Haemorrhagic septicaemia is a disease caused by Pasteurella multocida serotype B: 2 and E: 2. The organism causes acute, highly fatal septicaemic disease with high morbidity and mortality in cattle and more susceptible in buffaloes. Lipopolysaccharide can be found on the outer cell wall of the organism. Lipopolysaccharide is released during multiplication which leads to inflammatory reaction. It represents the endotoxin of P. multocida type B: 2 and responsible for toxicity in haemorrhagic septicaemia which plays an important role in the pathogenesis of the disease. Therefore, the aim of this study was to investigate the clinical signs, blood parameters, gross post mortem lesions and histopathology changes caused by P. multocida type B:2 immunogen lipopolysaccharide infections initiated through intravenous and oral routes of infection. 9 buffalo heifers were divided equally into 3 treatment groups. Group 1 was inoculated orally with 10 ml of phosphate buffer saline (PBS); Group 2 and 3 were inoculated with 10 ml of lipopolysaccharide broth intravenously and orally respectively. For the clinical signs, there were significant differences (p < 0.05) in temperature between the control, intravenous and oral group. In hematology and biochemistry findings, there were significant differences (p < 0.05) in erythrocytes, haemoglobin, PCV, MCV, lymphocytes, monocytes, eosinophils, GGT and albumin between the control, intravenous and oral group. However, there were no significant differences (p > 0.05) in the MCHC, leukocytes, band neutrophils, basophils, thrombocytes, plasma protein, icterus index, total protein, globulin and A:G ratio between intravenous and oral group. For Group 2 buffaloes, there were gross lesions in the lung, trachea, heart, liver, spleen, and kidney. In contrast, lesions were only observed in the lung, trachea and liver of Group 3 buffaloes. There were significant differences (p < 0.05) in hemorrhage and congestion; necrosis and degeneration; and inflammatory cells infiltration between experimental groups and control group. However, there were no significant differences (p > 0.05) in edema lesion between groups. In conclusion, this study is a proof that oral route infection of P. multocida type B:2 immunogen lipopolysaccharide can be used to stimulate host cell responses where oral vaccine through feed could be developed in the near future.
Although more than 100 genome sequences of Pasteurella multocida are available, comprehensive and complete genome sequence analysis is limited. This study describes the analysis of complete genome sequence and pathogenomics of P. multocida strain PMTB2.1. The genome of PMTB2.1 has 2176 genes with more than 40 coding sequences associated with iron regulation and 140 virulence genes including the complete tad locus. The tad locus includes several previously uncharacterized genes such as flp2, rcpC and tadV genes. A transposable phage resembling to Mu phages was identified in P. multocida that has not been identified in any other serotype yet. The multi-locus sequence typing analysis assigned the PMTB2.1 genome sequence as type ST101, while the comparative genome analysis showed that PMTB2.1 is closely related to other P. multocida strains with the genomic distance of less than 0.13. The expression profiling of iron regulating-genes of PMTB2.1 was characterized under iron-limited environment. Results showed significant changes in the expression profiles of iron-regulating genes (p < 0.05) whereas the highest expression of fecE gene (281 fold) at 30 min suggests utilization of the outer-membrane proteins system in iron acquisition at an early stage of growth. This study showed the phylogenomic relatedness of P. multocida and improved annotation of important genes and functional characterization of iron-regulating genes of importance to the bacterial growth.
One hundred and fourteen strains of Pasteurella multocida were isolated from different domestic animals species (cattle, buffalo, sheep, goat, pig, rabbit, dog, cat), avian species (chicken, duck, turkey) and wild animals (deer, tiger, orang utan, marmoset). The serogroups of P. multocida were determined by both conventional capsular serotyping and a multiplex PCR assay targeting specific capsular genes. Based on the conventional serotyping method, the 114 strains of P. multocida were subtyped into 55 species-specific (untypeable strains) P. multocida, 15 serogroup A, 23 serogroup B and 21 serogroup D. Based on the multiplex PCR assay on the specific capsular genes associated with each serogroup, the 114 strains were further divided to 22 species-specific P. multocida (KMT1 - 460 bp), 53 serogroup A (A - 1,044 bp), 33 serogroup B (B - 760 bp) and 6 serogroup D (D - 657 bp). No serogroup E (511 bp) or F (851 bp) was detected among the Malaysian P. multocida. PCR-based typing was more discriminative and could further subtype the previously untypeable strains. Overall, there was a significant and positive correlation between both methods in serogrouping P. multocida (r = 0.7935; p<0.4893). Various serogroups of P. multocida were present among the livestock with 75% of the strains belonging to serogroups A or B. PCR serotyping was therefore a highly species-specific, sensitive and robust method for detection and differentiation of P. multocida serogroups compared to conventional serotyping. To the best of our knowledge, this is the first report from Malaysia of the application of a PCR to rapidly define the species-specific P. multocida and its serogroups as an important zoonotic pathogen in Malaysia.
