Pharmacokinetic behaviour and pharmacokinetic-pharmacodynamic integration of doxycycline in rainbow trout (Oncorhynchus mykiss) after intravascular, intramuscular and oral administrations.

OBJECTIVE: Doxycycline (DO) has been used in fish for a long time, but there are some factors that have not yet been clarified regarding its pharmacokinetic (PK) and pharmacodynamic (PD) properties. Therefore, the aim of this study was to investigate the PK and PK/PD targets of DO after 20 mg/kg intravascular (IV), intramuscular (IM) and oral (OR) gavage administration in rainbow trout (Oncorhynchus mykiss). METHODS: Plasma samples were collected at specific time points and subsequently analysed by HPLC-ultraviolet. The PK/PD indices were calculated based on the MIC90 (Aeromonas hydrophila and Aeromonas sobria) values obtained for the respective bacteria and the PK parameters obtained for DO following both IM and OR administration. RESULTS: After IV administration, the elimination half-life (t1/2 ʎz), area under the concentration vs. time curve (AUC), apparent volume of distribution at steady-state and total body clearance of DO were 34.81 h, 723.82 h µg/mL, 1.24 L/kg and 0.03 L/kg/h, respectively. The t1/2λz of the DO was found to be 37.39 and 39.78 h after IM, and OR administration, respectively. The bioavailability was calculated 57.02% and 32.29%, respectively, after IM and OR administration. The MIC90 of DO against A. hydrophila and A. sobria was 4 µg/mL. The PK/PD integration showed that DO (20 mg/kg dose) for A. hydrophila and A. sobria with MIC90 ≤4 µg/mL achieved target AUC/MIC value after IM administration. CONCLUSIONS: These results suggest that when rainbow trout was treated with 20 mg/kg IV and IM administered DO, therapeutically effective concentrations were reached in the control of infections caused by A. hydrophila and A. sobria.

Dietary effects of Saccharomyces cerevisiae and Allium sativum on growth, antioxidant status, hepatic and intestinal histoarchitecture, expression of growth- and immune-related genes, and resistance of Oreochromis niloticus to Aeromonas sobria.

This study investigated the benefits of yeast, Saccharomyces cerevisiae and/or garlic, Allium sativum supplementation in diets of Nile tilapia with regard to growth, antioxidant status, hepatic and intestinal histoarchitecture, expression of growth- and immune-related genes, and resistance to Aeromonas sobria infection. Fish (with an initial weight of 9.43 ± 0.08 g) were allocated to twelve hapas, organized into four triplicate treatment groups defined as control (no supplementation), yeast (4 g/kg diet), garlic (30 g/kg diet), and a mixture of both. This trial continued over a 60-day feeding period. Results revealed that combined treatment (yeast + garlic) demonstrated the most promising outcomes regarding growth, with significantly higher final body weights, weight gains, and specific growth rates compared to other groups. Moreover, this combination enhanced hepatic antioxidant status, as evidenced by elevated levels of reduced glutathione and activities of catalase and superoxide dismutase enzymes, reflecting improved defense against oxidative stress. Histological assessments of the livers and intestines demonstrated structural enhancements in yeast and garlic treatments, suggesting improvements in organ health. In comparison to the control, the gene expression analyses unveiled increased expression of growth-related (igf-1 and ghr1) and immune-related (il-10, lyz, and hep) genes in the test groups, indicating a possible reinforcement of the growth and immune responses. The combined treatment also showed the highest resistance to A. sobria infection, as evidenced by improved survival rates and lower mortality compared with the other groups. These findings highlight the benefits of a combination of both yeast and garlic as a dietary supplementation regimen. In conclusion, this study suggests that the combined treatment regimen could be considered an effective strategy to promote the health and productivity of Nile tilapia under production conditions.

Dietary effects of microalga Tetraselmis suecica on growth, antioxidant-immune activity, inflammation cytokines, and resistance of Nile tilapia fingerlings to Aeromonas sobria infection.

