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The profitability of the olive flounder (Paralichthys olivaceus) aquaculture industry in Korea depends on high production and maintenance of flesh quality, as consumers prefer to eat raw flounders from aquaria and relish the raw muscles as ‘sashimi’. For sustaining high production, easy-to-deliver and efficient vaccination strategies against serious pathogens, such as viral hemorrhagic septicemia virus (VHSV), is very important as it cause considerable losses to the industry. Whereas, a safe and non-invasive vaccine formulation that is free from unacceptable side-effects and does not devalue the fish is needed to maintain flesh quality. We previously developed a squalene–aluminium hydroxide (Sq + Al) adjuvanted VHSV vaccine that conferred moderate to high protection in flounder, without causing any side effects when administered through the intraperitoneal (IP) injection route. However, farmers often demand intramuscular (IM) injection vaccines as they are relatively easy to administer in small fishes. Therefore, we administered the developed vaccine via IP and IM routes and investigated the safety and persistency of the vaccine at the injection site. In addition, we conducted a comparative analysis of vaccine efficacy and serum antibody response. The clinical and histological observation of the IM and IP groups showed that our vaccine remained persistence at the injection sites for 10–17 weeks post vaccination (wpv), without causing any adverse effects to the fish. The relative percentage of survival were 100% and 71.4% for the IP group and 88.9% and 92.3% for the IM group at 3 and 17 wpv, respectively. Thus, considering the persistency period (24 wpv) and both short and long-term efficacy of our vaccine, the present study offers an option to flounder farmers in selecting either IM or IP delivery strategy according to their cultured fish size and harvesting schedule — IM vaccination for small-sized fish and IP vaccination for table-sized fish.

Trauma remains a major public health problem worldwide, marked as the fourth leading cause of death among all diseases. Trauma patients who survived at initial stages in the Emergency Department (ED), have significantly higher chances of mortality due to sepsis associated complications in the ICU at the later stage. There is paucity of literature regarding the role of circulating monocytes subsets and development of sepsis complications following trauma haemorrhagic shock (THS). The study was conducted to investigate the circulating level of monocyte subsets (Classical, Inflammatory, and Patrolling) and its functions in patients with acute post-traumatic sepsis. A total 72, THS patients and 30 age matched healthy controls were recruited. Blood samples were collected at different time points on days 1, 7, and 14 to measure the serum levels of cytokines by Cytometric bead assay (CBA), for the immunophenotyping of monocytes subsets, and also for the cell sorting of monocytes subsets for the functional studies. The circulating levels of monocytes subsets were found to be significantly differs among THS patients, who developed sepsis when compared with others who did not. The levels of patrolling monocytes were elevated in THS patients who developed sepsis and showed negative correlation with Sequential organ failure assessment (SOFA) score on days 7 and 14. Classical monocytes responded strongly to bacterial TLR-agonist (LPS) and produced anti-inflammatory cytokines, whereas patrolling monocytes responded with viral TLR agonist TLR-7/8 (R848) and produced inflammatory cytokines in post-traumatic sepsis patients. In conclusion, this study shows disparity in the behaviour of monocytes subsets in patients with acute post-traumatic sepsis.

Vibrio strains (n=5) were isolated from a pooled liver samples of golden mahseer, Torputitora (n=10) showing symptoms of hemorrhagic septicemia in cultured conditions. From biochemical analysis, partial PCR amplification of 16S rRNA genes (~1400bp) and sequencing, the isolates were identified as Vibrio anguillarum. One of the 5 isolates was designated as a strain number MHJL 248 and submitted to the NCBI (MN900589). The phylogenetic analysis of the V. anguillarum MHJL 248 showed >99% similarity with V. anguillarum strain HA (KF413427). V. anguillarum MHJL 248 was positive to arginine, oxidase, catalase, string test, ONPG, siderophore, sensitive to O/129 vibriostatic disc (10 and 150 pg) grown on medium containing varying NaCl concentrations (0.5% to 2.0%), showing intermediate resistance to ampicillin (10 pg) and a-hemolytic on 5% sheep blood agar plate. The major histopathological changes recorded in the kidney, liver, gill and intestine collected from natural infection were cellular degeneration, necrosis, cytoplasmic vacuolation, degeneration of glomerulus, dilation and increasing Bowman's space and glomerular contraction. Vibrio anguillarum MHJL 248 was found resistant to cefoxitin (30 pg), ciprofloxacin (5 pg), imipenem (10 pg), levofloxacin (5 pg) and nalidixic acid (30 pg). The minimum inhibitory concentration of oxytetracycline, erythromycin and florfenicol against V. anguillarum MHJL 248 were 16, 0.5 and 0.125 pg mL−1, respectively. Transmission electron microscopy (negatively stained) of V. anguillarum MHJL 248 demonstrated the presence of monotrichous polar flagellum and capsular structure that can be correlated to the pathogenicity of the bacterium. This is the first case of a new host (T. putitora) record of V. anguillarum infection in coldwater aquaculture practices in India. Seeing the severity of infection of V. anguillarum in aquaculture worldwide, further study needs to be focused on the development of efficient measures to control its infection in coldwater aquaculture.

Background and Aim: Respiratory infection due to Mannheimia haemolytica and Pasteurella multocida are responsible for huge economic losses in livestock sector globally and it is poorly understood in ovine population. The study aimed to investigate and characterize M. haemolytica and P. multocida from infected and healthy sheep to rule out the involvement of these bacteria in the disease. Materials and Methods: A total of 374 healthy and infected sheep samples were processed for isolation, direct detection by multiplex PCR (mPCR), and antibiotic susceptibility testing by phenotypic and genotypic methods. Results: Overall, 55 Pasteurella isolates (27 [7.2%] M. haemolytica and 28 [7.4%] P. multocida) were recovered and identified by bacteriological tests and species-specific PCR assays. Significant correlation between the detection of M. haemolytica (66.6%) with disease condition and P. multocida (19.1%) exclusively from infected sheep was recorded by mPCR. In vitro antibiotic susceptibility testing of 55 isolates revealed higher multidrug resistance in M. haemolytica (25.9%) than P. multocida (7.1%) isolates. Descending resistance towards penicillin (63.6%), oxytetracycline (23.6%), streptomycin (14.5%), and gentamicin (12.7%) and absolute sensitivity towards chloramphenicol were observed in both the pathogens. The antibiotic resistance genes such as strA (32.7%) and sul2 (32.7%) associated with streptomycin and sulfonamide resistance, respectively, were detected in the isolates. Conclusion: The study revealed the significant involvement of M. haemolytica together with P. multocida in ovine respiratory infection and is probably responsible for frequent disease outbreaks even after vaccination against hemorrhagic septicemia in sheep population of Karnataka, southern province of India.

Vaccine candidate tests were carried out in the Laboratory of Microbiology, Faculty of Agriculture ULM; and the Laboratory of Molecular Genetics, Faculty of Science and Technology Airlangga University. The field test was conducted in Tampakang Village, Hulu Sungai Utara district and Bati-Bati village, Tanah Laut district that was including sterility test through vaccine mediated Brain Heart Infusion (BHI) inoculation. Safety testing was applied in experimental animals (mice and swamp buffaloes). Serum was tested serologically using Enzyme Linked Immuno Sorbent Assay (ELISA). Vaccine safety test showed 100% of mice and buffalo are surviving without clinical symptoms according to the characteristics of hemorrhagic septicemia disease. ELISA test indicated vaccinations increase antibody production, namely the Optical Density (OD) ƛ 450 ƞm 0.292 before vaccination to be 0.748 and 1.576 after vaccination and to be 1.821 after boosted. Based on sterilization test, safety test, and vaccine potency test, the vaccine candidate from cell wall proteins of P. multocida local isolate can be used to prevent hemorrhagic septicemia disease that infect swamp buffalo both on laboratory and field experiment.

