Efficacy of ulvan on immune response and immuno-antioxidant gene modulation in Labeo rohita against columnaris disease.

This study reports the effect of ulvan enriched diet on the influence of growth, changes in hemato-biochemical indices, improvement of antioxidant system, enhancement of innate-adaptive immunity and modification of immuno-antioxidant genes expression in Labeo rohita against Flavobacterium columnaris. The weight gain (WG) was significantly high (P > 0.05) in unchallenged normal and challenged fish fed with diets enriched with 25 and 50 mg kg-1 ulvan; the FCR was better (P > 0.05) when fed with 50 mg kg-1 enriched diet. In normal fish fed with or without ulvan supplementation was noted 100% survival rate (SR). In both groups, the red blood cell (RBC) and while blood cell (WBC) counts increased significantly (P > 0.05) when fed with 50 mg kg-1 ulvan diet whereas the hemoglobin (Hb) level increased significantly on being fed with 25 and 50 mg kg-1 ulvan diets. The SOD activity was enhanced significantly in both groups fed with any dose of ulvan diets whereas the MDA and GPx activity increased only with 25 and 50 mg kg-1 ulvan diets. The phagocytic (PC) activity significantly increased with any enriched diet and control diet groups while the respiratory burst (RB) activity increased only with 50 mg kg-1 ulvan diet. The alternate complement pathway (ACP), activity of lysozyme (Lyz), and immunoglobuline M (IgM) were better in both groups fed with 50 mg kg-1 ulvan diet. The SOD and GPx antioxidant gene expression were significantly high in both groups fed with any ulvan diet while the Nrf2 gene expression was high with 50 mg kg-1 ulvan diet. The IL-1ß, TNFα, hepcidin, Lyz, and IgM cytokines or proteins mRNA expression were significant in both groups fed with all ulvan supplement diet whereas the ß-2M expression was significant only with 50 mg kg-1 ulvan diet. The present research indicates that both L. rohita groups fed with 50 mg kg-1 ulvan diet significantly improved growth, antioxidant system, immune defense system, and immuno-antioxidant related gene expression against F. columnaris.

Tenacibaculosis induction in the Senegalese sole (Solea senegalensis) and studies of Tenacibaculum maritimum survival against host mucus and plasma.

Tenacibaculum maritimum, the aetiological agent for marine tenacibaculosis, is one of the most significant pathogens that threaten Senegalese sole, Solea senegalensis (Kaup), aquaculture. Because no immersion challenge with T. maritimum has been reported previously for this flatfish species, this study aimed to optimize bacterial yields as well as to establish a challenge model for tenacibaculosis induction. Several approaches were performed to optimize bacterial culture conditions, including treatment with non-ionic surfactants, detergents, cellulase hydrolysis and strong shaking. A prolonged bath challenge was performed for 24 h under two different temperatures, 16 and 23 °C. Moreover, mucus and plasma bactericidal activities against T. maritimum were also assessed. Culturing bacteria with strong shaking and continuous shaking provided suitable culture conditions to obtain higher bacterial yields without aggregation and fluctuation, contrary to most other treatments that showed a huge amount of bacterial aggregates. A prolonged bath method for 24 h, without skin or gill scarification, was considered suitable for disease induction with high mortality rates. Moreover, data regarding mucus and plasma bactericidal activities suggested that there is a lack of host innate immune response against T. maritimum or that this particular pathogen presents evading strategies against Senegalese sole.

In vivo activity of cefquinome against Riemerella anatipestifer using the pericarditis model in the duck.