The dietary effects of the green microalga Tetraselmis suecica (TS) on the growth, digestive enzymes, immune and antioxidant responses, genes expression, and disease resistance of Nile tilapia (Oreochromis niloticus) fingerlings were investigated. This microalga was mixed with the diet' ingredients at doses of 0.0 (the control), 5, 10, 15, and 20 g/kg diet and then fed to fish daily for 84 days. After the feeding trial, fish were experimentally challenged with Aeromonas sobria, infection and fish mortalities were recorded for another 10 days. Dietary TS significantly (p < 0.05) enhanced growth, digestive enzymes activities, and blood proteins, particularly at the level of 15 g/kg diet. Feeding the fish on 15 TS/kg feed exhibited highest mRNA expressions of GH and IGF-1 genes as well as SOD, CAT, and GPx genes compared to other TS groups. Moreover, highest levels of hepatic antioxidant and immune indices were found in the treatment of 15 g TS/kg feed. Significant downregulation of IL-1ß and IL-8 genes expression and significant upregulation of IL-10 gene expression were observed in TS-fed fish, principally in fish groups fed on 15-20 g TS/kg feed. Conversely, hepatic malondialdehyde levels, blood glucose, and the activities of transaminases (ALT and AST) were significantly (p < 0.05) decreased in fish fed with 15-20 g TS/kg diet. Serum bactericidal activity against A. sobria was significantly higher in TS-fed fish groups, and its highest levels were found in treatments of 15-20 g/kg diet. Of interest, the survival rates of fish groups fed diets with 10-20 g TS/kg feed were higher after the challenge with A. sobria infection than the control group. Accordingly, we can conclude that supplementing fish diets with a 15 g TS/kg diet enhanced the growth, antioxidant and immune activities, and resistance of Nile tilapia fingerlings to possible A. sobria infection.

Chitosan neem nanocapsule enhances immunity and disease resistance in nile tilapia (Oreochromis niloticus).

Finding eco-friendly alternatives for antibiotics in treating bacterial diseases affecting the aquaculture sector is essential. Herbal plants are promising alternatives, especially when combined with nanomaterials. Neem (Azadirachta indica) leaves extract was synthesized using a chitosan nanocapsule. Chitosan neem nanocapsule (CNNC) was tested in-vitro and in-vivo against the Aeromonas sobria (A. sobria) challenge in Nile tilapia. A preliminary experiment with 120 Nile tilapia was conducted to determine the therapeutic dose of CNNC, which was established to be 1 mg/L. A treatment study was applied for seven days using 200 fish categorized into four groups (10 fish/replicate: 50 fish/group). The first (control) and second (CNNC) groups were treated with 0 and 1 mg/L CNNC in water without being challenged. The third (A. sobria) and fourth (CNNC + A. sobria) groups were treated with 0 and 1 mg/L CNNC, respectively, and challenged with A. sobria (1 × 107 CFU/mL). Interestingly, CNNC had an in-vitro antibacterial activity against A. sobria; the minimum inhibitory concentration and minimum bactericidal concentration of CNNC against A. sobria were 6.25 and 12.5 mg/mL, respectively. A. sobria challenge caused behavioral alterations, skin hemorrhage, fin rot, and reduced survivability (60%). The infected fish suffered a noticeable elevation in the malondialdehyde level and hepato-renal function markers (aspartate aminotransferase, alanine aminotransferase, and creatinine). Moreover, a clear depletion in the level of the antioxidant and immune indicators (catalase, reduced glutathione, lysozymes, nitric oxide, and complement 3) was obvious in the A. sobria group. Treatment of the A. sobria-challenged fish with 1 mg/L CNNC recovered these parameters and enhanced fish survivability. Overall, CNNC can be used as a new versatile tool at 1 mg/L as a water treatment for combating the A. sobria challenge for sustainable aquaculture production.

Interdisciplinary approach to solve unusual mortalities in the European common frog (Rana temporaria) in two high-mountain ponds affected by climate change.

The global decline in amphibian populations is a major environmental issue. Chytridiomycosis, Ranaviruses and the red-leg syndrome have been identified in unusual mortality events. However, these infections do not account for all causes of declining amphibian populations. Moreover, several cases of amphibian mortality are difficult to solve without resorting to an interdisciplinary approach. Two cases of unusual mortality in Rana temporaria occurred at two high-mountain ponds (northwest Italy) in April and May 2021. Water and frog samples were analysed to understand the possible causes responsible for the unusual mortalities. Results of the main physicochemical (pH, conductivity, dissolved oxygen, chemical and biochemical oxygen demand) and nutrient (ammonia/ammonium, nitrite, nitrate, total phosphorus) parameters revealed a good condition of the water quality, with the absence of the main cyanotoxins (microcystins/nodularins). However, unseasonably high spring water temperatures were recorded in both ponds (12.73 °C and 14.21 °C for Frog Pond and Selleries Pond, respectively). Frogs (n = 50; snout-vent length: 7.0-9.8 cm; body mass: 85-123 g) collected from Frog Pond mainly presented bumps on the ventral cavity and dermal ulceration associated with the isolation of Carnobacterium maltaromaticum. On the other hand, frogs (n = 5; snout-vent length: 8.0-9.1 cm; body mass: 87-92 g) from Selleries Pond presented petechiae and dermal ulcerations on the rear limbs associated with the isolation of Aeromonas salmonicida and A. sobria. In both mortality events, the interdisciplinary approach revealed an association between frog mortalities and the isolation of bacteria. Isolated bacteria are considered opportunistic pathogens, and the high values of the water temperature has certainly led a stress on the frogs, favouring the spread of bacteria and the death of the frogs. Further studies are needed to assess the pathophysiological effects of the opportunistic bacteria here isolated, clarifying the interactions between emerging pathogens and climate change.

