Pasteurella Multocida Strains Research Articles

Genital Form of Pasteurellosis in Breeding Turkeys Infected During Artificial Insemination and Isolation of an Unusual Strain of Pasteurella multocida

A genital and potentially fatal form of Pasteurella multocida infection was reported on two turkey-breeding farms on which birds were vaccinated against Pasteurella multocida. Both outbreaks were linked to the use of semen from young vaccinated toms with a history of respiratory pasteurellosis followed by treatment during rearing. Typing by agar gel immunodiffusion and rapid slide agglutination of P. multocida isolated from cloacal swabs was completed by multilocus sequence typing. Restriction enzyme analysis showed that that the isolates were clonal. They belonged to sequence type (ST) 30, described in chickens, cats, and ducks. This strain differed in sequence type from the ones used in the vaccine (ST8, ST60, ST53, and ST235), which might have limited its effectiveness. No contamination of the semen (n = 30) was found, suggesting fecal contamination during semen collection.

Pharmacokinetic/pharmacodynamic relationship of marbofloxacin against Pasteurella multocida in a tissue‐cage model in yellow cattle

The fluoroquinolone antimicrobial drug marbofloxacin was administered to yellow cattle intravenously and intramuscularly at a dose of 2 mg/kg of body weight in a two-period crossover study. The pharmacokinetic properties of marbofloxacin in serum, inflamed tissue-cage fluid (exudate), and noninflamed tissue-cage fluid (transudate) were studied by using a tissue-cage model. The in vitro and ex vivo activities of marbofloxacin in serum, exudate, and transudate against a pathogenic strain of Pasteurella multocida (P.multocida) were determined. Integration of in vivo pharmacokinetic data with the in vitro MIC provided mean values for the area under the curve (AUC)/MIC for serum, exudate, and transudate of 155.75, 153.00, and 138.88, respectively, after intravenous dosing and 160.50, 151.00, and 137.63, respectively, after intramuscular dosing. After intramuscular dosing, the maximum concentration/MIC ratios for serum, exudate, and transudate were 21.13, 9.13, and 8.38, respectively. The ex vivo growth inhibition data after intramuscular dosing were fitted to the inhibitory sigmoid Emax equation to provide the values of AUC/MIC required to produce bacteriostasis, bactericidal activity, and elimination of bacteria. The respective values for serum were 17.25, 31.29, and 109.62, and slightly lower values were obtained for transudate and exudate. It is proposed that these findings might be used with MIC50 or MIC90 data to provide a rational approach to the design of dosage schedules which optimize efficacy in respect of bacteriological as well as clinical cures.

Pharmacokinetic/pharmacodynamic relationship of cefquinome against Pasteurella multocida in a tissue-cage model in yellow cattle

The cephalosporin antimicrobial drug cefquinome was administered to yellow cattle intravenously (i.v.) and intramuscularly (i.m.) at a dose of 1mg/kg of body weight in a two-period crossover study. The pharmacokinetic (PK) properties of cefquinome in serum, inflamed tissue-cage fluid (exudate), and noninflamed tissue-cage fluid (transudate) were studied using a tissue-cage model. The in vitro and ex vivo activities of cefquinome in serum, exudate, and transudate against a pathogenic strain of Pasteurella multocida (P.multocida) were determined. A concentration-independent antimicrobial activity of cefquinome was confirmed for levels lower than 4×MIC. Integration of in vivo pharmacokinetic data with the in vitro MIC provided mean values for the time that drug levels remain above the MIC (T>MIC) in serum was 14.10h after intravenous and 14.46h after intramuscular dosing, indicating a likely high level of effectiveness in clinical infections caused by P.multocida of MIC 0.04μg/mL or less. These data may be used as a rational basis for setting dosing schedules, which optimize clinical efficacy and minimize the opportunities for emergence of resistant organisms.

Evidence of circulation of an epidemic strain of Pasteurella multocida in Jiangsu, China by multi-locus sequence typing (MLST)

Pasteurella multocida, the causative agent of fowl cholera, is a serious threat to poultry farming. In this study, we isolated and identified 40 P. multocida strains in fowl cholera outbreaks in Jiangsu province, China. The identified P. multocida was further characterized using multi-locus sequence typing (MLST). All of the 40 P. multocida strains studied are genetically identical and belong to the ST129 sequence type based on seven MLST loci. Our study provides evidence of a circulating epidemic strain of P. multocida in Jiangsu, China.