This paper describes the first documented outbreak of haemorrhagic septicaemia (HS) caused by Pasteurella multocida type B in cattle in Spain. This acute, highly fatal septicaemia causes major economic losses in cattle and buffaloes in many areas of Asia and Africa. In other species and in European countries it is an infrequently reported disease. Acute septicaemic pasteurellosis occurred in a free-range farm of 150 cattle and 70 beef calves in Southern Spain. Twenty-one calves and one cow were affected, of which three calves and the adult cow died. Postmortem examination revealed characteristic oedema in the ventral area of the neck and the brisket region, and widespread haemorrhages in all organs. Pure cultures of P. multocida were obtained from all tissues and organs studied. The aetiological agent was further confirmed by molecular and biochemical analysis as P. multocida capsular type B, biovar 3. Although the source of infection could not be determined, wildlife may play an important role. The use of tulathromycin in the initial stage of the disease might be related to the low morbidity and mortality of this outbreak. After using an autogenous vaccine no more cases of HS were observed.

Para-veterinary systems have arisen in numerous developing countries to address the low capacity of national veterinary services in meeting livestock health demands of mainly smallholder farmers. In Cambodia, the village animal health worker (VAHW) system was established in the early 1990s, involving short training programmes to equip VAHWs to provide basic animal health services for smallholder farmers, particularly the vaccination of cattle for haemorrhagic septicaemia (HS). However, there are increasing expectations that VAHWs provide village level disease surveillance information to the national veterinary services, despite their low-level disease diagnostic skills. To identify opportunities to improve the disease reporting system in Cambodia, a closed-ended cross-sectional study of VAHWs (n = 80) from two provinces was conducted in 2015, examining their contact frequency with district and provincial animal health authorities. Ordinal logistic regression was used to determine factors associated with increased frequency of VAHW contact with the district animal health authorities responsible for national disease reporting. Positive associations between income generated from VAHW activities (p = .01) and the frequency of visiting farmers (p

We evaluate the efficacy of recombinant DNA vaccine ABA392 against haemorrhagic septicaemia infection through intranasal administration route by targeting the mucosal immunity. The DNA vaccine was constructed and subjected to animal study using the Sprague Dawley (SD) rat. The study was divided into two major parts: (i) active and (ii) passive immunization studies, involving 30 animals for each part. Each group was then divided into five test groups: two test samples G1 and G2 with 50 and 100 µg ml-1 purified DNA vaccine; one positive control G5 with 106 CFU per ml formalin-killed PMB2; and two negative controls, G3 and G4 with normal saline and pVAX1 vector. Both studies were conducted for the determination of immunogenicity by total white blood cell count (TWBC), indirect ELISA and histopathological changes for the presence of the bronchus-associated lymphoid tissue (BALT). Our findings demonstrate that TWBC, IgA and IgG increased after each of the three vaccination regimes: groups G1, G2 and G5. Test samples G1 and G2 showed significant differences (P

Sudden death is usually the main finding in field animals during haemorrhagic septicaemia outbreaks caused by Pasteurella multocida type B:2 that causes acute, fatal and septicaemic disease in cattle and buffaloes. This situation may be due to failure in early detection of the disease where early treatment of antibiotics may improve the prognosis of the animal and other surviving animals. Thus, there is a grey area on the knowledge on the potential usage of pro-inflammatory cytokines and acute phase proteins as early biomarkers in the diagnosis of haemorrhagic septicaemia. In addition, exploration of the cerebrospinal fluid during infection has never been studied before. Therefore, this study was designed to fill up the grey areas in haemorrhagic septicaemia research. Twenty-one buffalo calves were divided into seven treatment groups where group 1 was inoculated orally with 10 mL of sterile phosphate-buffered saline pH 7 which act as a negative control group. Groups 2 and 3 were inoculated orally and subcutaneously with 10 mL of 1012 colony-forming unit of P. multocida type B:2. Group 4 and 5 buffaloes were inoculated orally and intravenously with 10 mL of lipopolysaccharide broth. Groups 6 and 7 were administered orally and subcutaneously with 10 mL of outer membrane protein broth. During the post-infection period of 21 days, blood and cerebrospinal fluid were sampled for the analyses of pro-inflammatory cytokines, acute phase proteins and cytological examination. Buffalo calves infected with P. multocida and its immunogens via different routes of inoculation showed significant changes (p

Loaches are popular in aquaculture in China due to their flesh having a high concentration of nutrients important for human health, such as amino acids, vitamins, minerals, and taurine. As such, they are used in Chinese traditional medicine to restore health and slow aging. However, the quick expansion of intensive loach culture to meet consumer demand has been accompanied by the increased prevalence of disease outbreaks, most often caused by Aeromonas spp. bacteria. We collected and identified pathogenic bacteria causing hemorrhagic septicemia in pond loach sourced from two provinces of China (Hubei and Hunan), and found three Aeromonas species: hydrophila (the most prevalent), sobria and veronii. We performed a bacterial challenge on the pond loach (Misgurnus anguillicaudatus) using the different concentrations of A. hydrophila in order to determine the median lethal dose, and found that the LD50 value was 2.14 × 107 CFU/fish. Hepcidin protein encoded by hepcidin antimicrobial peptide (hamp) gene is a key factor for iron regulation, enhancement of the host's innate immune system and acts as antimicrobial peptide. To better understand the functions of this protein in pond loach, we cloned it, conducted in silico physicochemical characterization, and studied its expression at the mRNA level both in healthy loach, and after a challenge with A. hydrophila. We found two very similar paralogs (a and b) of the hamp 1 gene, the open reading frames of which were 276 bp, both encoding an identical 91 amino acid putative peptide. The purified recombinant hepcidin protein exhibited antibacterial activity against all six studied bacterial species at 100–3200 arbitrary units/ml and can decrease 50% of the cumulation mortality of loaches which were injected by A. hydrophila. Both hamp 1 paralogs were basally expressed in most tissues, and significantly up-regulated in the liver, spleen and kidney after the bacterial challenge. The results of this study strongly indicate that hepcidin plays a role in defense mechanisms against pathogenic bacteria and inflammation in loaches.

Striped catfish (Pangasianodon hypophthalmus) farming in the Mekong Delta Vietnam (MKDVN) importantly contributes to national aquaculture export. Currently, however, diseases occur more frequently across the entire MKDVN region. One of the most common types is hemorrhagic septicemia caused by Aeromonas hydrophila. In this study, isolation and selection of the phages for control in vitro Aeromonas hydrophila were conducted. 24 phages were isolated from 100 striped catfish pond water samples. Next, lytic activity of these phages was clarified. Four phages with short latent period (about 25 to 40 min) and/or high burst size (about 67 to 94 PFU/ cell) were selected to evaluate their infection activity to different phage-resistant A. hydrophila strains. Two phages termed as TG25P and CT45P were subjected to the phage cocktail to inactivate A. hydrophila. Re-growth of the host bacteria appeared about eight hours after treatment. Usage of the phage cocktail that attach different host bacterial receptors is not always much effective than usage of single phage. This is the first report about phage therapy to control A. hydrophila isolated from striped catfish. Some challenges in the phage cocktail were shown to achieve strategies in prospective studies in the context of high antibiotic resistance of A. hydrophila.

