets Supplied with Capsicum (Capsicum annuum) Oleoresin

A 45-day feeding study was carried out to assess the immune-stimulatory effects of capsicum oleoresin when added to rainbow trout diets. A total of 450 fish (mean weight: 155.20 ± 1.96 g) were distributed into 400 L tanks (30 fish/tank) across five experimental groups: control (CT, 0%), C7 (0.7%), C14 (1.4%), C21 (2.1%), and C28 (2.8%). Each group consisted of three replicate tanks. At the end of this period, hemato-biochemical parameters, innate immune responses, and immune-related gene expression levels were evaluated, and a histological examination of head kidney and liver sections was conducted. Finally, fish in all groups were challenged with Lactococcus garvieae and observed for an additional 20 days. The results revealed that oleoresin supplementation enhanced the immune responses of the treated fish, which was evidenced by the increased globulin, total protein, respiratory burst activity, and total immunoglobulin levels. The highest expression levels of the il-8, il-1β, TGF-β, and SAA genes was noticed in the C7 group, as compared with the results for the other groups. The IgT gene expression levels were higher in all experimental groups than in the CT group, and this increase was at the highest level in the C28 group. Following the bacterial challenge, all experimental groups displayed higher survival rates compared to that of the CT group. These values were 75.93, 72.22, 46.30, 33.33, and 29.63% in the C7, C14, C21, C28, and CT groups, respectively, with the C7 group displaying the highest survival rate among the groups. The histological examination of liver and head kidney tissues revealed that higher doses (in the C21 and C28 groups) showed an increase in cytoplasmic vacuolization, which causes adverse effects on fish health. However, the C7 group displayed normal histological structure in both tissues. Taken together, the most favorable immune responses were achieved in the C7 group, suggesting that 0.7% oleoresin could be applied to rainbow trout to boost immunity and protect the fish from diseases.

Variation in ovine serum vitamin D concentrations from late summer to early spring and during immune challenge with bacterial endotoxin

Vitamin D is most known for its role in the development and maintenance of the skeletal system. However, recent studies have found that vitamin D modulates the immune system, and at sufficient levels supports optimal immune function. Previous research has examined how vitamin D levels change seasonally and during immune stress challenges in both humans and cattle, but not sheep. Our study aimed to investigate the changes in the vitamin D status of sheep throughout the year in Southern Ontario as well as in response to a bacterial endotoxin challenge. Vitamin D was found to be significantly lower in spring when compared to summer and early winter levels. No significant differences were found in vitamin D levels among any measured time points during the immune challenge, although a trending increase was found 4 hr post-challenge. Overall, this study demonstrated how vitamin D levels in sheep change throughout the year, and slightly during endotoxin challenge. Given the importance of vitamin D in regulating optimal immune function, more investigation of these changes in vitamin D levels is warranted in terms of supporting optimal immune function.

Condensed tannins from black wattle as a promising nutritional additive for Nile tilapia: Growth, immune and antioxidant responses, and gut morphology

The present study aimed to investigate the potential of condensed tannins (CT), derived from black wattle (Acacia mearnsii) tree bark, as a nutritional additive for Nile tilapia (Oreochromis niloticus) juveniles. CT are known for their antioxidant and immune-modulating properties, and their inclusion in aquafeeds may enhance fish health and performance. In this study, six diets were formulated: one control diet (Con) with no additives, and five experimental diets supplemented with CT extract at concentrations of 150, 250, 500, 750, and 1000 mg/kg (CT150, CT250, CT500, CT750, CT1000, respectively). The selection of these doses was based on previous studies indicating that lower concentrations (below 1000 mg/kg of diet) of tannins may offer benefits, while higher concentrations could exhibit antinutritional effects. After a 90-day feeding trial, fish fed the CT150 diet exhibited significantly higher growth compared to the control group. Additionally, fish in the CT150 group showed higher plasma lysozyme activity, while myeloperoxidase activity and hemolytic activity of the complement system were significantly higher in all tannin-fed groups compared to the control. Notably, CT150-fed fish demonstrated liver antioxidant responses comparable to or better than the control group, with no significant increase in lipid peroxidation, suggesting antioxidant protection. Histological analysis revealed a significant increase in intestinal villi density in the CT150 and CT500 groups compared to the control. While all groups showed an increase in absorption surface area (ASA), this increase was statistically significant only in the CT250, CT500, and CT750 groups when compared to the control. Furthermore, the CT150 diet led to the highest survival rate (80 %) following a bacterial challenge. These findings suggest that dietary supplementation with 150 mg/kg of CT from black wattle bark provides the most beneficial effects on growth, immune response, and survival in Nile tilapia juveniles.