Molecular typing of haemorrhagic septicaemia-associated Pasteurella multocida isolates from Pakistan and Thailand using multilocus sequence typing and pulsed-field gel electrophoresis

A comparative genetic study of 23 field isolates and vaccine strains of Pasteurella multocida associated with haemorrhagic septicaemia cases from Pakistan and Thailand was done using pulsed field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). The MLST sequence type (ST) for all 20 of the 23 isolates tested was 122. The PFGE results showed one band difference between the Pakistani and the Thai isolates. Sequence type 122 is the dominant associated profile with haemorrhagic septicaemia (HS) cases in South Asia. The study supports the concept of using PFGE for short-term epidemiology and MLST for long-term epidemiology.

Phenotypic and Genotypic Characterisation of Pasteurella Multocida Strains Isolated from Pigs in Poland

Abstract A total of 319 Pasteurella multocida (Pm) strains isolated from pigs in Poland were examined. Phenotypic characterisation included: biochemical tests (to determine species, subspecies, and biovar), capsular typing, and antimicrobial susceptibility. Genotypic characterisation included detection of the toxA gene by PCR. All tested Pm strains were classified as Pm subsp. multocida: 87.2% biovar 3, 10.7%-2 and 0.9%-12. One strain was classified as biovar 1. Three strains of Pm did not suit any of the biovars. Using capsular typing methods, 77% of Pm strains isolated from nasal swabs belonged to type D and 33% to type A. Among Pm strains isolated from internal organs, 59.5% belonged to type A and 40.5% to type D. All the isolates showed a high susceptibility to β-lactams: ampicillin and amoxicilin with clavulonic acid (97.8%), penicillin (86.7%), doxicilline (100%), oxytetracycline (97.8%), and tetracycline (93.2%). It was found that all strains were susceptible to norfloxacin, 97.8% to enrofloxacin, and 95.6% to SxT. 24.4% and 15.6% of the strains were resistant to linco-spectin and tiamulin, respectively. The presence of toxA gene was confirmed by PCR in 20.8% of the strains isolated from nasal swabs and 29.1% of isolates from internal organs.

Some Studies on Rabbit Hemorrhagic Septicemia Vaccines in Egypt

In this study, a trivalent autogenous inactivated P.multocida vaccine was prepared. The prepared vaccine contained three different P. multocida strains which were previously isolated from different localities at Sharkia Governorate and identified by biochemical and molecular methods. The protective effect of the prepared vaccine was evaluated through estimation of the humeral immune response using indirect heamagglutination test (IHA); challenge infection and field evaluation. Pasteurella multocida strains were inactivated with formalin (0.5%) and adjuvanted with montanide oil (ISA -70). Repeated immunization raised up the specific antibody level eight times in vaccinated rabbits. It conferred protection against challenge with homologous strains in commercial rabbits. In the field study a positive effect of vaccination on nasal colonization with P. multocida was found.

Seven strains of mice as potential models of bovine pasteurellosis following intranasal challenge with a bovine pneumonic strain of Pasteurella multocida A:3; comparisons of disease and pathological outcomes

The Gram-negative bacterium Pasteurella multocida causes pneumonic and systemic pasteurellosis in bovids for which vaccines are either unavailable or inadequate. The work assessed whether an intranasal P. multocida challenge in mice might provide a model of infection for future vaccine development work. Clinical, pathological and biochemical responses were compared in seven strains of mice challenged with a virulent bovine pneumonic isolate of P. multocida A:3. Six mouse strains (Porton, CD-1, BALB/c, VM, C57BL/10 and C57BL/6) developed clinical signs of pneumonic disease and variable pneumonic lesions 41–70h post-infection. In contrast, mouse strain RIII became septicaemic within 36h post-infection. Concentrations of plasma acute phase proteins and serum lipopolysaccharide increased in all mice after infection, and the main or interaction effect of mouse strain and infection status was statistically significant (P<0.05). Responses in C57BL/10 mice showed close similarity to bovine pneumonic and in RIII mice to bovine systemic pasteurellosis.

Draft Genome Sequences of Two Virulent Serotypes of Avian Pasteurella multocida.

Here we report the draft genome sequences of two virulent avian strains of Pasteurella multocida. Comparative analyses of these genomes were done with the published genome sequence of avirulent P. multocida strain Pm70.