Exploring the Interactive Effects of Thymol and Thymoquinone: Moving towards an Enhanced Performance, Gross Margin, Immunity and Aeromonas sobria Resistance of Nile Tilapia (Oreochromis niloticus).

Plant-derived bioactive compounds with promising nutritional and therapeutic attributes (phytogenics) are among the top priorities in the aquaculture sector. Therefore, the impact of thymol (Thy) and/or thymoquinone (ThQ) on the growth, immune response antioxidant capacity, and Aeromonas sobria (A. sobria) resistance of Nile tilapia was investigated. Four fish groups were fed a control diet and three basal diets supplemented with 200 mg/kg diet of Thy or ThQ and a blend of both Thy and ThQ at a level of 200 mg/kg diet each. At the end of the feeding trial (12 weeks), the tilapias were challenged intraperitoneally with virulent A. sobria (2.5 × 108 CFU/mL) harboring aerolysin (aero) and hemolysin (hly) genes. The results revealed that tilapias fed diets fortified with a combination of Thy and ThQ displayed significantly enhanced growth rate and feed conversion ratio. Notably, the expression of the genes encoding digestive enzymes (pepsinogen, chymotrypsinogen, α-amylase and lipase) and muscle and intestinal antioxidant enzymes (glutathione peroxidase, catalase and superoxide dismutase) was significantly upregulated in Thy/ThQ-fed fish. An excessive inflammatory response was subsided more prominently in the group administrated Thy/ThQ as supported by the downregulation of il-ß, il-6 and il-8 genes and in contrast, the upregulation of the anti-inflammatory il-10 gene. Remarkably, dietary inclusion of Thy/ThQ augmented the expression of autophagy-related genes, whilst it downregulated that of mtor gene improving the autophagy process. Furthermore, Thy/ThQ protective effect against A. sobria was evidenced via downregulating the expression of its aero and hly virulence genes with higher fish survival rates. Overall, the current study encouraged the inclusion of Thy/ThQ in fish diets to boost their growth rates, promote digestive and antioxidant genes expression, improve their immune responses and provide defense against A. sorbia infections with great economic benefits.

Dietary Salvia officinalis leaves enhances antioxidant-immune-capacity, resistance to Aeromonas sobria challenge, and growth of Cyprinus carpio.

The current perspective is a pioneer to assess the efficacy of Salvia officinalis leave powder (SOLP) on growth, intestinal enzymes, physiological and antioxidant status, immunological response, and gene expression of Common carp (Cyprinus carpio). We also looked into fish resistance after being challenged with Aeromonas sobria, a pathogenic zoonotic bacteria. Fish (N = 120) were fed four different experimental diets in triplicate for 8 weeks. The control diet (SOLP0 - without SOLP); meanwhile, the other three diets included SOLP of 2, 4, and 8 g kg-1 concentrations (SOLP2, SOLP4, and SOLP8), respectively. Findings demonstrated that fish fed SOLP4 and SOLP8 diets had better growth performance and improved digestion by noticeable enhancing lipase and amylase enzymes activity than other groups. Additionally, the antioxidant (superoxide dismutase and glutathione peroxidase) and immune activities (immunoglobulin M, nitric oxide, and antiprotease) clarified a significant increase (p < 0.05) in SOLP4 and SOLP8 groups. Enriched diets with SOLP4 and SOLP8 exhibited better expression of splenic genes (IL-1ß, IL-6, IL-10, TLR-2, and SOD), intestinal genes (Slc26a6) and (PepT1 or Slc15a1), and muscular genes (IGF-1 and SOD), while MSTN was down-regulated. After 8 weeks of the experimental trial, C. carpio challenged by A. sobria exhibited the highest cumulative mortality (66.67%), while SOLP8-dietary intervention showed the best results in enhancing the fish resistance against A. sobria by lessening mortalities to 13.33% followed by SOLP4 diet (20%). The outcomes indicate that the expression of splenic, muscular, and intestinal genes confirm the efficacy of SOLP on enhancing growth, digestion, and immune-antioxidant status, and recommend the potential use of SOLP especially at 4 g kg-1 level as a valuable natural economic diet additive in C. carpio culture for sustaining aquaculture.