Integrated multi-omics identifies pathways governing interspecies interaction between A. fumigatus and K. pneumoniae

Polymicrobial co- and superinfections involving bacterial and fungal pathogens pose serious challenges for diagnosis and therapy, and are associated with elevated morbidity and mortality. However, the metabolic dynamics of bacterial–fungal interactions (BFI) and the resulting impact on disease outcome remain largely unknown. The fungus Aspergillus fumigatus and the bacterium Klebsiella pneumoniae are clinically important pathogens sharing common niches in the human body, especially in the lower respiratory tract. We have exploited an integrated multi-omics approach to unravel the complex and multifaceted processes implicated in the interspecies communication involving these pathogens in mixed biofilms. In this setting, A. fumigatus responds to the bacterial challenge by rewiring its metabolism, attenuating the translational machineries, and by connecting secondary with primary metabolism, while K. pneumoniae maintains its central metabolism and translation activity. The flexibility in the metabolism of A. fumigatus and the ability to quickly adapt to the changing microenvironment mediated by the bacteria highlight new possibilities for studying the impact of cross-communication between competing interaction partners. The data underscore the complexity governing the dynamics underlying BFI, such as pronounced metabolic changes mounted in A. fumigatus interacting with K. pneumoniae. Our findings identify candidate biomarkers potentially exploitable for improved clinical management of BFI.

Efficacy of Feed-Based Genome-Free Bacterial Vaccine Against Aeromonas hydrophila Infection in Red Tilapia (Oreochromis sp.).

Aeromonas hydrophila causes motile Aeromonas septicemia (MAS), a disease with a high mortality rate in tilapia culture. Feed-based vaccines with the incorporation of inactivated whole-cell bacteria into the feed offer promising tools to control MAS. Currently, the incorporation of genome-free bacteria as bacterial vaccine through the implementation of SimCells® technology into the feed has become a particular interest. Background/Objectives: This study investigates the efficacy of a feed-based vaccine incorporating genome-free A. hydrophila (FBV-GFAH) against MAS infection in red tilapia. Methods: The vaccine was prepared and delivered at 5% fish body weight for three consecutive days in weeks 0 (prime vaccination) and 2 (first booster vaccination), orally. Throughout a five-week experimental period, the immune-related genes (IL-1β, MHC-II, CD4, IgT, and IgM) expression in the hindgut and head kidney of the fish was determined using RT-qPCR assay. Lysozyme (serum) and overall IgM (serum, gut lavage, and skin mucus) productions were also detected. Results: Fish vaccinated with FBV-GFAH showed a significant (p ≤ 0.05) improvement in relative percent survival compared with unvaccinated fish following bacterial challenge. FBV-GFAH induced the expression of immune-related genes in the hindgut and head kidney, especially after booster vaccination. Furthermore, serum lysozyme activity and overall IgM production in serum, skin mucus, and gut lavage were also significantly (p ≤ 0.05) improved in the FBV-GFAH vaccinated fish than the unvaccinated fish. Conclusions: This study showed that FBV-GFAH is a promising feed-based vaccine technology to control MAS in cultured tilapia.

The RNA receptor RIG-I binding synthetic oligodeoxynucleotide promotes pneumonia survival.

Pneumonia is a worldwide threat to public health, demanding novel preventative and therapeutic strategies. The lung epithelium is a critical environmental interface that functions as a physical barrier to pathogen invasion while also actively sensing and responding to pathogens. We have reported that stimulating lung epithelial cells with a combination therapeutic consisting of a diacylated lipopeptide and a synthetic CpG oligodeoxynucleotide (ODN) induces synergistic pneumonia protection against a wide range of pathogens. We report here that mice deficient in TLR9, the previously described receptor for ODN, still displayed partial ODN-induced protection. This prompted us to seek an alternate ODN receptor, and we discovered by mass spectroscopy that the RNA sensor RIG-I could also bind DNA-like ODN. ODN binding by RIG-I resulted in MAVS-dependent pneumonia-protective signaling events. While RIG-I is essential to native defenses against viral infections, we report that therapeutic RIG-I activation with ODN promoted pathogen killing and host survival following both viral and bacterial challenges. These data indicate that maximal ODN-induced pneumonia protection requires activation of both the TLR9/MyD88 and RIG-I/MAVS signaling pathways. These findings not only identify what we believe to be a novel pattern recognition receptor for DNA-like molecules, but reveal a potential therapeutic strategy to protect susceptible individuals against lethal pneumonias during periods of peak vulnerability.

