Identification of virulence associated markers in the cell wall of pigeon Streptococcus gallolyticus strains

The cell wall protein profiles of 56 isolates of Streptococcus gallolyticus of differing virulence for pigeons were compared by SDS-PAGE. Additionally, Western blot analysis was performed on the cell wall proteins of 14 strains using sera of pigeons, experimentally infected with A(+)T1 or A(-)T2 strains of S. gallolyticus. The profile of silver stained gels exhibited a complex array of 20-50 bands ranging from less than 6.5-210kDa. A band with molecular mass of 114kDa was only observed in isolates that belonged to the highly virulent A(+)T1, A(+)T2, A(+)T3 and A(-)T1 culture supernatant groups. A band with a slightly higher molecular mass (115kDa) as well as a 207kDa band were only detected in isolates that belonged to the moderately A(-)T3 or low A(-)T2 virulent culture supernatant groups. The 114 and 115kDa band were recognised by all homologous and heterologous pigeon sera used whereas the 207kDa band was only recognised by sera of pigeons infected with a A(-)T2 strain. These findings may indicate that the 114, 115 and 207kDa bands are useful as additional virulence associated markers for pigeon S. gallolyticus strains.     

Prevalence of streptococcus bovis in racing pigeons

Summary

The prevalence of S. bovis in the intestinal tract of healthy racing pigeons was determined. Crop and cloaca swab samples obtained from 810 pigeons from 14 different lofts and from 122 pigeons that were presented for routine health control were examined for the presence of S. bovis. Pooled faecal samples were also obtained from pigeons in 82 different pigeon lofts. S. bovis was isolated from crop or cloaca samples of approximately 40 % of pigeons of all ages by direct culture and from 80 % of the pooled faecal samples by enrichment culture.

In a longitudinal study, crop and cloaca samples were collected every 3 months from pigeons in seven different pigeon lofts. The prevalence of S. bovis in these pigeons ranged from 0 to 100 %. The carriage rate was not related to the season or to the age of the pigeons.

The prevalence of S. bovis in organ lesions of pigeons examined at necropsy was investigated over a 35‐month period. S. bovis was isolated from 10 % of the birds examined. The incidence of S. bovis septicaemia was significantly higher in January to August than in September to December. It was concluded that S. bovis is an opportunistic pathogenic agent in pigeons.     

Brucella canis induces canine CD4+ T cells multi-cytokine Th1/Th17 production via dendritic cell activation

Brucella canis is a small intracellular Gram-negative bacterium that frequently leads to chronic infections highly resistant to antibiotic therapy in dogs. Also, it causes mild human brucellosis compared to other zoonotic Brucella spp. Herein we characterize the cellular immune response elicited by B. canis by analysing human and canine CD4⁺ T cells after stimulation with autologous monocyte-derived dendritic cells (MoDCs). Human and canine B. canis-primed MoDCs stimulated autologous CD4⁺ T cells; however, a Th1 response was triggered by human MoDCs, whereas canine MoDCs induced Th1/Th17 responses, with increased CD4⁺ T cells producing IFN-γ and IL-17A simultaneously. Each pattern of cellular response may contribute to host susceptibility, helping to understand the differences in B. canis virulence between these two hosts. In addition, other aspects of canine immunology are unveiled by highlighting the participation of IL-17A-producing canine MoDCs and CD4⁺ T cells producing IFN-γ and IL-17A.     