Hemorrhagic septicemia is an acute, highly fatal disease that affects goldfish (Carassius auratus). To gain a better understanding of related immune genes, the transcriptomes of the skin and head kidney of goldfish suffering hemorrhagic septicemia were sequenced, assembled, and characterized. Based on functional annotation, an extensive and diverse catalog of expressed genes were identified in both the skin and head kidney. As two different organs, pair-wise comparison identified 122/77 unigenes up/down-regulated (two-fold change with P

Aeromonas hydrophila causes disease in fish known as Motile Aeromonas Septicemia (MAS), also named as bacterial hemorrhagic septicemia. In this study, a pathogenic A. hydrophila strain was isolated from common carp Cyprinus carpio L., which were suffering from severe hemorrhagic septicemia. According to the phylogenetic analysis derived from 16S rDNA sequence, the isolate formed a single branch in the A. hydrophila group, named AhHN1. Artificial infection results indicated that AhHN1 showed strong pathogenicity in C. carpio and the LD50 was 1.38 × 106 CFU/fish, the clinical symptoms and pathological features of infected fish were similar to those observed in natural infections. The antimicrobial susceptibility testing revealed that AhHN1 resistance to more than 13 kinds of antimicrobial agents. However, the AhHN1 strain exhibited an extremely sensitivity to enrofloxacin, the in vitro activities of enrofloxacin were subsequently investigated and drug selection window (MSW) was 0.0016-0.0125 µg/ml. Pharmacokinetics data showed that plasma concentration of enrofloxacin was 0.0016, 0.0148 and 0.0282 µg/ml at 24 hr after orally administered with 2.5, 5 and 10 mg/kg enrofloxacin. Moreover, dosing once a day of 2.5, 5 and 10 mg/kg enrofloxacin, which the relative protection ratio (RPS) was amounted to 33.3, 66.7, and 83.3%, respectively. Therefore, 5 mg/kg enrofloxacin was considered to be the rational regimen for controlling AhHN1 infection in C. carpio in the countries where the use of enrofloxacin is permitted in aquaculture. The aim of this study was to establish a scientific medication regimen for the prevention and therapy of the mutidrug-resistant A. hydrophila infection.

Hemorrhagic septicemia of mandarin fish (Siniperca chuatsi) was mainly caused by Aeromonas hydrophila which was an opportunistic pathogen. In recent years, the disease has caused tremendous economic loss with high morbidity and mass mortality in the mandarin fish breeding industry. Histopathological analysis and the immune related gene expression profiles of mandarin fish (S. chuatsi) infected with A. hydrophila were investigated in this study. Transmission electron microscopy (TEM) images showed that the cells of A. hydrophila densely covered with a mass of fimbriae. Histopathological analysis revealed that inflammation, vacuolization and extensive necrosis existed in the gill, liver, spleen and head kidney of the diseased fish. Quantitative real-time PCR was performed to measure mRNA expression levels for six immune related genes in mandarin fish after A. hydrophila infection. The transcriptional analysis of these immune related genes demonstrated that the expression levels of major histocompatibility complex class II (MHC II), T cell receptor α (TCRα), tumor necrosis factor α (TNFα), CC chemokine 3, interleukin 8 (IL-8) and Hepcidin were strongly up-regulated in spleen and head kidney of mandarin fish post-infection. These results will contribute to further study on the pathogenesis and host defensive system in A. hydrophila infection.

Background Foot-and-mouth disease (FMD) and Haemorrhagic septicemia (HS) are two important diseases that are known to have caused significant economic losses to the cattle industry. Accordingly, vaccinations have been recognized as an efficient method to control and prevent both of the above-mentioned diseases. This study aimed to determine the immune response to FMD virus antigens and the recombinant outer membrane protein of HS (rOmpH) of Pasteurella multocida in cattle administered as a combination vaccine and compare antibody titers with the two vaccines given independently, under field conditions. Dairy cattle were divided into three groups. Each group was immunized with different vaccine types according to the vaccination program employed in this study. Antibody responses were determined by indirect ELISA, liquid phase blocking ELISA (LPB-ELISA) and viral neutralization test (VNT). Furthermore, the cellular immune responses were measured by lymphocyte proliferation assay (LPA). Results The overall antibody titers to HS and FMDV were above cut-off values for the combined FMD-HS vaccine in this study.The mean antibody titer against HS after the first immunization in the combined FMD-HS vaccine groups was higher than in the HS vaccine groups. However, no statistically significant differences (p > 0.05) were observed between groups. Likewise, the antibody titer to the FMDV serotypes O/TAI/189/87 and Asia 1/TAI/85 determined by LPB-ELISA in the combined vaccine were not statistically significantly different when compared to the FMD vaccine groups. However, the mean VNT antibody titer of combined vaccine against serotype O was significantly higher than the VN titer of FMD vaccine groups (p p p > 0.05). Conclusions The combined FMD-HS vaccine formulated in this study could result in high both antibody and cellular immune responses without antigenic competition. Therefore, the combined FMD-HS vaccine can serve as an alternative vaccine against both HS and FMD in dairy cattle under field conditions.

Haemorrhagic septicaemia caused by Aeromonas hydrophila in striped catfish (Pangasianodon hypophthalmus) is one of the most important aquatic diseases in the Mekong Delta, Vietnam. However, antibiotic-resistant A. hydrophila strains have become popular and resulted in inadequate control of the disease in striped catfish farms. This study investigates the protective efficacy of bacteriophage PVN02 against haemorrhagic septicaemia in striped catfish via oral administration. The phage-containing pellets were prepared by spraying the phage solution on food pellets at 20 ml/kg. The rate of phage desorption from the food pellets into the water was very low; the phage titres in the water were approximately log 1.0 PFU/ml or undetectable. The in vivo experiment evaluating the protective efficacy of PVN02 against haemorrhagic septicaemia in striped catfish was conducted using 21 groups of 1,260 fish in 50-L plastic tanks in triplicate. The catfish were fed twice daily with phage-sprayed pellets. Different densities of bacterial suspensions were added into the tanks for 24 hr. Without the existence of the phage, the highest mortality rate was 68.3 ± 2.9% at the highest density of bacterial suspension. In contrast, the mortality rate at the highest density of bacterial suspension was significantly reduced to 8.33 ± 2.9% or 16.67 ± 2.9% at the phage dose of log 6.2 ± 0.09 or log 4.2 ± 0.09 PFU/g. This study provides a very practical manner of applying phage therapy to prevent disease in large-scale striped catfish farms.

Diseased Anabas testudineus exhibiting signs of tail-rot and ulcerations on body were collected from a fish farm in Assam, India during the winter season (November 2018 to January 2019). Swabs from the infected body parts were streaked on sterilized nutrient agar. Two dominant bacterial colonies were obtained, which were then isolated and labelled as AM-31 and AM-05. Standard biochemical characterisation and 16S rRNA and rpoB gene sequencing identified AM-31 isolate as Aeromonas hydrophila and AM-05 as Aeromonas jandaei. Symptoms similar to that of natural infection were observed on re-infecting both bacteria to disease-free A. testudineus, which confirmed their virulence. LC50 was determined at 1.3 × 104 (A. hydrophila) and 2.5 × 104 (A. jandaei) CFU per fish in intraperitoneal injection. Further, PCR amplification of specific genes responsible for virulence (aerolysin and enterotoxin) confirmed pathogenicity of both bacteria. Histopathology of kidney and liver in the experimentally-infected fishes revealed haemorrhage, tubular degeneration and vacuolation. Antibiotic profiles were also assessed for both bacteria. To the best of our knowledge, the present work is a first report on the mortality of farmed climbing perch naturally-infected by A. hydrophila as well as A. jandaei, with no records of pathogenicity of the latter in this fish.