TNF-alpha gene promoter's hypomethylation mediates a pro-inflammatory phenotype in peripheral blood monocytes from apical periodontitis individuals.

Epigenetic regulation of the key inflammatory genes plays a crucial role in controlling monocyte/macrophage-mediated local and systemic responses to bacterial challenges. However, it has not been addressed in apical periodontitis (AP). We aimed to explore the methylation pattern of the TNF-α gene promoter and its association with the inflammatory phenotype of peripheral blood monocytes from individuals with AP and controls. A cross-sectional study was conducted, including otherwise healthy individuals with AP (n = 25) and controls (n = 29). Monocytes were isolated from the volunteer's blood samples using a Ficoll gradient followed by negative immunoselection. RNA and DNA were extracted. The DNA methylation profiles of the TNF-α gene promoter region were analyzed using bisulfite sequencing PCR. The mRNA expression levels of DNA methyltransferases 3a (DNMT3a) and Ten Eleven Translocation enzymes 1(TET1) were assessed by qPCR. A fraction of primary monocytes was also cultured for 24 h, and the supernatant was collected to measure cytokine levels through a Luminex assay. Generalized structural equation models (GSEM) evaluated the association between AP, DNA methylation, and TNF-α protein expression controlled for potential covariates. Models included the effect of the methylation of TNF-α gene promoter as a mediator of the association between AP and TNF-α protein expression levels. Monocytes from AP individuals exhibited a heightened secretion of TNF-α and IL-1β and hypomethylation of the TNF gene promoter (p < .05). AP diagnosis was associated with the TNF-α gene promoter´s hypomethylated profile and enhanced pro-inflammatory cytokine levels, while lower methylation of the gene promoter region and -163 CpG single site mediated TNF-α overexpression (p < .05). DNA hypomethylation at the TNF-α gene mediates a proinflammatory phenotype in monocytes from AP patients, supporting a role in the systemic response.

Nano-Bacillus amyloliquefaciens as a dietary intervention in nile tilapia (Oreochromis niloticus): Effects on resistance to Aeromonas hydrophila challenge, immune-antioxidant responses, digestive/absorptive capacity, and growth

The present investigation proposed an innovative trial at the probable beneficial effects of nano-Bacillus amyloliquefaciens (NBA) dietary incorporation on the Nile tilapia (Oreochromis niloticus). The investigation included the impact on the growth, digestive functions, immune-antioxidant indices, and resistance to Aeromonas hydrophila challenge. A total of 135 fish (35.14 ± 0.12 g) were equally allocated into three groups (45 fish/group; 15 fish/replicate) in triplicates for 70 days. The control, NBA2, and NBA4 groups were fed basal diets enriched with 0, 102, and 104 CFU/kg NBA, respectively. Following the feeding experiment, all experimental groups were injected with 0.1 mL (1.5 × 106) A. hydrophila, and the fish mortalities were observed for 14 days. The outcomes showed that dietary NBA (NBA4 followed by the NBA2 diet) augmented the growth variables (final body weight, total weight gain, and specific growth rate) and condition factor and declined the feed conversion ratio. The intestinal digestive enzyme (amylase and lipase) and growth hormone levels were increased and the serum glucose level was decreased by dietary NBA. Furthermore, NBA diets enhanced the immune (total protein, globulin, lysozyme, complement 3, myeloperoxidase, and phagocytic activity) and antioxidant (superoxide dismutase, catalase, and total antioxidant capacity) parameters. The intestinal histology revealed no pathological lesions with a significant improvement in the intestinal histomorpho-measures (villus height and width, villus surface area, lamina propria thickness, and tunica muscularis thickness) by NBA diets in a dose-dependent manner. In the fish intestine, the B. amyloliquefaciens count was increased in the NBA groups (NBA4 group followed by NBA2 group) with no discernible difference in the total bacterial count. Fish resistance to the A. hydrophila challenge was increased by increasing the survival % in the NBA4 group (91.70 %) followed by the NBA2 group (83.30 %) compared to the control group (70.80 %). Overall, dietary NBA (especially 104 CFU/kg diet) could be a promising feed supplement in the Nile tilapia diets for improving their growth, health, and resistance to bacterial challenges.