Haemorrhagic septicaemia (HS) is an acute disease of cattle and buffaloes caused by Pasteurella multocida 6:B. Outbreaks of the disease have been closely associated with carrier animals that transmit the organism to susceptible animals during stressful condition. This study was conducted to determine whether goats exposed intranasally to P. multocida 6:B can transmit the organism to contact goats. Thirty-six healthy local Katjang goats were divided into four groups and goats of groups 1 and 3 were each inoculated intranasally with a 1-ml inoculum that contained 1 x 10(9) CFU/ml of live P. multocida 6:B. Following the exposure, all goats of groups 3 and 4 were injected with dexamethasone at the rate of 1 mg/kg for three consecutive days. At the end of the dexamethasone treatment, goats of groups 1 and 2 were commingled but kept separate from goats of groups 3 and 4, which were commingled in another pen. Three surviving goats from each group were killed on days 7, 14 and 21 post-exposure for postmortem examination. Naso-pharyngeal mucus and heart blood were collected on swabs. Tissues from lungs, lymph nodes and tonsils were collected for bacteriological isolation and identification. Only one goat of group 3 died 6 days post-exposure showing clinical signs and lesions typical of HS. Other goats showed mild signs of upper respiratory tract infection. Goats of all groups developed acute mild pneumonic lesions, however, those treated with dexamethasone had significantly (P < 0.05) more extensive lesion scoring based on the lesion scoring system. P. multocida 6:B was isolated from the nasal mucosa and lung lesions of exposed and contact goats not treated with dexamethasone. Exposed and contact goats treated with dexamethasone carried the organism for 21 days. P. multocida isolation from heart blood was made only from exposed and contact goats treated with dexamethasone. P. multocida was isolated from the lymph node of the goat that died during the experiment.
Pasteurella multocida B:2 is a Gram-negative organism causing haemorrhagic septicaemia (HS) in buffaloes. It causes severe pulmonary infection, leading to infiltration of numerous macrophages and neutrophils. Despite the inflammatory response, buffaloes succumb to HS. This study aims to evaluate the in-vitro efficacy of macrophages and neutrophils of buffalo following exposure to P. multocida B:2. In-vitro infections were done using 107 cfu/ml of P. multocida B:2 for Group 1, Escherichia coli for Group 2 and Mannhaemia haemolytica A:2 for Group 3 cells. The inoculated cell cultures were harvested at 0, 30, 60 and 120 min post-exposure and the phagocytic, killing and cell death rates were determined. Both phagocytosis and killing rates of all bacteria increased over time. Phagocytosis involved between 71% and 73% neutrophils and between 60% and 64% macrophages at 120 min. Killing rate of all bacteria involved between 76% and 79% for neutrophils and between 70% and 74% for macrophages at 120 min. Death rate of neutrophils ranged between 67% in Group 3, and 88% in Group 1 at 120 min, significantly (p 0.05) than Group 2. Similar pattern was observed for death rate of macrophages. The phagocytosis and killing rates of P. multocida B:2 were similar to other bacterial species used in this study but more neutrophils and macrophages were dead following infection by P. multocida B:2 than M. haemolytica A:2.
In vitro experiments were undertaken to study the adhesion and colonization to tracheal mucosa, lung and aorta explants from freshly killed rabbits of two different strains of Pasteurella multocida. Serotype A:3 (capsulated, fimbriae +, haemagglutination -, dermonecrotic toxin -) isolated from a rabbit with rhinitis, and serotype D:1 (non-capsulated, fimbriae +, haemagglutination +, dermonecrotic toxin +) isolated from a dead rabbit with septicaemia, were used. When the explants were observed under the scanning electron microscope, the type D strain was highly adherent to trachea and aorta explants compared to the type A strain. Adhesion to lung explants was best achieved by the type A strain after 45 min incubation, but after 2 h incubation no significant difference was observed between the strains. Our data indicate that the presence of fimbriae and the absence of capsule seem to enhance the adherence of P. multocida type D strain to tracheal tissue. The capsular material of P. multocida type A strain and the toxin of the type D strain seem to influence the adherence to lung tissue in rabbit. Adhesion of strain D to aorta may indicate the expression of receptors on the endothelium to that strain and may also explain the ability of certain strains to cause septicaemia.