Hemorrhagic septicemia is a fatal disease of cattle and buffalo all over the world, including Pakistan, and it causes heavy economic losses every year. The poor farmers cannot bear this loss in the form of less milk production and heavy expenditures on the animal treatment. An enriched alum-precipitated vaccine with reduced dose was prepared and standardized, and safety testing of the enriched vaccine was performed in Swiss albino mice as well as in natural hosts. In this experiment, a total of 36 cattle both male and female, of different age groups ranging from 4 months to 4 years, were used. All these animals belonged to all major cattle breeds of Pakistan, including Sahiwal, Red Sindhi (crossbred and purebred), Dhani (crossbred and purebred), Lohani (crossbred and purebred), and Cholistani, and exotic breeds, including Holstein, Friesian and Jersey. These animals were examined for current immune titer prior to vaccination. Animals were vaccinated subcutaneously with 2 mL and 4 mL dose of the new vaccine and were observed for any untoward reaction for 48 h. All the animals were kept under close observation for the next 30 days, and all were found safe. The experiment was designed to reduce the dose of the vaccine to 2 mL by using BHI as a growth medium, as well as to increase the number of doses prepared in the same infrastructure, hence reducing the cost of vaccine production. The study proved that a vaccine with increased biomass in reduced dose is safe in local as well as in exotic breeds of cattle.

The objective of this study was to estimate sensitivity (Se) and specificity (Sp) of a novel enzyme-linked immunosorbent assay (ELISA) test (using a coating antigen from Pasteurella multocida M-1404 via heat extraction) and an indirect hemagglutination (IHA) test for detection of Hemorrhagic septicemia (HS) in dairy cows, under Thai conditions, using a Bayesian approach. Dairy cow sera with a total of 1236 samples from 44 farms were tested with the two tests to detect immune responses against the HS. Percentages of positive samples for the ELISA and IHA tests were 73% (901/1236) and 70% (860/1236), respectively. Estimated sensitivity and estimated specificity of the ELISA test were 90.5% (95% posterior probability interval (PPI) = 83.2–95.4%) and 70.8% (95% PPI = 60.8–79.8%), respectively. Additionally, estimates for the Se and Sp values of the IHA test were 77.0% (95% PPI = 70.8–84.1%) and 51.1% (PPI = 36.8–66.3%), respectively. The estimated prevalence of the disease was 71.7% (95% PPI = 62.7–82.6%). These results demonstrate that the ELISA test can be a useful tool for the detection of the presence of an antibody against the HS in dairy cows. Notably, the cows in this area indicated a high percentage of exposure to Pasteurella multocida.

Background Pasteurella multocida is the etiological agent responsible for several diseases in a wide range of hosts around the world and thus, causes serious economic losses. Acute septicemia associated with capsular type B P. multocida has recently emerged in Europe and continuous outbreaks of these acute processes have been described in Spain since they were first detected in pigs in 2009 and cattle in 2015. The scarcity of studies on the antimicrobial susceptibility of this capsular type of P. multocida and growing concern about the general increase of antimicrobial resistance mean that studies related to the performance of type B P. multocida against antibiotics are necessary to establish accurate treatments and to monitor antimicrobial resistances. Results Seventy-six isolates of P. multocida type B from pigs and cattle with acute septicemia were tested for susceptibility to 10 different antimicrobials. Bovine isolates were susceptible to all the antibiotics we tested except for lincomycin (94.4% of isolates were resistant). However, the antimicrobials we tested were less effective against swine isolates, of which none were susceptible to lincomycin. Furthermore, 29.3% swine isolates were resistant to tetracycline, 27.6% to penicillin, 20.7% to oxytetracycline, 17.3% to chloramphenicol, 15.5% to gentamicin, and 3.4% to enrofloxacin; no resistance to ceftiofur was detected. No multidrug resistant isolates were detected from cattle, while 25.86% of swine isolates were resistant to three or more antibiotic classes. Conclusions In this study, the lower resistance rates and multidrug resistant isolates reported for P. multocida type B derived from cattle compared to those isolated from pigs may be related to the increased use of antibiotics in the porcine industry in Spain. Lincomycin is not recommended for the treatment of acute septicemia in pigs or cattle, rather, the use of ceftiofur, enrofloxacin, or gentamicin is indicated as an emergency treatment in the early stages of disease; once the susceptibility results are known, the use of tetracyclines, penicillin, or chloramphenicol should be prioritized. The increase in multidrug resistant isolates and antimicrobial resistance rates indicates that more attention should be paid to prevention as well as the responsible use of antibiotics.

Hemorrhagic septicemia (HS) is an important infectious disease in cattle and buffaloes, caused by Pasteurella multocida B:2 and E:2. The intranasal recombinant OmpH-based vaccine was successfully used to protect dairy cattle from HS in a previous study. Thus, this study aimed to examine the protective ability of that vaccine among buffaloes. Four groups of Thai swamp buffaloes received different vaccines and were labeled as 100 or 200 μg of the rOmpH with CpG-ODN2007, commercial HS bacterin vaccine, and nonvaccinated control groups. Sera and whole blood were collected to examine the antibody levels and cellular immune response using indirect ELISA and MTT assay, respectively. Challenge exposure was performed with virulent P. multocida strain M-1404 serotype B:2 on day 72 of the experiment. The antibody titers to P. multocida among immunized buffaloes were significantly higher than in the control group (p<0.01), especially the 200 μg of the rOmpH group. The stimulation index (SI) of the intranasally vaccinated groups revealed significantly higher levels than the nonvaccinated group (p<0.01), but not different from the intramuscularly commercial HS vaccine. The clinical signs and high fever were observed after challenge exposure in the nonvaccinated group, while it was not observed among the 200 μg of rOmpH immunized buffaloes. The other immunized groups showed partial protection with transient fever. In conclusion, the rOmpH-based intranasal vaccine could elicit protective ability and induce antibody- and cell-mediated immune response against virulent P. multocida strain among swamp buffaloes.

Aim This study aimed to create rapid characterization and genotyping of Pasteurella multocida (PM) protocol using modern molecular biology techniques. Materials and methods Thirty bacterial isolates were characterized by capsular and somatic identification using conventional procedure followed by multiplex polymerase chain reaction (PCR), restriction endonucleases analysis (REA), and finally confirmed by sequence analysis. Two local vaccine strains and two field isolates were identified as PM Type A and B. Results A total of 30 isolates were found positive for PM either morphologically and biochemically; however, multiplex PCR technique identified only 22 isolates as Pasteurella species using universal primers while 8 isolates were found negative for PM. 12 of 22 isolates (54%) were characterized at the same reaction into PM Type A, five isolates (23%) were Type B and the rest five isolates (23%) of tested isolates were negative for Types A, B, and D. Hemorrhagic septicemia Type B: 2 or B: 5 could be identified somatically within PM capsular serogroup B using PCR technique. Somatic characterization of PM was done using REA that could identify all PM Type A into A:1 and all PM Type B into B: 2. These protocols were verified for its accuracy and reliability by sequence analysis of two vaccine strains of PM Type A and B that were characterized previously by biochemical and serological methods as well as two selected isolates from the 22 positive isolates representing PM Type A and B. Conclusion PCR and REA could confirm the identity of PM and provide a rapid and reliable characterization in comparison with biochemical analysis and conventional serotyping that may take up to 2 weeks. Hence, they can reduce the time needed for polyvalent vaccine production and when the reference antisera are unavailable. Moreover, the identity of Omp-H for vaccine and field strains may provide better data to control Pasteurellosis in Egypt.