Transcriptomic and bioinformatic analysis of Histone H2A-genes in Lates calcarifer and Oreochromis niloticus upon Vibrio vulnificus challenge

Histone H2A-derived antimicrobial peptides (AMPs) have been recognized for their antimicrobial properties and role in host defense mechanisms. The present study investigated the transcriptional responses of two histone H2A-derived AMPs, Lc-Histone from Lates calcarifer and GIFT-Histone from Oreochromis niloticus, in response to Vibrio vulnificus infection. The genes encoding these peptides were amplified and sequenced from fingerling stage fishes, followed by in silico analysis and phylogenetic characterization using ClustalW. Computational methods were used to assess their functional attributes, and in vivo bacterial challenge assay was employed to evaluate the efficacy. Sequence alignment identified conserved histone motifs, supporting their evolutionary conservation. The antimicrobial peptide database predicted membrane interaction potential, suggesting antimicrobial activity, alongside biofilm-active properties and efficient expression in heterologous protein systems. Expression profiling of Histone H2A-derived AMP genes under Gram-negative bacterial stimulation revealed distinct temporal dynamics, indicative of their innate immune response involvement. Significant gene expression variations were observed between L. calcarifer and O. niloticus. These baseline findings enhance understanding of host defense mechanisms and bacterial interactions in aquaculture. The antimicrobial potential of histone H2A-derived AMPs highlights their significance as therapeutic agents and warrants further investigation into their mechanisms of action and optimal usage in aquaculture settings.

Dietary vitamin E (α-tocopherol acetate) modulates growth, digestive enzymes, histopathology, and vulnerability of Nile tilapia, Oreochromis niloticus to Aeromonas hydrophila infection

Vitamin E is a fat-soluble vitamin that plays a vital function in several biological processes, and fish cannot synthesize it to meet their requirement. So, 56-day research was conducted to examine the influence of vitamin E (vit-E) (α-tocopherol acetate) on the Nile tilapia's growth, digestive enzymes, hematology, histology, and susceptibility to Aeromonas hydrophila. A total of 450 mono-sex Acclimated Nile tilapia (Oreochromis niloticus) were haphazardly dispersed into 30 aquaria, each with a capacity of 100 liters (15 fish/aquarium) to exemplify five groups with six replicates. A control diet (30 % protein) was enriched with 0.0 (E0), 150 (E150), 300 (E300), 600 (E600), and 1200 (E1200) mg/kg feed. Fish (13.5 ± 0.12 g) were given the trial diets until obvious satiation thrice daily for 56 days. At the end of the feeding trial, growth performance, digestive enzymes, hematology, and histology for the mid-intestine were examined. Subsequently, twenty fish from each treatment were challenged to contagion with Aeromonas hydrophila bacteria, and fish mortality was recorded for a further 14 days. At the end of the bacterial challenge, histology for the mid-intestine, liver, and spleen tissues was examined. Growth performance, feed utilization, and digestive enzyme secretion (proteases, lipase, and α-amylase) were substantially (P < 0.5) improved with raising vit-E levels in fish feeds up to E1200. Increasing the vit-E doses improved fish gut histomorphology by increasing the count and size of intestinal folds bordered by well-arranged enterocytes. The body composition was not influenced by dietary vitamin E, except lipid content, which increased substantially as vitamin E levels increased. Fish fed with vita-E enriched diets had higher resistance to A. hydrophila infection; however, the control group exhibited the greatest fish mortality rate (80 %), while the lowest rate was observed at E1200 (30 %). Hepatic and spleen tissues in the control group (E0) showed severe congestion and degeneration, whereas vit-E-treated fish groups progressively recovered normal histomorphology depending on the vit-E doses. Finally, this research recommends feeding Nile tilapia on vit-E, particularly 1200 mg/kg feed, to enhance its performance, welfare status, and resistance to A. hydrophila contagion.

Enriched Diet With Orange Essential Oil Citrus sinensis for Tambaqui Colossoma macropomum Promotes Growth Performance and Resistance Against Aeromonas hydrophila.