Ensayo in vitro de extractos de Lippia graveolens y Lippia alba contra Aeromonas spp aisladas de tilapia (Oreochromis niloticus) / In vitro assay of Lippia graveolens and Lippia alba extracts against Aeromonas spp isolated from tilapia (Oreochromis niloticus) / Ensaio in vitro de extratos da Lippia graveolens e Lippia alba sobre Aeromonas spp isolados de tilápia (Oreochromis niloticus)

Resumen El presente estudio evaluó la actividad antimicrobiana de extractos y aceites esenciales de Lippia graveolens y Lippia alba, frente a nueve cepas de Aeromonas spp., aisladas de Oreochromis niloticus. Los extractos crudos se obtuvieron por la técnica de percolación y a través de la extracción de CO2 supercrítico, mientras que los aceites esenciales se realizaron mediante la técnica de hidrodestilación empleando un equipo Clevenger. La actividad antimicrobiana para cada extracto y aceite esencial se verificó mediante el método de difusión en disco a las 24 horas. El aceite esencial de L. graveolens mostró mayor efectividad para inhibir patógenos bacterianos de peces (100%), en comparación con los extractos. Se obtuvo un diámetro de inhibición que osciló entre 25,20 a 36,94 mm. Los extractos de fluido supercrítico y de la técnica de percolación con acetato de etilo y ciclohexano presentaron la misma efectividad (77,78%). El extracto crudo obtenido con etanol al 95% mostró efecto antimicrobiano limitado (22,22%), presentando el menor halo de inhibición (8,34 y 9,57 mm). Por otro parte, L. alba mostró menor actividad antibacteriana. El aceite esencial inhibió únicamente el 66,67% de las bacterias patógenas, presentando un halo de inhibición que oscila entre 10,68 a 16,29 mm. El resultado de este estudio indica que los aceites esenciales de L. graveolens y L. alba son una alternativa prometedora para el control del crecimiento de Aeromonas spp.

Abstract This study evaluated the antimicrobial activity of Lippia graveolens and Lippia alba extracts and essential oils against nine strains of Aeromonas spp., isolated from Oreochromis niloticus. The crude extracts were obtained by percolation technique and CO2 supercritical fluid extraction, while the essential oils by applying hydro-distillation technique using a Clevenger apparatus. The antimicrobial activity for each extract and essential oils was verified through the disc diffusion method at 24 hours. The essential oil of L. graveolens showed higher effectiveness to inhibit fish bacterial pathogens (100%) than the extracts, revealing an inhibition zone diameter that ranged from 25.20 - 36.94 mm. The extracts from supercritical fluid and from the percolation technique with ethyl acetate and cyclohexane presented the same effectiveness (77.78%). The crude extract obtained with ethanol 95% showed limited antimicrobial effect (22.22%), presenting the smallest inhibition zone (ranged from 8.34 to 9.57 mm). On the other hand, L. alba displayed a lower antibacterial activity, being the essential oil 66.67% effective, presenting an inhibition zone ranging between 10.68 to 16.29 mm. The result of this study indicates that essential oils from both L. graveolens and L. alba offer a promising alternative for the control of Aeromonas spp. growth.

Resumo Neste estudo avaliou-se a atividade antimicrobiana de extratos e óleos essenciais de Lippia graveolens e Lippia alba, contra nove cepas de Aeromonas spp., isoladas de Oreochromis niloticus. Os extratos brutos foram obtidos pela técnica de percolação e por extração supercrítica com CO2, enquanto os óleos essenciais foram obtidos por hidrodestilação em aparelho tipo Clevenger. A atividade antimicrobiana para cada extrato e óleo essencial foi verificada pelo método de disco-difusão em 24 horas. O óleo essencial de L. graveolens apresentou maior eficácia na inibição de patógenos bacterianos em peixes (100%), comparado aos extratos. Obteve-se um diâmetro de halo de inibição que variou entre 25,20 e 36,94 mm. Os extratos do fluido supercrítico e da percolação com acetato de etila e ciclohexano apresentaram a mesma eficácia (77,78%). O extrato bruto obtido com etanol 95% apresentou efeito antimicrobiano limitado (22,22%), apresentando o menor diámetro de halo de inibição (8,34 e 9,57 mm). Por outro lado, L. alba apresentou menor atividade antibacteriana. O óleo essencial inibiu apenas 66,67% das bactérias patogênicas, apresentando um halo de inibição que varia de 10,68 a 16,29 mm de diámetro. Os resultados deste estudo indica que os óleos essenciais de L. graveolens e L. alba são uma alternativa promissora para o controle do crescimento de Aeromonas spp.

Aeromonas sobria Serine Protease Degrades Several Protein Components of Tight Junctions and Assists Bacterial Translocation Across the T84 Monolayer.