Hemorrhagic septicemia (HS) is a globally renowned disease that affects cattle and buffaloes. Its effects on the reproductive system have not been previously studied. Thus, the present study aims to evaluate the pathological responses in pre-pubertal female buffalo infected with immunogens; lipopolysaccharide (LPS) and outer membrane protein (OMP) derived from P. multocida type B: 2. Fifteen healthy pre-pubertal female buffaloes of approximately 8 months old were selected and divided into five equal groups of 3 buffaloes each. Buffaloes in groups 1 and 2 were intravenously and orally inoculated with 10 mL of P. multocida LPS at 1× 1012 colony forming unit (cfu) ,while those in group 3 and group 4 were subcutaneously and orally inoculated with10 mL of P. multocida OMP at 1× 1012 colony forming unit (cfu). Buffaloes in group 5 were inoculated with 10mL of sterile phosphate buffered saline (PBS) PH7 and served as the negative control. During the post infection period, all buffaloes were examined for clinical signs throughout 21 days and surviving buffaloes were euthanized for postmortem evaluation. Histopathological evaluation of buffaloes inoculated with LPS showed a significantly higher occurrence (p

BACKGROUND/AIMS Increased risk of bacterial infection is common in cirrhotic patients with upper gastrointestinal bleeding (UGIB). Our study aimed to explore the association of the bacteremia with in-hospital mortality and risk factors of bacteremia in these patients. MATERIALS AND METHODS In our retrospective cohort study, we collected data for cirrhotic patients with UGIB admitted to our hospital between August 2010 and December 2010. The primary outcome was in-hospital mortality. The secondary outcome was bacteremia. A multivariate logistic regression analysis was performed to determine risk factors for mortality and bacteremia. RESULTS A total of 202 patients with cirrhosis presenting with UGIB at the emergency department (ED) were enrolled. Bacteremia was associated with a higher mortality rate (adjusted odds ratio [OR]: 9.7; 95% confidence interval [CI]: 1.9-50.6, p=0.007), whereas shock (systolic blood pressure 0% immature neutrophils of band form) were associated with bacteremia in cirrhotic patients with UGIB (adjusted OR: 5.3; 95% CI: 2.3-12.7, p

Pasteurella multocida serotype B:2 causes hemorrhagic septicemia in cattle and buffalo. The invasion mechanism of the bacterium when invading the bloodstream is unclear. This study aimed to characterize the effects of immunomodulatory molecules, namely dexamethasone and lipopolysaccharide, on the invasion efficiency of P. multocida serotype B:2 toward bovine aortic endothelial cells (BAECs) and the involvement of actin microfilaments in the invasion mechanism. The results imply that treatment of BAECs with lipopolysaccharide at 100 ng/mL for 24 h significantly increases the intracellular bacteria number per cell (p ＜ 0.01) compared with those in untreated and dexamethasone-treated cells. The lipopolysaccharide-treated cells showed a significant decrease in F-actin expression and an increase in G-actin expression (p ＜ 0.001), indicating actin depolymerization of BAECs. However, no significant differences were detected in the invasion efficiency and actin filament reorganization between the dexamethasone-treated and untreated cells. Transmission electron microscopy showed that P. multocida B:2 resided in a vacuolar compartment of dexamethasone-treated and untreated cells, whereas the bacteria resided in cellular membrane of lipopolysaccharide-treated cells. The results suggest that lipopolysaccharide destabilizes the actin filaments of BAECs, which could facilitate the invasion of P. multocida B:2 into BAECs.

To evaluate the safety and immunogenicity of combined hemorrhagic septicemia (HS) and blackleg (BL) vaccine, combined vaccine was produced in two different formulations (group one, G1 and group two, G2) and compared with safety and immunogenicity of monovalent Hs and BL vaccines in different doses. G1 combined vaccine was vaccinated in 4 and 2 ml in to five calves each while G2 combined vaccine, the monovalent HS and blackleg vaccines were vaccinated in 2 and 1 ml amount into to five calves each leaving ten non-vaccinated calves managed similarly as vaccinated ones. The safety test was made by giving single and double doses of experimental vaccines and double doses of monovalent vaccines in each case using two calves per dose group. The immunogenicity of BL vaccine component in the combined vaccine was evaluated by using 10 guinea pigs per group for six different doses of combined and monovalent blackleg vaccine, leaving 10 non-vaccinated controls. The study indicated that the protection against virulent challenges for animals vaccinated with G1 combined vaccine at 2 ml, G2 vaccine at 1 ml, monovalent HS and BL vaccine vaccinated at 1 ml doses was by far less than 90%. On the other hand, protection against experimental challenge for G1 vaccine vaccinated in 4 ml amount was 100% against both HS and blackleg virulent challenges while the protection against experimental challenge for G2 vaccine vaccinated in 2 ml amount was 66.67% against the HS virulent challenge and 90% against BL virulent challenge. So G1 combined vaccine vaccinated in 4 ml was found to be the best candidate vaccine according to this experiment which needs to be confirmed at field test before use for mass vaccination. Key words: Combined, hemorrhagic septicemia, blackleg, safety, immunogenicity.

Haemorrhagic septicemia (HS) is a contagious disease in cattle with high morbidity and mortality rates. HS vaccine in Thailand is an oil-adjuvant formulation, and is difficult to administer. The present study aimed to formulate and evaluate the protection in dairy calves conferred by immunization with an in-house intranasal HS vaccine. The intranasal vaccine was formulated in a total volume of 500 µl containing either 50 or 100 µg of the recombinant outer membrane protein H (rOmpH) of Pasteurella multocida strain M-1404 (serovar B:2), and 10 µg of Cytosine-phosphate-guanosine oligodeoxynucleotides (CpG-ODN) as a mucosal adjuvant. Intranasal immunizations were conducted three times at three-week intervals. The antibodies post-immunization were detected by indirect ELISA and demonstrated efficient in vitro activity in suppressing a P. multocida strain from the complement-mediated killing assay. An intranasal vaccine induced both the serum IgG and secretory IgA levels that were significantly higher than the level conferred by the parenteral vaccine (P

Hemorrhagic septicemia is a highly infectious and contagious disease caused by Pasteurella multocida serogroup B:2 in tropical Asian and African countries. The acute inflammatory responses induced by Pasteurella multocida are the main cause of death in hemorrhagic septicemia. Therefore, present study was undertaken to examine the blood cytokine expression profiles (TNF-α, IL-1β, and IL-6), bacterial colonization and histopathological changes of intraperitoneally and subcutaneously challenged vaccinated and unvaccinated mice with 102 CFU of P. multocida P52. The observations were made at 6, 12, 18, 24 h and 48 h intervals. Real-time PCR based blood cytokine profiles (TNF-α, IL-1β, and IL-6) measurement revealed a significantly higher amount of pro-inflammatory cytokines expression in the unvaccinated challenged group of mice than the vaccinated challenged group. There was heavy bacterial load in all organs of mice viz. trachea, lung, spleen, within 6 h of challenge in both vaccinated and unvaccinated group of mice, but bacterial load increased in the unvaccinated challenged group of mice with respect to time whereas the load were constant in the vaccinated challenged group. Histopathological changes were mild in the vaccinated challenged group of mice in comparison to the unvaccinated challenged group. There was no significant difference in the bacterial load, histopathological changes and cytokines expression when challenged through different routes.