The objective of this study was to evaluate the effect of dietary supplementation with Citrus sinensis (L.) Osbeck essential oil on the growth, immune system, and resistance to Aeromonas hydrophila infection in Colossoma macropomum fingerlings. The experiment was conducted with five treatments (control diet, Tween80 diet, and diets supplemented with 200, 400, and 800 mg L-1 of C. sinensis essential oil) with four replicates. At the end of the experimental period, growth parameters were measured, and blood samples were collected for thrombogram, leukogram, and phagocytic activity analysis. A bacterial challenge with A. hydrophila was conducted for 96 h. C. macropomum fingerlings that were fed with 400 and 800 mg L-1 of C. sinensis essential oil had the highest growth parameters, with final weights of 533.18 ± 2.03 mg and 531.91 ± 2.67 mg, respectively, and an increase in the number of thrombocytes, lymphocytes, monocytes, and neutrophils, as well as higher phagocytosis rates compared to the control group. Regarding the challenge, fish in the 400 and 800 mg L-1 treatments also exhibited the lowest cumulative mortality rate (26.66% ± 3.33%). Therefore, supplementation with C. sinensis essential oil promotes growth, improves health, and enhances resistance to A. hydrophila in C. macropomum fingerlings.

A novel murine wire injury based infectious endocarditis model aggravates aortic valve stenosis

Abstract Background With mortality rates up to 40% under optimal medical treatment, infectious endocarditis (IE) remains one of the deadliest infectious diseases worldwide. In the early stages of IE, diagnosis and treatment initiation are often difficult and research is limited due to unreliable models. Most murine IE models rely on permanent prosthetic wire placement and i.v. bacterial injection, thus making it inadequate to study IE pathophysiology with the aim to reflect the course of disease in humans. Purpose To develop a reproducible murine IE model, based on aortic valve injury preceding intravenous S. aureus bacterial challenge. Methods Using a wire-injury model we induced endothelial lesions. 72h after wire injury (WI), we performed intravenous bacterial challenge using S. aureus. Echocardiography was performed to confirm bacterial vegetations and assess valvular function. Cross sections of valvular leaflets were prepared for scanning electron microscopy (SEM). Macrophage, neutrophil and S. aureus-specific immunofluorescence staining was performed and visualized by immunofluorescence microscopy. Bacterial cultivation was performed from peripheral blood and valve tissue. Systemic immune response was analyzed using flow cytometry. Results WI induced endothelial damage in all mice verified using scanning electron microscopy imaging. The combination of wire injury and injection of 10^5 and 10^6 colonies-forming-units (CFU) S. aureus reliably caused IE validated via in vivo echocardiography as well as S.aureus immunofluorescence staining and SEM imaging. Mice undergoing bacterial challenge responded with significant neutrophilia in the blood. Using 10^6 CFU S. aureus was associated with an increased mortality, liver and kidney abscess formation due to systemic bacterial spreading compared to 10^5 CFU. Peak velocity across the aortic valve and aortic valve leaflet thickness as markers for aortic valve stenosis were increased in IE compared to WI only mice. Aortic regurgitation was more prevalent after bacterial challenge mediating increased left ventricular volumes, meanwhile ejection fraction was not altered. Immunofluorescence staining revealed a pro-inflammatory milieu including increased macrophage and neutrophil infiltration in IE compared to WI only mice. Conclusion In vivo echocardiography and ex vivo histological staining revealed reliable IE induction using our new model with S. aureus concentrations of 10^5 CFU. The additional bacterial challenge causes a more severe aortic valve stenosis compared to WI only mice, potentially due to an observed increase in valvular inflammation.Fig 1 Scanning electron microscopy after

In-vitro and in-vivo assessment of the bactericidal potential of peracetic acid and hydrogen peroxide disinfectants against A. hydrophila infection in Nile tilapia and their effect on water quality indices and fish stress biomarkers