Streptococcus gallolyticus infections in racing pigeons, a literature review

S. gallolyticus, formerly known as S. bovis is known since 1988 as a facultative pathogen of racing pigeons. Important clinical signs include acute mortality, inability to fly, lameness, weight loss and slimy green diarrhea. A pathognomonic sign at post mortem examination is the presence of well circumscribed areas of necrosis in the pectoral muscle. Furthermore tenosynovitis of the supracoracoid muscle and arthritis of the knee, shoulder and hock can be observed. In one study S. gallolyticus septicaemia was diagnosed in 10% of necropsied pigeons. Since S. gallolyticus was also isolated from nearly 40% of clinical healthy pigeons it is regarded as a facultative pathogen. Various biotypes, serotypes and culture supernatant phenotypes can be distinguished. Supernatant phenotypes are identified on the basis of the presence of either a T1, T2 or T3 protein triplet and the presence or absence of an extracellular A protein. S. gallolyticus strains with A protein are highly virulent, while strains with only T3 or T2 protein are of moderately or low virulence respectively. Fimbriae are only seen in highly virulent and some of the moderately virulent strains. Possible virulence factors include survival in macrophages, adhesion to cells and toxin production. Infection with serotype 1 and 2 induces some degree of protection against re-infection with serotype 1, which offers perspectives for the development of a vaccine. Experimentally ampicillin, doxycycline and erythromycin have shown therapeutic effects. For the treatment of clinical cases the use of ampicillin is advocated, together with hygienic measures, such as the use of grid floors and avoiding overcrowding.     

In vitro effects of dexamethasone on bovine CD25+CD4+ and CD25−CD4+ cells

This paper investigates the in vitro effect of dexamethasone on bovine CD25^highCD4⁺, CD25^lowCD4⁺ and CD25⁻CD4⁺ T cells. Only a small percentage of bovine CD25^highCD4⁺ (2–4%) and CD25^lowCD4⁺ (1–2%) cells expressed Foxp3. Dexamethasone caused considerable loss of CD25⁻CD4⁺ cells, but it increased the relative and absolute numbers of CD25^highCD4⁺ and CD25^lowCD4⁺ lymphocytes, while at the same time reducing the percentage of Foxp3⁺ cells within the latter subpopulations. Considering all these, as well as the intrinsically poor Foxp3 expression in bovine CD25⁺CD4⁺, it can be concluded that the drug most probably increased the number of activated non-regulatory CD4⁺ lymphocytes. It has been found that changes in cell number were at least partly caused by proapoptotic effect of the drug on CD25⁻CD4⁺ cells and antiapoptotic effect on CD25^highCD4⁺ and CD25^lowCD4⁺ cells. The results obtained from this study indicate that the involvement of CD4⁺ lymphocytes in producing the anti-inflammatory and immunosuppressive effect of dexamethasone in cattle results from the fact that the drug had a depressive effect on the production of IFN-γ by CD25⁻CD4⁺ cells. Secretion of TGF-β and IL-10 by CD4⁺ lymphocytes was not involved in producing these pharmacological effects, because the drug did not affect production of TGF-β and, paradoxically, it reduced the percentage of IL-10⁺CD4⁺ cells.     

Antigenic relationships of Moraxella bovis isolates recovered from outbreaks of infectious bovine keratoconjunctivitis in Argentina, Brazil, and Uruguay between 1983 and 2000

Cross-reactivity indices (CRIs) of 28 isolates of Moraxella bovis recovered from outbreaks of infectious bovine keratoconjunctivitis in Argentina (A, 11 isolates), Brazil (B, 7), and Uruguay (U, 10) between 1983 and 2000 were estimated. Hyperimmune sera were produced in rabbits and antibody titres determined with each isolate. Isolates showing CRIs³ 70 were placed in the same group. Group I had 13 isolates (A, 1; B, 6; U, 6); group II had 6 isolates (A, 4; U, 2); groups III, IV, and V had 2 isolates each, recovered in Argentina; group VI had 2 isolates, from Uruguay; and group VII had 1 isolate, from Brazil. The CRIs³ 70 between vaccine strains and isolates recovered before and after 1990 were 58% and 42%, 50% and 50%, and 33% and 67% with vaccine strains 2419, 2358, and 2439, respectively. Isolate 273, from Uruguay, showed CRIs > 70 with 78% of the isolates and is recommended as the vaccine strain.     

Comparison of the Virulence of Various Strains of Moraxella bovis

The relative virulence of various strains of Moraxella bovis (M. bovis) was studied using the eyes of mice and cattle. The investigation consisted of three separate experiments. Experiments I and II involved a study on the effects of (1) different methods of growth and (2) serial blood agar passaging on the virulence of M. bovis. Experiment III involved a study on the relative virulence of different strains of M. bovis and M. bovis-like organisms.