Characterization of Pasteurella multocida strains isolated from geese

Phenotypic and genotypic diversity of forty-two Pasteurella multocida isolates from geese were characterized by analysis of their capsular type, Heddleston serotype, biotype, fimbrial gene allele type, comparative outer membrane protein (OMP) electrophoresis patterns, and were analyzed using PCR for the presence of virulence-associated genes (toxA, tbpA, pfhA, hgbA, hgbB, nanH, nanB, fimA, hsf-1, and pmHAS). A sequence comparison of the thdF and rpoB housekeeping genes of twenty representative P. multocida strains from three different OMP groups demonstrated that seventeen strains were closely related phylogenetically to previously published strains of P. multocida subsp. multocida and P. multocida subsp. gallicida, and only three strains from geese were grouped with previously published strains of P. multocida subsp. septica. This study is the first report regarding the characterization of phenotypic and genotypic features of different P. multocida field strains isolated from geese with the different clinical representation of pasteurellosis and provide our overview on the usefulness of these in vitro tests for primary characterization of P. multocida strains for the epidemiological studies of fowl cholera.

Comparative genome analysis of an avirulent and two virulent strains of avian Pasteurella multocida reveals candidate genes involved in fitness and pathogenicity

BackgroundPasteurella multocida is the etiologic agent of fowl cholera, a highly contagious and severe disease of poultry causing significant mortality and morbidity throughout the world. All types of poultry are susceptible to fowl cholera. Turkeys are most susceptible to the peracute/acute forms of the disease while chickens are most susceptible to the acute and chronic forms of the disease. The whole genome of the Pm70 strain of P. multocida was sequenced and annotated in 2001. The Pm70 strain is not virulent to chickens and turkeys. In contrast, strains X73 and P1059 are highly virulent to turkeys, chickens, and other poultry species. In this study, we sequenced the genomes of P. multocida strains X73 and P1059 and undertook a detailed comparative genome analysis with the avirulent Pm70 strain. The goal of this study was to identify candidate genes in the virulent strains that may be involved in pathogenicity of fowl cholera disease.ResultsComparison of virulent versus avirulent avian P. multocida genomes revealed 336 unique genes among the P1059 and/or X73 genomes compared to strain Pm70. Genes of interest within this subset included those encoding an L-fucose transport and utilization system, several novel sugar transport systems, and several novel hemagglutinins including one designated PfhB4. Additionally, substantial amino acid variation was observed in many core outer membrane proteins and single nucleotide polymorphism analysis confirmed a higher dN/dS ratio within proteins localized to the outer membrane.ConclusionsComparative analyses of highly virulent versus avirulent avian P. multocida identified a number of genomic differences that may shed light on the ability of highly virulent strains to cause disease in the avian host, including those that could be associated with enhanced virulence or fitness.

A 39-kDa Capsular Protein is a Major Cross-Protection Factor as Demonstrated by Protection of Chickens with a Live Attenuated &lt;i&gt;Pasteurella multocida&lt;/i&gt; Strain of P-1059

The aim of this study was to show that a 39-kDa protein or OmpH of Pasteurella multocida strain P-1059 is essential for cross protection. Strain PBA322, a thinly capsulated strain of P. multocida strain P-1059, was used as a live vaccine in chickens. Strain PBA322 is a thinly capsulated strain in comparison with the parental strain P-1059. Chickens were vaccinated by single injection and then challenge-exposed with strains P-1059 or X-73 at two weeks post vaccination. Moreover, immune responses were also evaluated for both humoral and cellular immune response by ELISA and lymphocyte proliferation assay, respectively. The results showed that the live vaccine induced efficient immunity to protect chickens from challenge-exposure to the parent strain, but that the heterologous protection was poor. We concluded that the 39-kDa protein is essential for cross protection.

A Case of a Late and Atypical Knee Prosthetic Infection by No-Biofilm Producer Pasteurella multocida Strain Identified by Pyrosequencing

Prosthetic joint infections due to Pasteurella multocida are rarely but increasingly reported but no data on production of biofilm are available. We report the case of a woman with a late, haematogenous peri-prosthetic infection of cemented total knee arthroplasty caused by a strain of P. multocida identified by pyrosequencing and unable to produce biofilm. Comparison of clinical and laboratory findings with those reported in other patients evidenced differences mainly in the period of symptoms' onset and in the behaviour of some inflammatory markers.