Aeromonas sobria is a Gram-negative pathogen that causes food-borne illness. In immunocompromised patients and the elderly, A. sobria opportunistically leads to severe extraintestinal diseases including sepsis, peritonitis, and meningitis. If A. sobria that infects the intestinal tract causes such an extraintestinal infection, the pathogen must pass through the intestinal epithelial barrier. In our earlier study using intestinal cultured cells (T84 cells), we observed that an A. sobria strain with higher A. sobria serine protease (ASP) production caused a marked level of bacterial translocation across the T84 intestinal epithelial monolayer. Herein, we investigated the effect of ASP on tight junctions (TJs) in T84 cells. We observed that ASP acts on TJs and causes the destruction of ZO-1, ZO-2, ZO-3, and claudin-7 (i.e., some of the protein components constituting TJs), especially in the strains with high ASP productivity. Based on the present results together with those of our earlier study, we propose that ASP may cause a disruption of the barrier function of the intestinal epithelium as a whole due to the destruction of TJs (in addition to the destruction of adherens junctions) and that ASP may assist invasion of the pathogens from the intestinal epithelium into deep sites in the human body.

Comparison of Surgical Outcomes and Predictors in Patients with Monomicrobial Necrotizing Fasciitis and Sepsis Caused by Vibrio vulnificus, Aeromonas hydrophila, and Aeromonas sobria.

Background: Monomicrobial necrotizing fasciitis caused by Vibrio vulnificus, Aeromonas hydrophila, and Aeromonas sobria are often associated with high mortality rates. The purpose of this study was to compare the independent predictors related to outcomes between Vibrio vulnificus and Aeromonas species necrotizing fasciitis. Patients and Methods: Monomicrobial necrotizing fasciitis caused by Vibrio vulnificus (60 patients) and Aeromonas species (31 patients) over an 11-year period were reviewed retrospectively. Differences in mortality, patient characteristics, clinical presentations, and laboratory data were compared between the Vibrio vulnificus and Aeromonas species groups, and between the death and the survival subgroups of patients with Aeromonas species. Results: Six patients in the Vibrio vulnificus group (10%) and 11 in the Aeromonas species group (32.3%) died. Fifty-nine patents had bacteremia and 16 patients died (27.1%). Patients who had Vibrio vulnificus had a higher incidence of bacteremia. The patients who had Aeromonas species presenting with bacteremia were significantly associated with death. The death subgroup of patients with Aeromonas necrotizing fasciitis had a higher incidence of bacteremia, higher counts of banded leukocytes, lower platelet counts, lower total lymphocyte counts, and lower serum albumin level than the survival subgroup. Conclusions: Monomicrobial necrotizing fasciitis caused by Aeromonas species was characterized by more fulminating and higher mortality than that of Vibrio vulnificus, even after early fasciotomy and third-generation cephalosporin antibiotic therapy. Those risk factors, such as bacteremia, shock, lower platelet counts, lower albumin levels, and antibiotic resistance were associated with mortality, which should alert clinicians to pay more attention to and aggressively treat those patients with Aeromonas and Vibrio necrotizing fasciitis.

A Case of Severe Aeromonas Bacteremia with Necrotizing Fasciitis of Lower Limb and Fournier ' s Gangrene in a Post-Allogenic Unrelated Hematopoietic Stem Cell Transplant Recipient.

Bikram DasBackground Aeromonas is a water-dwelling Gram-negative bacillus primarily associated with gastrointestinal tract diseases. Aeromonas sobria causing gastroenteritis has been reported in India. In immunocompromised host, Aeromonas sobria can also present with severe necrotizing skin and soft tissue infection with a high mortality rate. We report a case of Aeromonas sobria sepsis with skin and soft tissue infection in the background of immunosuppression. Case Presentation Fifty-year-old male who underwent an unrelated donor peripheral stem cell transplant for relapsed pre-B acute lymphoblastic leukemia in complete clinical remission on graft versus host disease prophylaxis, post-white blood cell engraftment presented with acute onset lethargy, lower limb pain without fever, or any skin changes initially. He rapidly worsened clinically over few days and developed sepsis, multiorgan dysfunction with the appearance of erythema and blister over the lower limb, and Fournier's gangrene of scrotum. He was found to have Aeromonas sobria bacteremia with isolated resistance to carbapenems while sensitive to all other classes of antibiotics. Despite appropriate antibiotic therapy and supportive measures, he succumbed to death for this invasive bacterial disease. Conclusion Aeromonas should be considered a cause of sepsis in immunosuppressed hosts, especially those with hematological malignancy presenting with necrotizing skin and soft tissue infection. Considering the virulence of this pathogen, despite the very susceptible antibiogram, such patients must be managed aggressively. Early recognition of the disease with a combination of medical and surgical management might help to improve the outcome.

A fatal combination of disseminated strongyloidiasis with two bacterial infections in an immunocompromised host.