Cefquinome is a fourth-generation cephalosporin with broad-spectrum antibacterial activity, including activity against enteric gram-negative bacilli such as Riemerella anatipestifer. The pericarditis model was used to examine the pharmacodynamic characteristics of cefquinome against R. anatipestifer. Serum levels of cefquinome following the administration of different doses were determined by LC-MS/MS. Ducks with ca. 10(6) CFU/mL at the initiation of therapy were treated with cefquinome at doses that ranged from 0.0156 to 2 mg/kg of body weight/day (in 3, 6, 12, or 24 divided doses) for 24 h. The percentage of a 24-h dosing interval that the unbound serum cefquinome concentrations exceeded the MIC (fT > MIC) were the pharmacokinetic (PK)-pharmacodynamic (PD) parameter that best correlated with efficacy (R(2) 86.3% for R. anatipestifer, compared with 58.9% for the area under the concentration-time curve/MIC and 10.6% for peak/MIC). A sigmoid Emax model was used to estimate the magnitudes of the %fT > MIC associated with net bacterial stasis, a 1-log10 CFU reduction from baseline, and a 2-log10 CFU reduction from baseline; the corresponding values were (22.5 ± 1.3) %, (35.2 ± 4.5) %, and (42.4 ± 2.7) %. These data showed that treatment with cefquinome results in marked antibacterial effects in vivo against R. anatipestifer and that the host's immunity may also play a key role in the anti-infective therapy process.

Evaluation of the immune response in rainbow trout fry, Oncorhynchus mykiss (Walbaum), after waterborne exposure to Flavobacterium psychrophilum and/or hydrogen peroxide.

The immune response in rainbow trout fry against Flavobacterium psychrophilum was elucidated using an immersion-based challenge with or without prior exposure to hydrogen peroxide (H2O2). Samples were taken from the head kidney 4, 48, 125 and 192 h after immersion, and the regulation of several genes was examined. Bacterial load was assessed based on the presence of 16S rRNA and correlated with gene expression, and the levels of specific antibodies in the blood were measured 50 days post-infection. Separately, both H2O2 and F. psychrophilum influenced gene expression, and pre-treatment with H2O2 influenced the response to infection with F. psychrophilum. Pre-treatment with H2O2 also affected correlation between gene regulation and pathogen load for several genes. A delay in antibody production in H2O2-treated fish in the early phase of infection was indicated, but H2O2 exposure did not affect antibody levels 50 days post-infection. An increasing amount of F. psychrophilum 16S rRNA was found in the head kidneys of infected fish pre-treated with H2O2 relative to the F. psychrophilum group. The results show that a single pre-treatment with H2O2 impairs the response against F. psychrophilum and may intensify infection.

Research Note: Serological investigation of Ornithobacterium rhinotracheale infection in China.

Ornithobacterium rhinotracheale (ORT) has been associated with avian respiratory disease. On coinfection with other pathogens, ORT can cause serious health problems in avian species, leading to financial losses. To monitor the serologic prevalence of ORT in chicken flocks in China, 1,280 sera were collected to determine ORT antibodies among 64 flocks from 15 provinces of China using a commercial ELISA kit. The overall seroprevalence of ORT among the birds tested was 44.06%. In younger chickens, the serum positive rate was lower than that in older chickens, and with increased age, the serum positive rates increased. Older chickens had not only higher positive rates but also higher antibody levels. These data indicated that ORT infections were common in China. Because an ORT vaccine is currently not available, good disease management and biosecurity measures are required for effective disease control.

Development and evaluation of an experimental model of cutaneous columnaris disease in koi Cyprinus carpio.

A reproducible, experimental model of columnaris disease was developed to study the pathogenesis of cutaneous disease associated with Flavobacterium columnare infection in koi (Cyprinus carpio). In experimental infections, lesions were usually restricted to skin and fins; gill necrosis was not a consistent finding. Cytologic and histopathologic examinations provided a presumptive diagnosis of columnaris disease. Specific detection of F. columnare was done using the polymerase chain reaction and DNA in situ hybridization (ISH). Polymerase chain reaction allowed the detection of F. columnare in fresh biological material and in formalin-fixed, paraffin-embedded tissues. The DNA ISH technique allowed the identification and localization of F. columnare in formalin-fixed, paraffin-embedded tissues. Using these molecular techniques, F. columnare was readily detected in skin specimens from infected fish; however, the bacterium was infrequently detected in specimens of liver, kidney, and spleen. These observations suggest that columnaris disease generally presents as a cutaneous disease that is unassociated with systemic infection in koi. Hematologic studies indicated that most infected koi developed microcytic, normochromic, nonregenerative anemia and leukopenia characterized by lymphopenia, mild neutrophilia, and monocytosis. Biochemical changes in diseased fish included significant hyperglycemia, hyponatremia, and hypochloridemia.