This study aimed to assess the in vitro and in vivo disinfectant potential of peracetic acid (PAA) (1 mg/L) and hydrogen peroxide (H2O2) (20 mg/L) on the physicochemical and microbiological water quality parameters of fish aquaria, the microbial density of Nile tilapia muscular tissue, fish hepatic cortisol levels, and antioxidant biomarkers. In vitro, PAA and H2O2 reduced A. hydrophila colony viability by 5 log units after 30 and 5 min of contact time, respectively. PAA and H2O2 were added to aquaria water twice a week for the three-week experiment. Increased fish escape reflexes were observed only in the PAA group, which returned to normal within 10 min. No mortalities were reported in either the PAA or H2O2 groups. An in vivo experimental challenge with a pathogenic strain of A. hydrophila revealed a 20% reduction in mortality in the PAA group, with no mortalities in the H2O2 group. Cortisol levels and antioxidant markers were measured to assess the impact of PAA and H2O2 on fish health. Cortisol levels in the PAA and H2O2 groups were significantly higher than in the control group after disinfectant exposure, but they progressively returned to normal. A significant reduction in superoxide dismutase (SOD) and catalase (CAT) activity, along with considerably higher glutathione peroxidase (GPx) and malondialdehyde (MDA) enzymatic activity, was observed in the PAA and H2O2 groups compared to the control group. A substantial increase in total antioxidant capacity (TAC) was recorded in the PAA group. Physicochemical analyses revealed reduced pH and increased dissolved oxygen levels in the PAA and H2O2 groups. Microbiological analyses showed a significant reduction in bacterial density in water by 64% and 76% after 30 min of exposure to PAA and H2O2, respectively, with a non-significant increase in microbial count after bacterial challenge. Additionally, aerobic bacterial count, Aeromonas spp., and psychotropic bacterial count in fish muscle showed a significant reduction in the H2O2 group compared to the PAA and control groups before and after infection. The study concludes that regular application of PAA and H2O2 can temporarily reduce bacterial load in aquaria and fish muscle, regulate stress responses, and improve fish health by reducing A. hydrophila-induced infections and improving survival.

MdSVWC1, a new pattern recognition receptor triggers multiple defense mechanisms against invading bacteria in Musca domestica

BackgroundSingle-domain von Willebrand factor type C (SVWC) constitute a protein family predominantly identified in arthropods, characterized by a SVWC domain and involved in diverse physiological processes such as host defense, stress resistance, and nutrient metabolism. Nevertheless, the physiological mechanisms underlying these functions remain inadequately comprehended.ResultsA massive expansion of the SVWC gene family in Musca domestica (MdSVWC) was discovered, with a count of 35. MdSVWC1 was selected as the representative of the SVWC family for functional analysis, which led to the identification of the immune function of MdSVWC1 as a novel pattern recognition receptor. MdSVWC1 is highly expressed in both the fat body and intestines and displays acute induction upon bacterial infection. Recombinant MdSVWC1 binds to surfaces of both bacteria and yeast through the recognition of multiple pathogen-associated molecular patterns and exhibits Ca2+-dependent agglutination activity. MdSVWC1 mutant flies exhibited elevated mortality and hindered bacterial elimination following bacterial infection as a result of reduced hemocyte phagocytic capability and weakened expression of antimicrobial peptide (AMP) genes. In contrast, administration of recombinant MdSVWC1 provided protection to flies from bacterial challenges by promoting phagocytosis and AMP genes expression, thereby preventing bacterial colonization. MdSPN16, a serine protease inhibitor, was identified as a target protein of MdSVWC1. It was postulated that the interaction of MdSVWC1 with MdSPN16 would result in the activation of an extracellular proteolytic cascade, which would then initiate the Toll signaling pathway and facilitate the expression of AMP genes.ConclusionsMdSVWC1 displays activity as a soluble pattern recognition receptor that regulates cellular and humoral immunity by recognizing microbial components and facilitating host defense.

Passive immunoprophylaxis with Ccombodies against Vibrio parahaemolyticus in Pacific white shrimp (Penaeus vannamei)