The objectives of this study were to determine the prevalence, characterization and antibiotic resistance of Pasteurella multocida isolated from calves with respiratory infection in Iran. P. multocida was detected in 141/169 bovine respiratory infection cases on Iranian dairy and beef farms. P. multocida were grouped into serogroups A (126/141), D (12/141), and B (3/141). Of the P. multocida isolates, all harboured the psl, ompH, oma87, fimA, ptfA, nanB, and nanH genes, 139/141 had hsf-2, and 115/141 pfhA, and tadD. The isolates were most frequently resistant to penicillin G (43/141 resistant isolates; 30.5%) and streptomycin (31/141; 22%).
Clinical and pathological changes are described in groups of five goats pretreated with dexamethasone and then infected with a large dose of Pasteurella multocida B:2 (the cause of haemorrhagic septicaemia) by the intratracheal, subcutaneous or intranasal route (groups A, B and C, respectively). In group A, two goats died (on day 1 and 4 post-inoculation); in group B three died (days 2, 5 and 14); and in group C one died (day 20). The infecting organism was recovered from the four goats that died within < or =5 days. The major pulmonary lesions included acute pneumonia, congestion, oedema and hydrothorax. Subcutaneous oedema of the lower jaw and brisket, typically seen in cattle and buffalo, was absent in goats.
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
Sixteen 8- to 9-week-old Pasteurella multocida-free rabbits were divided into two equal groups. Eight rabbits in one group were inoculated intranasally with P. multoida type A:3. The other eight were inoculated intranasally with phosphate-buffered saline and used as controls. Nasal swabs taken before and after inoculation were cultured for bacterial isolation. Post-mortem nasal swabs and lung samples were cultured for bacteriological isolation. Nasal mucosa and lung samples were collected and processed for transmission electron microscopy. Pasteurella multocida was isolated from the nasal cavity of all infected rabbits and from the lungs of four infected rabbits. Degenerative ultrastructural changes in epithelial cells and endothelial cells were seen in the infected rabbits. Deciliation of the ciliated epithelium and hyperplasia of the goblet cells in the nasal mucosa were noted. Thickening of the alveolar septa due to hyperplasia of type II pneumocytes, swelling of the endothelial lining of capillaries and infiltration of inflammatory cells were also observed. Intracellular invasion of the nasal epithelial cells and of type II pneumocytes by the organism was observed. Coccobacilli were observed in membrane-bound vacuoles in the cytoplasm of these cells. The vacuoles were adjacent to the host-cell mitochondria and some of these vacuoles appeared to be fused to the mitochondrial membrane. Some type I pneumocytes with intracellular membrane-bound vacuoles containing bacterial cells showed protrusions, which appeared to detach into the alveolar lumina. These results indicated that P. multocida serotype A:3 in rabbits can invade the epithelial cell and cause structural changes in the interstitium, epithelium and endothelium. Heterophils and macrophages appear to play important roles in tissue injury.
BACKGROUND: Pasteurella multocida B:2 causes haemorrhagic septicaemia in cattle and buffaloes. However, buffaloes are found to be more susceptible to the infection than cattle. Upon infection, the pathogen rapidly spread from the respiratory tract to the blood circulation within 16-72 h, causing septicaemia. So far, limited study has been conducted to evaluate the response of endothelial cells of buffalo towards P. multocida B:2 and its lipopolysaccharide (LPS). This study aimed to evaluate the ultrastructural changes in the aortic endothelium of buffaloes (BAEC) following exposure to P. multocida B:2 and its endotoxin. The endothelial cells were harvested from the aorta of healthy buffaloes and were prepared as monolayer cell cultures. The cultures were divided into 3 groups before Group 1 was inoculated with 107 cfu/ml of whole cell P. multocida B:2, Group 2 with LPS, which was extracted earlier from 107 cfu/ml of P. multocida B:2 and Group 3 with sterile cell culture medium. The cells were harvested at 0, 6, 12, 18, 24, 36, and 48 h post-inoculation for assessment of cellular changes using transmission electron microscopy.