Lipopolysaccharide (LPS) of P. multocida B:2, a causative agent of haemorrhagic septicaemia (HS) in cattle and buffaloes, is considered as the main virulence factor and contribute in the pathogenesis of the disease. Recent studies provided evidences about the involvement of the nervous system in pathogenesis of HS. However, the role of P. multocida B:2 immunogens, especially the LPS is still uncovered. Therefore, this study was designed to investigate the role of P. multocida B:2 LPS to induce pathological changes in the nervous system. Nine eight-month-old, clinically healthy buffalo calves were used and distributed into three groups. Calves of Group 1 and 2 were inoculated orally and intravenously with 10 ml of LPS broth extract represent 1 × 1012 cfu/ml of P. multocida B:2, respectively, while calves of Group 3 were inoculated orally with 10 ml of phosphate buffer saline as a control. Significant differences were found in the mean scores for clinical signs, post mortem and histopathological changes especially in Group 2, which mainly affect different anatomic regions of the nervous system, mainly the brain. On the other hand, lower scores have been recorded for clinical signs, gross and histopathological changes in Group 1. These results provide for the first time strong evidence about the ability of P. multocida B:2 LPS to cross the blood brain barrier and induce pathological changes in the nervous system of the affected buffalo calves.

The present study tested the susceptibility and pathological changes of catla, Catla catla (Hamilton) infected with Edwardsiella tarda (ET-PG-29). The bacterium was isolated from the kidney of a diseased pangas catfish. To determine the median lethal dose (LD50), C. catla were challenged with this bacterium (108-103 CFU ml-1), and the LD50 was calculated as 105.5 CFU ml-1. Another set of healthy C. catla were injected intraperitoneally with the LD50 dose to induce edwardsiellosis. The clinical signs of the infected C. catla were observed and recorded. Tissues such as kidney, liver, intestine, heart, and gill from the infected fish with clinical signs of edwardsiellosis were used for histopathology. The clinical and gross signs were first visible at 1 d post-injection, and the infected fish showed typical signs of hemorrhagic septicemia. The most striking histopathological features were found in the kidney which showed multi-focal necrosis with the formation of granuloma indicating an inflammatory response against the pathogen. The intestine displayed goblet cell hyperplasia, the liver showed hydropic degeneration with hyperemic central veins, and there was inflammation of gill lamellae and cardiac myositis associated with leucocyte infiltration. Collectively, the results confirmed the susceptibility of C. catla to E. tarda infection and that this bacterium is a threat to C. catla in aquaculture practices. CORRESPONDING AUTHOR: T.B. Devi, D. Kamilya [+] Department of Fish Health and Environment College of Fisheries, Central Agricultural University Lembucherra, Tripura (w) – 799210, Tripura, India T.J. Abraham Department of Fishery Pathology and Microbiology, Faculty of Fishery Sciences, West Bengal University of Animal and Fishery Sciences 5 – Budherhat Road, Chakgaria, P. O – Panchasayar Kolkata – 700094, West Bengal, India

Inactivated bacterial vaccines may contain residual levels of formaldehyde, which may have adverse effects. Therefore, it is important to determine the residual level of formaldehyde to ensure product safety. A number of assays are available for the determination of residual free formaldehyde in inactivated vaccines, including acetyl acetone titration, ferric chloride titration, and the basic fuchsin test. In the current study, ferric chloride quantitative method was used to evaluate formaldehyde residue on 4 types of vaccine produced in the Bacterial Vaccine section in Veterinary Research Institute (VRI), Ipoh. Four types of vaccines tested were Hemorrhagic Septicemia oil (HS oil), Hemorrhagic Septicemia alum (HS alum) vaccine, Duck vaccine (DV), and Sheep and Goat Pasteurellosis vaccine (SGP). Results revealed formaldehyde residue in vaccines tested was between 1.289 g/L to 2.228 g/L (mean SD ±0.18). According to European Pharmacopeia, when formaldehyde has been used in the preparation, the concentration of free formaldehyde is not greater than 0.5 g/L. While the United States Code of Federal stated that it must not exceed 0.74 g/L unless a higher amount has been shown to be safe. In our study, all the vaccines has been tested for safety and potency test before being used on animals, and therefore, it is safe for vaccination purpose. Formaldehyde residue will be conducted for each batch of vaccines production to ensure its safety.

Mass mortality events in wildlife are a growing concern. Under conditions of rapid global change, opportunistic responses in bacterial commensals, triggered by environmental stressors, may be increasingly implicated in die‐offs. In 2015, over 200,000 saiga antelope died of hemorrhagic septicemia caused by the pathogen Pasteurella multocida serotype B. We use this case to explore die‐offs from commensal bacteria more broadly, looking at factors which might favor such extreme events. We review other recorded disease outbreaks caused by Pasteurellaceae organisms, firstly in saiga and secondly in other wild ungulates, and ask whether the 2015 die‐off was unprecedented in terms of mortality rates, numbers dead, spatial scale, and in the nature of the predisposing or environmental factors involved. We also compare these outbreaks with mass mortality events associated with commensal bacteria in wildlife more generally. We identify three additional major die‐offs in saiga in which Pasteurellaceae organisms may be implicated, of which one in 1988 closely resembles the 2015 hemorrhagic septicemia event. No other recorded cases in wild ungulates approach the magnitude of these cases for any of the metrics considered, possible exceptions being die‐offs in Mongolian gazelles, in which the role of these pathogens is poorly substantiated. Environmental triggers were the most commonly suggested factor leading to pathogenesis, with warm humid conditions most commonly associated with hemorrhagic septicemia. Life history may also be significant—saigas are migratory and the largest pasteurellosis outbreaks outside this species also occur in migratory species of bird or other temperate ungulates aggregating in large numbers. Cases provoked by other commensals tend to be small in magnitude. Exceptions involve interactions between multiple pathogens and climatic conditions or sets of climatic conditions acting on different stages of the host–pathogen life cycle, leading to time lags between infection and subsequent disease. Overall, the scale and rapidity of the saiga die‐offs appear unprecedented among mortality events caused by bacterial commensals in wild mammals. Experimental research into the genetics and microbiology of host–pathogen interactions upon changes in the external environment, and monitoring of animals and conditions at calving sites, may eventually reveal the underlying causes of these die‐offs.

Hemorrhagic septicemia (HS) is a highly fatal disease caused by Pasteurella multocida that often cause outbreaks in buffalo and cattle in India, and thus is a major cause of production losses. It is one of the livestock diseases with the highest mortality, and despite available vaccines, outbreaks still occur. To assess the seroprevalence in the state of Assam, Northeast India, 346 serum samples from cows from 224 randomly selected households, from both urban and rural areas of three districts, were tested with a commercial ELISA. In total 88 cows were seropositive (25.4%), and indigenous cattle were significantly more seropositive (33.5%) compared to the crossbred cattle (18.5%) (p = 0.002). Herd prevalence was 35.7%, and more rural farms (47.4%) were positive compared to the urban farms (23.6%) (p < 0.001). No other risk factors were identified in this study. Only one farm had vaccinated against HS, but there were no seropositive animals detected in that herd. This study shows that HS is highly prevalent in Assam. Considering the importance of dairy production in India, and the dependence of the rural Assam population on farming and livestock keeping, more extensive vaccination campaigns would be important.