The Vibrio parahaemolyticus strain causing acute hepatopancreatic necrosis disease (AHPND) in shrimp secretes toxins A and B (PirAVp/PirBVp). These toxins have been implicated in pathogenesis and are targets for developing anti-AHPND therapeutics or prophylactics that include passive immunization. We have previously reported that Ccombodies (recombinant hagfish variable lymphocyte receptor B antibodies; VLRB) targeting PirBVp conferred protection against V. parahaemolyticus in shrimp when administered as a feed supplement. In this study, we screened a phage-displayed library of engineered VLRBs for PirAVp-targeting Ccombodies that were mass-produced in a bacterial expression system. We then introduced these Ccombodies into the diet of Pacific white shrimp (Penaeus vannamei) over a seven-day period. Subsequently, the shrimp were exposed to a challenge with V. parahaemolyticus. Mortality rates were then observed and recorded over the following seven days. Administering shrimp feed supplemented with Ccombodies at a high dose (100 mg per 100 g feed) reduced mortality in recipient animals (2.96–5.19 %) statistically similar to mock-challenged control (1.48 %), but significantly different from the Ccombody-deficient control (74.81 %). This suggests that the Ccombodies provided strong protection against the bacterium. Feeding shrimp with a median dose (10 mg/100 g feed) gave statistically comparable low mortality (5.93–6.67 %) as the high dose. Reducing the Ccombody dose to 1 mg/100 g feed showed variable effects. Ccombody A2 showed mortality (11.85 %) significantly lower than that of the Ccombody-deficient group (74.81 %), suggesting that it can effectively protect against the bacterial challenge at a low dose. Our results demonstrate the ability of the phage-displayed VLRB library to generate antigen-specific Ccombodies rapidly and simply, with the expression of high protein levels in bacteria. The protective effect provided by these Ccombodies aligns with our earlier results, strongly supporting the use of VLRB antibodies as a substitute for IgY in passive immunoprophylaxis against AHPND in shrimp.

The Determination of the Rapid and Effective Activity of an Air Sanitizer against Aerosolized Bacteria Using a Room-Sized Aerobiology Chamber.

Air sanitization is an important non-pharmaceutical intervention for mitigating the risk of indoor pathogen spreading. A dipropylene glycol-containing air sanitizer was tested against aerosolized Staphylococcus aureus and Klebsiella pneumoniae. The bacteria, suspended in a soil load, were aerosolized using a six-jet Collison nebulizer with pressurized air. The 25-m3 (~900 ft3) aerobiology chamber was maintained at 22 ± 2 °C and 50 ± 5% relative humidity per the U.S. Environmental Protection Agency's 2012 Guidelines on air sanitizers. An initial 2-min air sample was collected from the chamber using a slit-to-agar sampler containing 150-mm Petri plates, with Trypticase soy agar (TSA) containing neutralizers to quench the microbicidal activity of the air sanitizer, to determine the initial bacterial challenge in the air. The air sanitizer was sprayed into the chamber from pressurized cans. Additional air samples were collected from the chamber over 10 min to detect surviving bacteria. The TSA plates were then incubated aerobically at 36 ± 1 °C for 90 ± 4 h and scored for bacterial colony-forming units. A 30-s spray of the air sanitizer reduced infectious S. aureus and K. pneumoniae titers by 3.0 log10 (99.9%) in 3.2 ± 0.3 min and 1.2 ± 0.0 min, respectively. Based on these findings, the EPA granted registration of the air sanitizer as the first product of its kind for indoor air sanitization.

Evaluation of inhibitory, immunomodulatory, survival, and growth effects of host-derived Weissella confusa on Macrobrachium rosenbergii challenged with Vibrio parahaemolyticus

Macrobrachium rosenbergii is a highly valuable prawn species in aquaculture due to its current growing demand in the market. However, various bacterial diseases caused by Vibrio parahaemolyticus have been observed to induce mortality in larval, juvenile, and adult stages of M. rosenbergii. While antibiotics are commonly used to manage these diseases, they contribute to antimicrobial resistance, a global concern with serious environmental and health implications. Consequently, the scientific community is exploring host-derived beneficial microorganisms as functional feed additives, offering eco-friendly alternatives that boost shrimp and prawn immunity. The purpose of this study was to examine the effects of host-derived beneficial microorganisms from the gut of M. rosenbergii, on the inhibition, immunomodulation, survival, and growth of M. rosenbergii when exposed to V. parahaemolyticus. The result of this study showed that Weissella confusa C6, a lactic acid bacterium isolated from the gut of M. rosenbergii, significantly inhibits the growth of V. parahaemolyticus R1 as demonstrated in agar well diffusion assay and co-culture assay. Additionally, it showed immunomodulatory effects, indicated by elevated total hemocyte count (THC) and phenoloxidase (PO) activity during the bacterial challenge test. Furthermore, supplementing the diet with W. confusa led to significant improvements in M. rosenbergii, including increased survival rates, weight gain, specific growth rate, better feed conversion ratio, and reduced density of V. parahaemolyticus in the prawn's gut. Therefore, it can be considered a viable option for use as a feed supplement in prawn farming, enhancing M. rosenbergii's immune system, disease resistance, and overall health.