Ornithobacterium rhinotracheale has neuraminidase activity causing desialylation of chicken and turkey serum and tracheal mucus glycoproteins.

Neuraminidases (sialidases) are virulence factors of several poultry pathogens. Ornithobacterium rhinotracheale is a well known poultry pathogen causing respiratory disease in chickens and turkeys all over the world. We investigated whether O. rhinotracheale has neuraminidase enzymatic activity (NEAC). We tested NEAC in 47 O. rhinotracheale strains isolated from turkeys and chickens in eight countries. All strains showed relatively strong NEAC and considerable levels of NEAC were detected also in "cell-free supernatants" of their pelleted cells. Zymography using neuraminidase-specific chromogenic substrate indicated that a protein with molecular mass of ~40kDa and isoelectric point (pI) of ~8.0 is a putative neuraminidase of O. rhinotracheale. Notably, the genome of the type strain of O. rhinotracheale, DSM 15997 contains a gene (Ornrh_1957) encoding a putative neuraminidase with such Mw (39.5 kDa) and pI (8.5). We sequenced a corresponding genomic region of 20 O. rhinotracheale strains and found five distinct types of the neuraminidase gene (termed nanO) sequences. Most diversified nanO sequence was found in two strains isolated from chickens in Hungary in 1995. Their nanO sequences differ from that of the type strain (LMG 9086(T)) in 27 nucleotides. O. rhinotracheale neuraminidase showed capacity to cleave sialic acid from chicken and turkey glycoproteins. It cleaved sialic acid from SAα(2-6)gal moiety of their serum proteins, including immunoglobulin G (IgG) and transferrin. O. rhinotracheale also desialylated chicken and turkey tracheal mucus glycoprotens with SAα(2-3)gal moieties. This study provides the first evidence that O. rhinotracheale has neuraminidase which can desialylate glycoproteins of its natural hosts.

Decreased phosphorylation of platelet vasodilator-stimulated phosphoprotein in periodontitis--a role of periodontal pathogens.

INTRODUCTION: Epidemiological studies indicate an association between periodontitis and cardiovascular disease, but the underlying mechanisms are poorly understood. Vasodilator-stimulated phosphoprotein (VASP) in its phosphorylated form represents a regulator of platelet function and an indicator for the sensitivity of platelets towards physiologically relevant antagonists of platelet function. As platelets and their activation state play a central role in the development of cardiovascular disease, this study aimed to investigate the influence of periodontal disease and periodontal pathogens on intraplatelet VASP-phosphorylation and platelet function. MATERIAL AND METHODS: Besides several markers of platelet activation, basal and PGE(1) induced intracellular VASP-phosphorylation were determined in platelets of periodontitis patients (n = 26) and healthy donors (n = 19). Furthermore, platelets from healthy donors were incubated with distinct periodontal pathogens and basal and PGE(1) induced VASP-phosphorylation was determined. RESULTS: Compared to controls, platelets of periodontitis patients showed a significant decrease in basal and PGE(1) induced VASP-phosphorylation. VASP-phosphorylation in platelets from periodontitis patients positive for Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis or Tannerella forsythia was significantly decreased compared to patients that were negative for these bacteria. Furthermore, VASP-phosphorylation in platelets isolated from healthy donors was affected by incubation with these periodontal pathogens. CONCLUSIONS: Our results provide evidence that periodontitis interferes with VASP-phosphorylation in human platelets, presumably as a consequence of a direct effect of periodontitis-associated bacteria. Decreased basal and PGE(1) induced VASP-phosphorylation might represent a mechanism responsible for enhanced platelet activation in periodontitis.