Molecular Epidemiologic Investigation of Polish Avian Pasteurella multocida Strains Isolated from Fowl Cholera Outbreaks Showing Restricted Geographical and Host-Specific Distribution

Molecular Epidemiologic Investigation of Polish Avian Pasteurella multocida Strains Isolated from Fowl Cholera Outbreaks Showing Restricted Geographical and Host-Specific Distribution

Molecular Epidemiologic Investigation of Polish Avian Pasteurella multocida Strains Isolated from Fowl Cholera Outbreaks Showing Restricted Geographical and Host-Specific Distribution

Molecular epidemiologic analyses of the 42 clinical isolates of Pasteurella multocida from various avian hosts (geese, ducks, turkeys, and laying hens) in Poland from 2001 to 2011, including a single reference strain, were performed by enterobacterial repetitive intergenic consensus (ERIC)-PCR, single primer PCR, and repetitive extragenic palindromic (REP)-PCR. Forty-two isolates were identified as P. multocida (serotype A). The majority of P. multocida strains were obtained from waterfowl clustered within one genotype, and they were not consistent with the genotypes obtained from the turkey strains. Pasteurella multocida showed genetic homogeneity between the host species, especially when isolated on the same farm. Some of the clones also were characteristic to the particular farm. The strains obtained in different regions represent distinct molecular patterns. The present findings demonstrate that some clones of P. multocida are restricted in geographical and host distribution. In addition, this study suggests that ERIC-PCR, single primer PCR, and REP-PCR are suitable techniques for studying the host adaptation of P. multocida and the epidemiology of fowl cholera.

Regulation of Toll-like receptor 4-mediated immune responses through Pasteurella multocida toxin-induced G protein signalling

BackgroundLipopolysaccharide (LPS)-triggered Toll-like receptor (TLR) 4-signalling belongs to the key innate defence mechanisms upon infection with Gram-negative bacteria and triggers the subsequent activation of adaptive immunity. There is an active crosstalk between TLR4-mediated and other signalling cascades to secure an effective immune response, but also to prevent excessive inflammation. Many pathogens induce signalling cascades via secreted factors that interfere with TLR signalling to modify and presumably escape the host response. In this context heterotrimeric G proteins and their coupled receptors have been recognized as major cellular targets. Toxigenic strains of Gram-negative Pasteurella multocida produce a toxin (PMT) that constitutively activates the heterotrimeric G proteins Gαq, Gα13 and Gαi independently of G protein-coupled receptors through deamidation. PMT is known to induce signalling events involved in cell proliferation, cell survival and cytoskeleton rearrangement.ResultsHere we show that the activation of heterotrimeric G proteins through PMT suppresses LPS-stimulated IL-12p40 production and eventually impairs the T cell-activating ability of LPS-treated monocytes. This inhibition of TLR4-induced IL-12p40 expression is mediated by Gαi-triggered signalling as well as by Gβγ-dependent activation of PI3kinase and JNK.Taken together we propose the following model: LPS stimulates TLR4-mediated activation of the NFĸB-pathway and thereby the production of TNF-α, IL-6 and IL-12p40. PMT inhibits the production of IL-12p40 by Gαi-mediated inhibition of adenylate cyclase and cAMP accumulation and by Gβγ-mediated activation of PI3kinase and JNK activation.ConclusionsOn the basis of the experiments with PMT this study gives an example of a pathogen-induced interaction between G protein-mediated and TLR4-triggered signalling and illustrates how a bacterial toxin is able to interfere with the host’s immune response.

Immunostimulant Effect of Egyptian Propolis in Rabbits

The present experiment was conducted to study the effect of ethanolic extract of Egyptian propolis given alone or in combination with inactivated Pasteurella multocida vaccine on rabbits challenged with a virulent strain of Pasteurella multocida. Fifty-six New-Zealand rabbits, 6–8 weeks old and non-vaccinated against pasteurellosis, were randomly divided into eight equal groups. The first group was kept as a control for the experiment. The other groups received different treatments with propolis extract, inactivated vaccine, or both. The experiment continued for seven weeks during which clinical signs, body weight, and mortality rate were monitored, and blood samples were collected weekly for evaluating the leukogram, serum biochemistry, and immune response in all groups of animals. At the end of the seventh week, the animals were subjected to challenge with a virulent strain of Pasteurella multocida. Two weeks later, tissue specimens were collected from different organs for histopathological examination. Results showed that rabbits of the groups treated with both propolis and the vaccine by different routes appeared healthy after challenge. It has been concluded that alcoholic extract of propolis administrated in combination with inactivated Pasteurella multocida vaccine has no adverse effects on the general health conditions and enhances immune response in rabbits.