Strains of M. bovis and M. bovis-like organisms varied in their pathogenicity for mice. However, different methods for preparation of exposure cultures of M. bovis did not influence the disease produced.

     

Virulence factors and genetic variability of uropathogenic Escherichia coli isolated from dogs and cats in Italy

In this study, the association between virulence genotypes and phylogenetic groups among Escherichia (E.) coli isolates obtained from pet dogs and cats with cystitis was detected, and fingerprinting methods were used to explore the relationship among strains. Forty uropathogenic E. coli (UPEC) isolated from dogs (n = 30) and cats (n = 10) in Italy were analysed by polymerase chain reaction (PCR) for the presence of virulence factors and their classification into phylogenetic groups. The same strains were characterized by repetitive extragenic palindromic (REP)- and enterobacterial repetitive intergenic consensus (ERIC)-PCR techniques. We found a high number of virulence factors such as fimbriae A, S fimbriae (sfa) and cytotoxic necrotizing factor 1 (cnf1) significantly associated with phylogenetic group B2. We demonstrated a high correlation between α-hemolysin A and pyelonephritis C, sfa, and cnf1 operons, confirming the presence of pathogenicity islands in these strains. In addition, UPEC belonging to group B2 harboured a greater number of virulence factors than strains from phylogenetic groups A, B1, and D. REP- and ERIC-PCR grouped the UPEC isolates into two major clusters, the former grouping E. coli strains belonging to phylogenetic group B2 and D, the latter grouping those belonging to groups A and B1. Given the significant genetic variability among the UPEC strains found in our study, it can be hypothesized that no specific genotype is responsible for cystitis in cats or dogs.     

Antimicrobial susceptibility and molecular characterization of macrolide resistance of Mycoplasma bovis isolates from multiple provinces in China

Mycoplasma bovis has spread widely throughout the world via animal movement and has become an important pathogen of bovine respiratory disease. However, the minimum inhibitory concentrations of antimicrobials for Mycoplasma bovis have not been studied in China. The objective of this study was to determine the prevalence and antibiotic resistance of Mycoplasma bovis isolated from young cattle with respiratory infection in China. Mycoplasma bovis was detected in 32/45 bovine respiratory infection outbreaks at beef farms in 8 provinces in China. The isolates were susceptible or had medium sensitivity to ciprofloxacin, enrofloxacin and doxycycline, but were frequently resistant to macrolides (13/32, 41%). An A2058G (Escherichia coli Numbering) mutation located in the rrnA operon in domain V of 23S rRNA was observed in strains that were resistant to macrolides. This single mutations at the rrnA operon in domain V of 23S rRNA may play an important role in the resistance of Mycoplasma bovis strains to macrolides.     

Serotype distribution and production of muramidase-released protein, extracellular factor and suilysin by field strains of Streptococcus suis isolated in the United States

Streptococcus suis is an important swine pathogen and a zoonotic agent. Differences in virulence have been noted among the 33 described serotypes, serotype 2 being considered the most virulent. In this study, we aimed at assessing the serotype distribution and the production of virulence-associated markers by strains recovered from diseased pigs in the United States (U.S.). Results showed that among the 100 strains evaluated, serotype 3 (20% of the isolates) and serotype 2 (17%) were the most prevalent. We then investigated the presence in these isolates of the genes sly, epf and mrp, encoding the virulence-associated markers suilysin (SLY), extracellular factor (EF) and muramidase-released (MRP) protein, respectively. The effective production of the markers by the strains was also verified. Results showed that the presence of the gene did not always correlate with actual expression of the respective protein. In the case of MRP, this was due, in most cases, to frameshift mutations at the 5′ end of the gene resulting in premature stop codons. The most prevalent phenotypes among U.S. strains were MRP⁺EF⁻SLY⁻ (40%) and MRP⁻EF⁻SLY⁺ (35%). Serotype distribution greatly differed from that reported in several European countries, as did the production of virulence markers, particularly for serotype 2. On the other hand, our results for the U.S. S. suis isolates are similar to those reported for Canadian strains, suggesting a common status in North America.     