INTRODUCTION: Strongyloides stercoralis is an intestinal nematode that is endemic in tropical countries. It can have a variable presentation ranging from asymptomatic eosinophilia in immunocompetent hosts to disseminated disease with sepsis in immunocompromised hosts. CASE REPORT: We report a case of chronic diarrhoea and decreased appetite in a 53-year-old man. He was a chronic alcoholic with diabetes, hypertension and dyslipidaemia and had earlier been treated for pulmonary tuberculosis. He was treated symptomatically for loose stools at a primary health care facility without relief. Following referral to our tertiary care centre, microscopic examination of the stool showed numerous larvae and a few eggs of Strongyloides stercoralis. Additionally, Aeromonas sobria was isolated from stool culture. The patient was discharged following improvement with a combination therapy of ivermectin, albendazole and ciprofloxacin. However, within 3 days, he was readmitted and succumbed to Escherichia coli sepsis. CONCLUSION: Strongyloidiasis can be diagnosed easily using a very simple but often neglected investigation, namely stool microscopy. This provides an early diagnosis, based on which prompt treatment with the appropriate antihelminthics can be started, thereby reducing the probability of disseminated infection. Disseminated strongyloidiasis is a medical emergency with a poor prognosis, especially in an immunocompromised state. Such patients should be treated aggressively with antihelminthics. They must be monitored for sufficient duration in the hospital for early signs of complication. Their discharge from hospital should be planned based on a negative stool microscopy report in addition to clinical improvement, so as to decrease the mortality reported for both untreated and treated individuals.

Aeromonas sobria Induces Proinflammatory Cytokines Production in Mouse Macrophages via Activating NLRP3 Inflammasome Signaling Pathways.

Aeromonas sobria, a common conditional pathogenic bacteria, is widely distributed in the environment and causes gastroenteritis in humans or septicemia in fish. Of all Aeromonas species, A. sobria is the most frequently isolated from human infections especially in immunocompromised subjects. Innate immunity is the first protection system of organism to resist non-specific pathogens invasion; however, the immune response process of hosts against A. sobria infection re\mains unexplored. The present study established an A. sobria infection model using primary mouse peritoneal macrophages (PMφs). The adherence and cytotoxicity of A. sobria on PMφs were determined by May-Grünwald Giemsa staining and LDH release measurement. Pro-inflammatory cytokine expression levels were measured using qPCR, western blotting, and ELISA methods. We also investigated the levels of ASC oligomerization and determined the roles of active caspase-1 in IL-1ß secretion through inhibition assays and explored the activated pattern recognition receptors through immunofluorescence. We further elucidated the roles of activated inflammasome in regulating the host's inflammatory response through inhibition combined with ELISA assays. Our results showed that A. sobria induced lytic cell death and LDH release, whereas it had no adhesive properties on PMφs. A. sobria triggered various proinflammatory cytokine transcription level upregulation, and IL-1ß occupied the highest levels. The pro-IL-1ß protein expression levels increased in a dose-dependent manner with MOI ranging from 1 to 100. This process was regulated by ASC-dependent inflammasome, which cleavage pro-IL-1ß into active IL-1ß p17 with activated caspase-1 p20. Meanwhile, the expression levels of NLRP3 receptor significantly increased, location analysis revealed puncta-like surrounding nuclear, and inhibition of NLRP3 inflammasome downregulated caspase-1 activation and IL-1ß secretion. Blocking of NLRP3 inflammasome activation through K+ efflux and cathepsin B or caspase approaches downregulated A. sobria-induced proinflammatory cytokine production. Overall, these data indicated that A. sobria induced proinflammatory cytokine production in PMφs through activating NLRP3 inflammasome signaling pathways.

Aeromonas sobria regulates proinflammatory immune response in mouse macrophages via activating the MAPK, AKT, and NF-κB pathways.

Aeromonas sobria, a Gram-negative bacterium that can colonize both humans and animals, is found in a variety of environments, including water, seafood, meat, and vegetables (Cahill, 1990; Galindo et al., 2004; Song et al., 2019). Aeromonas spp. are conditionally pathogenic bacteria in aquaculture, which can rapidly proliferate, causing disease and even death in fish, especially when the environment is degraded (Neamat-Allah et al., 2020, 2021a, 2021b). In developing countries, Aeromonas spp. have been associated with a wide spectrum of infections in humans, including gastroenteritis, wound infections, septicemia, and lung infections (San Joaquin and Pickett, 1988; Wang et al., 2009; Su et al., 2013). Infections caused by Aeromonas spp. are usually more severe in immunocompromised individuals (Miyamoto et al., 2017). The presence of a plasmid encoding a ß|-lactamase in A. sobria that confers resistance to ß-lactam antibiotics poses a huge challenge to the treatment of diseases caused by this microorganism (Lim and Hong, 2020). Consequently, an in-depth understanding of the interaction between A. sobria and its hosts is urgently required to enable the development of effective strategies for the treatment of A. sobria infections.

Myo-Inositol Restores Tilapia's Ability Against Infection by Aeromonas sobria in Higher Water Temperature.