Time-course investigation of infection with a low virulent Pasteurella multocida strain in normal and immune-suppressed 12-week-old free-range chickens

Twelve-week-old indigenous chickens, either immune-suppressed using dexamethasone (IS) or non-immune-suppressed (NIS), were challenged with a low virulent strain, Pasteurella multocida strain NCTC 10322T, and developed clinical signs and pathological lesions typical of chronic fowl cholera. NIS birds demonstrated much more severe signs of fowl cholera than IS birds. With few exceptions, signs recorded in IS and NIS birds were of the same types, but significantly milder in the IS birds, indicating that immune suppression does not change the course of infection but rather the severity of signs in fowl cholera. P. multocida signals by fluorescent in situ hybridization (FISH) were observed between 1 h and 14 days in the lungs, trachea, air sacs, liver, spleen, bursa of Fabricius and caecal tonsils, while signals from other organs mostly were observed after 24 h. More organs had FISH signals in NIS birds than in IS birds and at higher frequency per organ. Many organs were positive by FISH even 14 days post infection, and it is suggested that these organs may be likely places for long-term carriage of P. multocida following infection. The present study has demonstrated the spread of P. multocida in different tissues in chickens and distribution of lesions associated with chronic fowl cholera, and pointed to a decrease of pathology in IS birds. Since dexamethasone mostly affects heterophils, the study suggests that these cells play a role in the development of lesions associated with chronic fowl cholera in chickens.

Characterization of Pasteurella multocida strains isolated from cattle and buffaloes in Karachi, Pakistan

Pasteurella multocida is a Gram-negative, non-motile coccobacillus. The organism is of special significance in the tropical countries including Pakistan where only in Punjab Province, it caused heavy economics losses as Rs.2.17 billion per year. We Isolated and identified the disease causing serotypes in Pakistan by using cultural, morphological and biochemical tests. Pathogenicity was confirmed by inoculation of the isolates in rabbits. Serotyping of the isolates was done and our isolates were found to be P. multocida , B: 2 serotype. Polymerase chain reaction (PCR) of 16s ribosomal deoxyribonucleic acid (rDNA) was done which amplified 1500 bp deoxyribonucleic acid (DNA) fragment. DNA sequencing and phylogenetic analysis of the gene revealed that our isolates are different from each other and they belong to two different groups. Whereas these isolates are closely related with P. multocida, sub species multocida, P. multocida and sub species septica.

PmST3 from Pasteurella multocida encoded by Pm1174 gene is a monofunctional α2–3-sialyltransferase

Pasteurella multocida (Pm) strain Pm70 has three putative sialyltransferase genes including Pm0188, Pm0508, and Pm1174. A Pm0188 gene homolog in Pm strain P-1059 encodes a multifunctional α2-3-sialyltransferase, PmST1, that prefers oligosaccharide acceptors. A Pm0508 gene homolog in the same strain encodes a monofunctional sialyltransferase PmST2 that prefers glycolipid acceptors. Here, we report that the third sialyltransferase from Pm (PmST3) encoded by gene Pm1174 in strain Pm70 is a monofunctional α2-3-sialyltransferase that can use both oligosaccharides and glycolipids as efficient acceptors. Despite the existence of both Pm0188 and Pm0508 gene homologs encoding PmST1 and PmST2, respectively, in Pm strain P-1059, a Pm1174 gene homolog appears to be absent from Pm strains P-1059 and P-934. PmST3 was successfully obtained by cloning and expression using a synthetic gene of Pm1174 with codons optimized for Escherichia coli expression system as the DNA template for polymer chain reactions. Truncation of 35 amino acid residues from the carboxyl terminus was shown to improve the expression of a soluble and active enzyme in E. coli as a C-His(6)-tagged fusion protein. This sialidase-free monofunctional α2-3-sialyltransferase is a useful tool for synthesizing sialylated oligosaccharides and glycolipids.