4th International Conference on Computational and Experimental Science and Engineering (ICCESEN) -- OCT 04-08, 2017 -- Kemer, TURKEY WOS: 000478894500021 Fish diseases are a major obstacle to the development of the fisheries industry. Aeromonas sobria is an infectious waterborne bacterium that causes ulcers, tail rot and hemorrhagic septicemia in fishes and resistant to many existing antibiotics. In this context, A. sobria-AgNPs were synthesized by A. sobria using AgNO3. A. sobria-AgNPs were characterized using UV-Vis spectroscopy, and a peak was obtained at a range of 420-480 nm. A. sobria-AgNPs were evaluated for antibacterial activities against different fish pathogens. The highest antibacterial activity was observed against A. hydrophila, E. cloacae and E. coli. The lower activity was found against C. braakii and E. hermannii, but against H. alvei, P. rettger and M. morganii subsp. sibonii no zone of inhibition was recorded. The results indicated that the A. sobria-AgNPs can be used to develop antibacterial agent and as a therapeutic agent in the fishing industry and water disinfection. The antibacterial efficacy against the fish pathogen A. hydrophila of silver nanoparticles is a hope for possible application as a disinfectant or antimicrobial agent for better fish health management. Ahi Evran University Scientific Research Projects Coordination UnitAhi Evran University [FEF.E2.17.038] This work was supported by the Ahi Evran University Scientific Research Projects Coordination Unit. Project Number FEF.E2.17.038.

Haemorrhagic septicaemia (HS) is an acute, fatal, septicaemic disease of cattle and buffaloes caused by one of two specific serotypes of Pasteurella multocida B:2 and E:2 in Asian and African, respectively. It is well known that HS affect mainly the respiratory and digestive tracts. However, involvement of the nervous system in pathogenesis of HS has been reported in previous studies without details. In this study, nine buffalo calves of 8 months old were distributed into three groups. Animals of Group 1 and 2 were inoculated orally and subcutaneously with 10 ml of 1 × 10(12) cfu/ml of P. multocida B:2, respectively, while animals of Group 3 were inoculated orally with 10 ml of phosphate buffer saline as a control. All calves in Group 1 and Group 3 were euthanised after 504 h (21 day) post-infection, while calves in Group 2 had to euthanise after 12 h post-infection as they develop sever clinical signs of HS. Significant differences were found in Group 2 in the mean scores of clinical signs, gross and histopathological changes which mainly affect different anatomic regions of the nervous system. In addition, successful bacterial isolation of P. multocida B:2 were obtained from different sites of the nervous system. On the other hand, less sever, clinical, gross and histopathological changes were found in Group 1. These results provide for the first time strong evidence of involving of the nervous system in pathogenesis of HS, especially in the peracute stage of the disease.

We report the case of a 56-year-old female patient, with a three-day history of hematemesis, melena, abdominal wall hematoma and epistaxis associated with thrombocytopenia and anemia. Idiopathic thrombocytopenic purpura was diagnosed and she was treated with dexamethasone for four days. The patient developed acute respiratory failure with signs of systemic inflammatory response. Blood and pleural fluid cultures grew Pasteurella canis. This is the first case, to our knowledge, of P. canis empyema associated with hemorrhagic septicemia without epidemiological background and the third case of septicemia caused by P. canis reported in the literature.

Lactococcosis is a hyperacute hemorrhagic septicemia disease caused by Lactococcus garvieae, which is an emerging pathogen in global fish farming. Between 2016 and 2018, rainbow trout (Oncorhynchus mykiss) from five farms that presented outbreaks were sampled as part of a Mexican surveillance program for the detection of fish diseases. Fourteen L. garvieae isolates were recovered from sampled fish, as confirmed by biochemical tests, 16S rRNA gene sequencing, and clinical and histological insights. The biochemical and protein profiles of the isolates obtained were homogeneous. Repetitive extragenic palindromic&mdash, (REP)&mdash, and enterobacterial repetitive intergenic consensus sequence PCR (ERIC-PCR) analyses established weak genetic heterogeneity. Rainbow trout challenged with two of the isolates used at different bacterial concentrations (10&minus, 2 and 10&minus, 4 CFU/mL) showed melanosis, and hemorrhages were observed in the fins, liver, kidney, and spleen. Isolates were obtained from all of the organs sampled, including from surviving fish, as either pure or mixed cultures. The present study is the first to confirm the presence of L. garvieae as the agent of severe lactococcosis outbreaks in the two primary Mexican states for trout farming.

Aeromonas spp. are the causative agents of motile Aeromonas septicemia (MAS) in fish, as well as gastroenteritis and infections in humans. The antigen diversity between Aeromonas spp. and serotypes greatly challenges the identification of this zoonotic pathogen in environmental, food, and clinical samples. In the present study, six conserved peptides in the outer membrane of Aeromonas were selected and used as immunogens after conjugation to produce monoclonal antibodies (mAbs). Peptides 1 and 4 with the amino sequences of “VYDKDGTTFD” and “GGFKGKLSYQTND”, effectively elicited cross-reactive antibodies against Aeromonas in mice. Based on the mAbs 1B1 and 14H5, the developed sandwich immunoassay detected twelve Aeromonas spp. species, including A. hydrophila (8/8), A. veronii (7/7), A. caviae, A. sobria, A. dhakensis, A. media, A. popoffii, A. punctata subsp. caviae, A. bivalvium, A. lacus, A. jandaei, A. bestiarum and A. salmonicida (1/2). The detection limit of these strains primarily ranged from 1.69 × 104 to 4.57 × 105 CFU mL−1. No cross-reactivity was observed with the other tested strains. The sensitivity and specificity of this method was 96% (26/27) and 100% (15/15), respectively. Furthermore, real sample analyses detected river water samples containing 105 CFU mL−1 of A. hydrophila or A. caviae, and detected Carassius auratus samples showing hemorrhagic septicemia after enrichment in buffered peptone water. The established immunoassay may be promising as an effective screening method for monitoring Aeromonas spp. in environmental and food samples.

Haemorrhagic septicaemia (HS) is a well-known high fatality septicaemic disease happening among bovines. The disease is caused by the Pasteurella multocida serotype B:2 bacteria. P. multocida B:2 has high mortality and morbidity rates and is spread through the intranasal and oral routes in bovines. In this study, our aim was to investigate the efficacy of the recombinant protein vaccine, ABA392/pET30a via intranasal inoculation by targeting the mucosal immunity. The constructed recombinant protein vaccine ABA392/pET30a was subjected to an animal study using Sprague Dawley rats. The study was divided into two parts: active and passive immunization studies. Both studies were carried out through the determination of immunogenicity (using Total White Blood Cell (TWBC) Count with Indirect ELISA) and histopathogenicity, analyzing (Bronchus Associated Lymphoid Tissue (BALT) formation) in lungs. As a result, the IgA and IgG development of both tested groups: group 1 (50μg/mL protein vaccine) and group 2 (100μg/mL protein vaccine) showed equivalent with the positive control group 4 (formalin-killed P. multocida B:2). However, there was a significant difference when compared with the negative control group 3 (normal saline). These results demonstrate that both the protein vaccine at the concentration 50μg/mL and 100μg/mL have the same efficacy as the commercially available positive control vaccine. From the studies, higher concentration of protein vaccine at 100μg/mL showed higher development of both IgA and IgG compared to 50μg/mL protein vaccine. Higher and rapid development of IgA compared to IgG showed that mucosal immunity has been induced through the intranasal administration of the protein vaccine. In addition, leucocytosis was observed at each dose of vaccination showed that the protein vaccine is capable to induce the immune responses of the host. Histopathogenicity studies of the vaccinated groups showed more BALT formation and no severe lesions after challenge compared to the negative control group. Besides, no inflammatory onsite or anaphylactic responses were observed after the intranasal inoculation which proved to be safer and provided longer lasting immunity. Therefore, recombinant protein vaccine ABA392/pET30a could be a potential candidate for intranasal administration which can provoke mucosal immunity against HS disease.