Bacterial infection presents severe challenge to tilapia farming, which is largely influenced by water temperature. However, how water temperature determines tilapias' survival to infection is not well understood. Here, we address this issue from the perspective of metabolic state. Tilapias were more susceptible to Aeromonas sobria infection at 33°C than at 18°C, which is associated with differential metabolism of the fish. Compared to the metabolome of tilapia at 18°C, the metabolome at 33°C was characterized with increased an tricarboxylic acid cycle and a reduced level of myo-inositol which represent the most impactful pathway and crucial biomarker, respectively. These alterations were accompanied with the elevated transcriptional level of 10 innate immune genes with infection time, where il-1b, il-6, il-8, and il-10 exhibited a higher expression at 33°C than at 18°C and was attenuated by exogenous myo-inositol in both groups. Interestingly, exogenous myo-inositol inactivated the elevated TCA cycle via inhibiting the enzymatic activity of succinate dehydrogenase and malate dehydrogenase. Thus, tilapias showed a higher survival ability at 33°C. Our study reveals a previously unknown relationship among water temperature, metabolic state, and innate immunity and establishes a novel approach to eliminate bacterial pathogens in tilapia at higher water temperature.

Envenoming by king cobras (Ophiophagus hannah) in Vietnam with cardiac complications and necrotizing fasciitis.

King Cobra (Ophiophagus hannah) bite is well-known for its potentially fatal neurotoxicity. However, fatalities still occur, despite specific antivenom and respiratory support. Cardiovascular disturbances, which have attracted little attention in published reports of O. hannah envenoming, could contribute to fatality. We present two cases of confirmed O. hannah envenoming in Southern Vietnam in which there were cardiac abnormalities including arrhythmias and electrocardiographic changes, as well as elevated markers of myocardial damage. Cardiac pacing was required. One patient developed critical multi-organ dysfunctions partly explained by extensive necrotizing fasciitis/myositis originating from an Aeromonas sobria wound infection. This resulted in rhabdomyolysis, disseminated intravascular coagulation and acute kidney injury. Specific antivenom reversed neurotoxic effects of envenoming. Additional therapeutic interventions included antibiotics, surgical debridement, continuous renal replacement therapy and therapeutic plasma exchange. Both patients eventually made full recoveries. Apart from the critical problem of rapidly evolving and severe neurotoxicity, our case reports also emphasises the risk of cardiotoxic envenoming, and the complications of an overwhelming secondary bacterial wound infection. We suggest a practical approach to diagnosis and management.

Immunological Responses and the Antioxidant Status in African Catfish (Clarias gariepinus) Following Replacement of Dietary Fish Meal with Plant Protein.

African catfish (Clarias gariepinus) were subjected to a 30-day feeding trial to determine the appropriateness of using plant protein (PP) (soybean and sunflower meal) as a fishmeal (FM) replacement in the diet and its effects on immune status, antioxidant activity, pituitary adenylate cyclase-activating polypeptide (PACAP) gene expression, and disease resistance. A total of 150 C. gariepinus (51.01 ± 0.34 g) were randomly distributed among five groups in triplicate. Five experimental diets were formulated to replace 0 (control), 33.5, 50, 66.5, and 100% FM with soybean and sunflower meal to form the experimental diets (R0, R33.5, R50, R66.5, and R100, respectively). After 30 days, the diet containing PP for FM had no significant impact on total, and differential leukocyte counts determined at the end of the feeding period. The total globulin concentration showed significantly greater differences in the following order R0 > R33.5 > R50 > R66.5 > R100. The R0 group had the highest concentration of serum γ-globulin, while R100 had the lowest concentration. The antioxidant status complements 3 (C3), lysozyme activity (LYZ), and antiprotease activity were not significantly different between R0, R33.5, and R50 groups, while they were significantly lower in R100. The serum nitric oxide activity (NO) exhibited significantly greater differences in the following order R0 > R33.5 > R50 > R66.5 > R100. PACAP was significantly higher in the R33.5 group. The highest cumulative mortality caused by Aeromonas sobria was recorded in the R100 group (60%) and the lowest in the R0 group (30%). In conclusion, the results indicate that the immunological responses and antioxidant status of C. gariepinus were not affected when they consumed a diet with FM replaced by up to 50% with PP (SBM and SFM) with methionine and lysine supplementation, but total globulin, NO, and cumulative mortality were impaired with a diet containing a 100% FM replacement.

Aeromonas sobria necrotizing fasciitis complicating neonatal sclerosing cholangitis.

Aeromonas species can cause gastroenteritis, soft tissue infections, hepatobiliary tract and other infections. While most reported soft tissue infections have been attributed to Aeromonas hydrophila, we report a case of necrotizing fasciitis caused by Aeromonas veronii biovar sobria. Prompt and accurate diagnosis cum treatment could help save children with Aeromonas soft tissue infections from developing a bacteremia with extended morbidity and mortality.