RESULTS: The BAEC of Groups 1 and 2 demonstrated moderate to severe endothelial lysis, suggestive of acute cellular injury. In general, severity of the ultrastructural changes increased with the time of incubation but no significant difference (p > 0.05) in the severity of the cellular changes between Groups 1 and 2 was observed in the first 18 h. The severity of lesions became significant (p
This study aimed to determine the effect of intranasal exposure to low doses of Pasteurella multocida B:2 on survival of goats challenged with high doses of the same organism. Eighteen goats were selected and divided into three groups. Goats of group 1 were exposed intranasally twice, with a two-week interval, to 7 x 10(6) cfu/ml of live P. multocida B:2. Goats of group 2 were not exposed to P. multocida B:2 but were kept together with the exposed group 1. Goats of group 3 remained as unexposed controls and were kept separated from the other two groups. Serum samples were collected at weekly intervals to determine the antibody levels. At week 5 post exposure, all goats were challenged subcutaneously with 3.7 x 10(10) cfu/ml of live P. multocida B:2. Following challenge exposure, 8 (67%) goats (4 goats from each of groups 1 and 2) were killed owing to haemorrhagic septicaemia. Four goats were killed peracutely within 48 h post challenge, while the other four goats were killed acutely between 2 and 4 days post challenge. None of the goats of group 3 were killed for haemorrhagic septicaemia. Goats of groups 1 and 2 showed significantly (p < 0.05) higher antibody levels following the first intranasal exposure to P. multocida B:2. However, only group 1 retained the significantly (p < 0.05) high antibody levels following a second intranasal exposure, and remained significantly (p < 0.05) higher than groups 2 and 3 at the time of challenge. P. multocida B:2 was successfully isolated from various organs of goats that were killed between 1 and 4 days post challenge.
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.
Pasteurella multocida causes pneumonic pasteurellosis and hemorrhagic septicemia (HS) in large ruminants. In this study, we determined the complete genome sequence of P. multocida strain PMTB2.1 capsular serotype A isolated from buffaloes that died of septicemia.
A previously healthy woman in her mid-70s presented with right upper quadrant abdominal pain, fever, intermittent chills and malaise for 1 week. She was clinically septic with raised inflammatory markers. Her blood culture revealed Pasteurella multocida, which was susceptible to penicillin and amoxicillin-clavulanic acid. CT of liver revealed an abscess of 8.0×7.9×8.5 cm at the left lobe of the liver. However, the abscess was not amenable for surgical or radiological drainage. She was a farmer and had close contact with her pet cats. She was occasionally scratched by her cats when caring for them. The liver abscess resolved completely without drainage after prolonged antimicrobial therapy of 109 days. She commenced on 63 days of intravenous antimicrobials and 46 days of oral amoxicillin-clavulanic acid. This case illustrated P. multocida bacteraemia with a large liver abscess in an immunocompetent adult after non-bite exposure.
Pasteurella multocida serotypes B:2 and E:2 are the main causative agents of ruminant hemorrhagic septicemia in Asia and Africa, respectively. Pasteurella multocida strain PMTB was isolated from a buffalo with hemorrhagic septicemia and has been determined to be serotype B:2. Here we report the draft genome sequence of strain PMTB.
We report here the first high-quality draft genome sequence of Pasteurella multocida sequence type 128, which was isolated from the infected finger bone of an adult female who was bitten by a domestic dog. The draft genome will be a valuable addition to the scarce genomic resources available for P. multocida.
The aim of this study was to investigate the clinico-pathology and haemato-biochemistry alterations in buffaloes inoculated with Pasteurella multocida type B:2 immunogen outer membrane protein via subcutaneous and oral routes. Nine buffalo heifers were divided equally into 3 treatment groups. Group 1 was inoculated orally with 10 mL of phosphate buffer saline (PBS); Group 2 and 3 were inoculated with 10 mL of outer membrane protein broth subcutaneously and orally respectively. Group 2 buffaloes showed typical haemorrhagic septicaemia clinical signs and were only able to survive for 72 h of the experiment. However, Group 3 buffaloes were able to survive throughout the stipulated time of 21 days of experiment. There were significant differences (p 0.05) in edema between groups except for the lung. This study was a proof that oral route infection of Pasteurella multocida type B:2 immunogen outer membrane protein can be used to stimulate host cell.