Weissella ceti, a Gram-positive nonmotile bacterium, is currently an emerging pathogen within rainbow trout (Oncorhynchus mykiss) farms in China, Brazil, the United States, and Japan. This study is the first to isolate, identify, and characterize W. ceti isolates from rainbow trout farmed in Mexico. In late 2015, a severe disease outbreak caused a 60% mortality rate among 20,000 fish. The diseased rainbow trout (100-300 g average) exhibited severe cachexia, body darkening, abdominal distension, exophthalmia, haemorrhages, and corneal opacity. Internally, diseased fish had pale gills; multifocal, disseminated whitish spots on the liver; haemorrhages in the swim bladder, ovary, and on the parietal surface of the muscle; and hearts with pseudo-membrane formation. Histologically, lesions were characterized by corneal oedema, degenerative and necrotic hepatitis, and meningitis. A brain (W-1) and kidney (W-2) isolate were identified as W. ceti through polyphasic taxonomy, which included phenotypic characterization and 16S rRNA sequencing. RAPD and ERIC-PCR analyses demonstrated genetic homogeneity among the Mexican isolates. Virulence tests in rainbow trout through intraperitoneal W. ceti injections at concentrations of 1 × 104 , 1 × 105 , and 1 × 106 CFU per fish resulted in cumulative mortality rates of 25%, 62.5%, and 87.5%, respectively, as well as the same clinical signs of hemorrhagic septicaemia as were recorded for the natural outbreak. The present report is the first to confirm the presence of W. ceti in Mexico, thus extending the known geographical distribution of this pathogen across the Americas.

A retrospective epidemiological case-control study was performed in Karachi, Pakistan, from January to April 2013. The owners of 217 dairy cattle and buffalo farms from six different locations in Karachi were interviewed. The aim of the study was to identify risk factors associated with the presence of haemorrhagic septicaemia (HS). Farms with a history of at least one instance of sudden death in a dairy animal during 2012 and a positive clinical HS diagnosis (made by local veterinarians) were defined as cases. Farms having no history of sudden deaths in 2012 were defined as controls. Univariable analyses were initially conducted, and factors with P ≤ 0.25 were offered to a multivariable logistic regression model to identify putative risk factors. The final multivariable logistic model contained five factors. Vaccination was found to be a protective factor (OR = 0.22) along with the length of time cattle were kept on farm (months). For every extra month cattle were kept, the odds of HS disease were reduced by a factor of 0.9. In contrast, for every extra animal in a herd, the risk of infection increased by a factor of 1.01. Supplying underground water and the presence of foot and mouth disease on the farm increased the risk by 2.90 and 2.37, respectively. To understand the epidemiology of HS in Karachi dairy herds, more in-depth research is required to study the risk and protective factors identified in this survey and to evaluate risk mitigation strategies, where possible.

The aim of the present study was to identify and compare the variations among clinical of P. multocida isolates originating from different hosts, sheep (n = 4), buffalo (n = 4) and cattle (n = 4). These isolates were obtained from various locations in the Fayoum governorate, Egypt and they were analyzed by morphological, API kit and RAPD PCR at molecular level to check the distribution of Pneumonia and hemorrhagic septicemia in different farm animals. The chosen isolates exhibited luxuriant growth on blood agar with translucent grayish or yellowish green colonies. However, they showed no growth on MacConkey and no hemolysis of blood; also isolates were positive for oxidase, catalase and H2S production, negative of urease and can ferment glucose, fructose and glucose but can't ferment lactose. Antibiotic sensitivity showed that isolates were sensitive to tetracycline, clindamycin and erythromycin but resistant to ampicillin, trimethoprim/sulfamethoxazole and flucloxacillin. To determine the genetic diversity nine random primers OPA-01  OPA09 were used to which generate different profiles. The distinct RAPD profiles of p. multocida strains showed specificity with regard to the host and the disease induced.

Background: Pasteurella multocida a Gram-negative bacterium has been identified as the causative agent of many economically important diseases in a wide range of hosts. Hemorrhagic septicemia is a disease caused by P. multocida serotype B:2 and E:2. The organism causes acute, a highly fatal septicemic disease with high morbidity and mortality in cattle and more susceptible in buffaloes. Therefore, the aim of this study was to investigate the clinical signs, blood parameters, post mortem and histopathology changes caused by P. multocida Type B:2 infections initiated through the oral and subcutaneous routes. Methods: Nine buffalo heifers were divided equally into 3 treatment groups. Group 1 was inoculated orally with 10 ml of phosphate buffer saline; Groups 2 and 3 were inoculated with 10 ml of 1012 colony forming unit of P. multocida Type B:2 subcutaneously and orally respectively. Results: There was a significant difference (p

We report the serotyping of foot-and-mouth disease virus (FMDV) and Pasteurella multocida from Indian gaurs which were concurrently infected with foot-and-mouth disease (FMD) and haemorrhagic septicaemia. Bannerghatta biological park (BBP), a national park located in the outskirts of Bengaluru city, Karnataka, India, is bordered by several villages. These villages witnessed massive outbreaks of FMD which spread rapidly to the herbivores at BBP. Post-mortem was conducted on carcasses of two Indian gaurs that died with symptoms of FMD. The salient gross findings included extensive vesicular lesions on the tongue, gums, cheeks, upper palate and hooves. Haemorrhagic tracheitis and ecchymotic haemorrhages on the heart were characteristic. The vesicular lesions of oral cavity were positive for 'O' type of FMD virus by sandwich enzyme-linked immuno sorbent assay (ELISA). The heart blood and spleen samples yielded growth of pure cultures of P. multocida. The isolates were typed as P. multocida type B using KTSP61 and KTT72 primers yielding specific amplicons of 620 bp. The phylogenetic analysis of the isolates was carried by sequencing of 1.4-Kbp nucleotides on the 16S ribosomal RNA (rRNA) gene of the isolates.

Aeromonas hydrophila is the most common water fish pathogen and cause diseases such as hemorrhagic septicemia, dropsy, ulceration and asymptomatic septicemia. A. hydrophila secretes many extracellular products (ECPs) which contribute to effective infection, wide distribution and great adaptability to environmental changes. Crude ECPs of A. hydrophila CK257, a strain used in this study, exhibits a toxic activity to the animals including mouse, rabbit and fish. Toxic symptoms were indicated by tissue damage and skin injuries in animal. When ECPs were subcutaneously injected to animals, skin damages were observed, appearing like necrosis. Preliminary research demonstrated that the active factors are protein component. The crude ECPs were collected after ammonium sulfate precipitation of cell-free culture supernatant. ECPs were fractionated with the use gel filtration chromatography. Five ECP fractions were obtained, of which one fraction was found to be toxic to goldfish. MALDI-TOF analyses provided two interesting proteases called M35 and M28. Both M35 and M28 are known as metalloprotease. Accordingly, proteins in an active fraction exhibited caseinolytic activity. These proteins were difference of caseinolytic activity under different metallic ions. Also active fraction has elastolytic activity. These results suggested that peptidase M28 and M35 may be a candidate factor for tissue necrosis activity about infection with A. hydrophila.