Evaluation of the live vaccine efficacy of virulence plasmid-cured,and phoP- or aroA-deficient Salmonella enterica serovar Typhimurium in mice

We evaluated the protective efficacy of 94-kb virulence plasmid-cured, and phoP- or aroA-deficient strains of Salmonella enterica serovar Typhimurium (ΔphoP or ΔaroA S. Typhimurium) as oral vaccine candidates in BALB/c mice. Two weeks after the completion of 3 oral immunizations with 1 × 10⁸ colony-forming units (CFU) of virulence plasmid-cured, and ΔphoP or ΔaroA S. Typhimurium at 10-day intervals, S. Typhimurium lipopolysaccharide (LPS)-specific mucosal secretory immunoglobulin A (s-IgA) antibody titers were detected in the cecal homogenate, bile and lung lavage fluid, but not in the intestinal lavage fluid. In addition, the increases in S. Typhimurium LPS-specific immunoglobulin G (IgG) and IgA antibody titers in the serum were also observed 2 weeks after completing 3 oral immunizations with virulence plasmid-cured, and ΔphoP or ΔaroA S. Typhimurium. The series of 3 oral immunizations protected the mice against an oral challenge with 5 × 10⁸ CFU of the virulent strain of S. Typhimurium, suggesting that both the virulence plasmid-cured, and ΔphoP and ΔaroA S. Typhimurium strains are promising candidates for safe and effective live S. Typhimurium vaccines.     

S‐ketamine versus racemic ketamine in dogs: their relative potency as induction agents

ObjectiveTo determine the potency ratio between S-ketamine and racemic ketamine as inductive agents for achieving tracheal intubation in dogs.Study designProspective, randomized, ‘blinded’, clinical trial conducted in two consecutive phases.Animals112 client-owned dogs (ASA I or II).MethodsAll animals were premedicated with intramuscular acepromazine (0.02 mg kg⁻¹) and methadone (0.2 mg kg⁻¹). In phase 1, midazolam (0.2 mg kg⁻¹) with either 3 mg kg⁻¹ of racemic ketamine (group K) or 1.5 mg kg⁻¹ of S-ketamine (group S) was administered IV, for induction of anaesthesia and intubation. Up to two additional doses of racemic (1.5 mg kg⁻¹) or S-ketamine (0.75 mg kg⁻¹) were administered if required. In phase 2, midazolam (0.2 mg kg⁻¹) with 1 mg kg⁻¹ of either racemic ketamine (group K) or S-ketamine (group S) was injected and followed by a continuous infusion (1 mg kg minute⁻¹) of each respective drug. Differences between groups were statistically analyzed via t-test, Fisher exact test and ANOVA for repeated measures.ResultsDemographics and quality and duration of premedication, induction and intubation were comparable among groups. During phase 1 it was possible to achieve tracheal intubation after a single dose in more dogs in group K (n = 25) than in group S (n = 16) (p = 0.046). A dose of 3 mg kg⁻¹ S-ketamine allowed tracheal intubation in the same number of dogs as 4.5 mg kg⁻¹ of racemic ketamine. The estimated potency ratio was 1.5:1. During phase 2, the total dose (mean ± SD) of S-ketamine (4.02 ±1.56 mg kg⁻¹) and racemic ketamine (4.01 ± 1.42) required for tracheal intubation was similar.Conclusion and clinical relevanceRacemic and S-ketamine provide a similar quality of anaesthetic induction and intubation. S-ketamine is not twice as potent as racemic ketamine and, if infused, the potency ratio is 1:1.     

Vaccination of calves with Mycobacteria bovis Bacilli Calmete Guerin (BCG) induced rapid increase in the proportion of peripheral blood γδ T cells