Characterization of the Pelodiscus sinensis polymeric immunoglobulin receptor (P. sinensis pIgR) and its response to LPS and Aeromonas sobria.

The polymeric immunoglobulin receptor (pIgR) is one of the most vital components of mucosal immunity that plays a pivotal role in mediating transcytosis of polymeric immunoglobulin (pIg) on epithelial surfaces for protection against invading pathogens. Herein, we cloned the full-length cDNA of Pelodiscus sinensis pIgR, designated as P. sinensis pIgR, made of an open reading frame (ORF) of 1848 bp, molecular weight of 68.2 kDa and estimated isoelectric point of 7.00. The deduced P. sinensis pIgR sequence had a leader peptide, extracellular region containing four immunoglobulin-like domains (Ig like domains), transmembrane and intracellular regions comparable with other vertebrates. P. sinensis pIgR contained four Ig like domains that corresponded with mammalian D1, D3, D4 and D5 similar with reptile and avian Ig like domains. It had 40 potential phosphorylation sites, four putative N-glycosylation sites and several motifs resembling mammalian pIgR motifs. Phylogenetic analysis showed a close relationship between P. sinensis pIgR with avian and reptile pIgRs. P. sinensis pIgR basal levels were higher in the esophagus, small intestine and intestinnum crissum than in other organs of health turtles. Intragastric delivery of LPS and Aeromonassobria led to significant upregulation of P. sinensis pIgR in tissues of the gastrointestinal tract. A polyclonal anti- P. sinensis pIgR antibody produced in rabbit reacted with the recombinant P. sinensis pIgR protein expressed in Escherichia coli in Western blot. These studies demonstrate the existence and immune response of P. sinensis pIgR to stimulation in mucosal organs in Chinese soft-shelled turtles.

Motile aeromonad septicemia (MAS) at Cyprinus carpio L, 1758 (Actinopterygii: Cyprinidae) in Lake Tödürge (Sivas/Turkey) / [Septicemia móvel por aeromonada (SMA) em Cyprinus carpio L, 1758 (Actinopterygii: Cyprinidae) no lago Tödürge (Sivas, Turquia)]

In this study, fish's morphologic and anatomic lesions caused by motile aeromonad septicemia (MAS) depending on environmental stress in carp, Cyprinus carpio population living in Lake Tödürge were identified. Various morphological and anatomical deformations and lesions were observed in the body of approximately 17% (252 fish specimens) of a total of 1488 carp samples. Bacteria are grown from all wipe samples. Bacterial colonies have a gray-white appearance with round, convex and smooth edges. 15-20 cfu colonies were observed in each aerop culture. As a result of analysis of wet wipe samples from infected fish's skin, gill, kidney and liver, it is determined that the bacteria which causes septicemia is Aeromonas sobria from the Aeromonadaceae family (with 99.2% confidence value). No bacteria were grown in cultures except A. sobria. Some symptoms of the infection are inflammation on different parts of the fish bodies, eruption on skin and scales, dermal necrosis, degeneration at soft rays of the fins, exophthalmos, and purulent liquid accumulation in the abdominal cavity, etc. Infected fish were most commonly encountered in July and August (water temperature above 20ºC), the lowest in October and November (water temperature below 10ºC).(AU)

Neste estudo, foram identificadas lesões morfológicas e anatômicas causadas por septicemia móvel por aeromônios (MPA), dependendo do estresse ambiental da carpa, a população de Cyprinus carpio que vive no lago Tödürge foi identificada. Várias deformações e lesões morfológicas e anatômicas foram observadas no corpo de aproximadamente 17% (252 amostras de peixes) de um total de 1488 amostras de carpa. As bactérias são cultivadas a partir de todas as amostras de limpeza. As colônias bacterianas têm uma aparência branco-acinzentada, com bordas arredondadas, convexas e lisas. Foram observadas 15-20 colônias de UFC em cada cultura de aerop. Como resultado da análise de amostras de lenços umedecidos da pele, brânquias, rins e fígado de peixes infectados, é determinado que a bactéria que causa a septicemia é a Aeromonas sobria, da família Aeromonadaceae (com valor de confiança de 99,2%). Nenhuma bactéria foi cultivada em culturas, exceto A. sobria. Alguns sintomas da infecção são inflamação em diferentes partes dos corpos dos peixes, erupção na pele e escamas, necrose dérmica, degeneração aos raios moles das barbatanas, exoftalmia e acúmulo de líquido purulento na cavidade abdominal, entre outros. Os peixes infectados eram encontrados com maior frequência em julho e agosto (temperatura da água acima de 20ºC), e eram menos comumente encontrados em outubro e novembro (temperatura da água abaixo de 10ºC).(AU)