Changes in the proportion of peripheral blood T cell subsets after subcutaneous inoculation of cattle with Mycobacterium bovis Bacille Calmette-Guerin (BCG) were studied. Calves were injected with approximately 8 × 10⁶ BCG bacillus and blood samples collected at weekly intervals for flow-cytometric analyses to determine the proportion of CD4⁺, CD8⁺ and γδ T cells. In addition, whole blood samples were stimulated in vitro with M. bovis purified protein derivative (PPD) and the secreted IFN-γ quantified by ELISA. Results showed cellular and cytokine changes which could be categorized into three phases. The first phase occurred within the first 2 weeks after vaccination involving an increase in proportion of WC1⁺ γδ T cells and a concomitant increase in the secretion of IFN-γ. These two responses peaked at 2 weeks and waned thereafter. The second phase involved an increase in the CD4/CD8 ratio as a result of an increase in the proportion of CD4⁺ T cells between 4 and 6 weeks. The third phase involved a decrease in the CD4/CD8 ratio due to an increase in the proportion of CD8⁺ T cells between 8 and 10 weeks. Surprisingly, the IFN-γ response was associated with changes in the γδ rather than the CD4⁺ or CD8⁺ T cells, suggesting that this cytokine was secreted by γδ-T cells. These results are consistent with the reported ability of γδ T cells to act rapidly and bridging the innate and classically adaptive immune responses.     

Diversity in bacterium-host interactions within the species Helicobacter heilmannii sensu stricto

Helicobacter (H.) heilmannii sensu stricto (s.s.) is a zoonotic bacterium that naturally colonizes the stomach of dogs and cats. In humans, this microorganism has been associated with gastritis, peptic ulcer disease and mucosa associated lymphoid tissue (MALT) lymphoma. Little information is available about the pathogenesis of H. heilmannii s.s. infections in humans and it is unknown whether differences in virulence exist within this species. Therefore, a Mongolian gerbil model was used to study bacterium-host interactions of 9 H. heilmannii s.s. strains. The colonization ability of the strains, the intensity of gastritis and gene expression of various inflammatory cytokines in the stomach were determined at 9 weeks after experimental infection. The induction of an antrum-dominant chronic active gastritis with formation of lymphocytic aggregates was shown for 7 strains. High-level antral colonization was seen for 4 strains, while colonization of 4 other strains was more restricted and one strain was not detected in the stomach at 9 weeks post infection. All strains inducing a chronic active gastritis caused an up-regulation of the pro-inflammatory cytokine IL-1β in the antrum. A reduced antral expression of H⁺/K⁺ ATPase was seen in the stomach after infection with 3 highly colonizing strains and 2 highly colonizing strains caused an increased gastrin expression in the fundus. In none of the H. heilmannii s.s.-infected groups, IFN-γ expression was up-regulated. This study demonstrates diversity in bacterium-host interactions within the species H. heilmannii s.s. and that the pathogenesis of gastric infections with this microorganism is not identical to that of an H. pylori infection.     

Development of a tandem repeat-based multilocus typing system distinguishing Babesia bovis geographic isolates

Mini- and microsatellite sequences have proven to be excellent tools for the differentiation of strains and populations in several protozoan parasites due to their high variability. In the present work we have searched the genome of the tick-transmitted bovine hemoprotozoon Babesia bovis for tandem repeats (TRs) that could be useful for a multilocus typing system. Hundred and nineteen sequences were shortlisted and tested in five common B. bovis reference isolates originating from distinct geographic locations of North and South America: Texas, USA (T2Bo), Mexico (RAD and Mo7), and Santa Fe and Salta, Argentina (R1A and S2P, respectively). Satellite sequences were PCR-amplified using specific primers, separated by polyacrylamide gel electrophoresis, visualized by silver staining and sized. Fourteen TR sequences could be reliably amplified in all isolates and displayed length polymorphism. All primers used were specific for B. bovis and did not amplify genomic DNA from the bovine host or from Babesia bigemina, the principal co-infecting bovine parasite in the Americas, allowing their future use in field surveys. The 14 satellite markers identified are distributed throughout the four chromosomes of B. bovis as follows: chromosome 1 (n = 3), chromosome 2 (n = 2), chromosome 3 (n = 5), and chromosome 4 (n = 4). Within the five B. bovis isolates we identified nine satellite marker loci with two alleles, three with three alleles, one with four and another with five alleles. In comparison to Theileria parva, a bovine hemoprotozoan that pertains to the same piroplasmida order and owns a genome of similar size, the number of polymorphic TRs and the average number of alleles per TR locus seem to be significantly reduced in the B. bovis genome. Furthermore, the ratio of micro- to minisatellites in both B. bovis and T. parva is considerably lower than in other eukaryotes, as confirmed by bioinformatic analysis. The multilocus genotype of the five B. bovis isolates was assessed and the genetic distance between each other determined followed by cluster analysis based on neighbor joining. The resulting phenogram showed that B. bovis isolates segregated into three clusters according to their geographic origin. The presented marker system is suitable to explore various parameters of B. bovis populations such as genetic diversity, infection dynamics and their structure under different epidemiological situations, which are of crucial importance for improved control strategies.     

Comparison of the effects of propofol or alfaxalone for anaesthesia induction and maintenance on respiration in cats

ObjectiveTo compare the effects of propofol and alfaxalone on respiration in cats.Study designRandomized, ‘blinded’, prospective clinical trial.AnimalsTwenty cats undergoing ovariohysterectomy.MethodsAfter premedication with medetomidine 0.01 mg kg⁻¹ intramuscularly and meloxicam 0.3 mg kg⁻¹ subcutaneously, the cats were assigned randomly into two groups: group A (n = 10) were administered alfaxalone 5 mg kg⁻¹ minute⁻¹ followed by 10 mg kg⁻¹ hour⁻¹ intravenously (IV) and group P (n = 10) were administered propofol 6 mg kg⁻¹ minute⁻¹ followed by 12 mg kg⁻¹hour⁻¹ IV for induction and maintenance of anaesthesia, respectively. After endotracheal intubation, the tube was connected to a non-rebreathing system delivering 100% oxygen. The anaesthetic maintenance drug rate was adjusted (± 0.5 mg kg⁻¹ hour⁻¹) every 5 minutes according to a scoring sheet based on physiologic variables and clinical signs. If apnoea > 30 seconds, end-tidal carbon dioxide (Pe′CO₂) > 7.3 kPa (55 mmHg) or arterial haemoglobin oxygen saturation (SpO₂) < 90% occurred, manual ventilation was provided. Methadone was administered postoperatively. Data were analyzed using independent-samples t-tests, Fisher's exact test, linear mixed-effects models and binomial test.ResultsManual ventilation was required in two and eight of the cats in group A and P, respectively (p = 0.02). Two cats in both groups showed apnoea. Pe′CO₂ > 7.3 kPa was recorded in zero versus four and SpO₂ < 90% in zero versus six cats in groups A and P respectively. Induction and maintenance dose rates (mean ± SD) were 11.6 ± 0.3 mg kg⁻¹ and 10.7 ± 0.8 mg kg⁻¹ hour⁻¹ for alfaxalone and 11.7 ± 2.7 mg kg⁻¹ and 12.4 ± 0.5 mg kg⁻¹ hour⁻¹ for propofol.Conclusion and clinical relevanceAlfaxalone had less adverse influence on respiration than propofol in cats premedicated with medetomidine. Alfaxalone might be better than propofol for induction and maintenance of anaesthesia when artificial ventilation cannot be provided.     

Anaesthetic sparing effect of local anaesthesia of the ovarian pedicle during ovariohysterectomy in dogs

ObjectiveTo evaluate the effect of local anaesthesia of the mesovarium on end-tidal isoflurane (Fe′_iso) concentration and vital parameters during canine ovariohysterectomy.Study designProspective, randomized, blinded study.AnimalsTwenty client-owned dogs undergoing elective ovariohysterectomy. Mean age 1.7 (±0.53, SD) years and mean body weight 21 kg (±5.9, SD).MethodsPre-medication was with intravenous acepromazine (0.02 mg kg⁻¹) and methadone (0.1 mg kg⁻¹). Anaesthesia was induced with propofol and maintained with isoflurane in oxygen. One group (n = 10) received local infiltration of the mesovarium with 0.5 mL lidocaine 2% and one group (n = 10) with 0.5 mL NaCl 0.9%. Heart (HR) and respiratory rates (fr), invasive mean arterial blood pressure (MAP) and Fe′_isowere recorded. The Fe′_iso was adjusted according to changes in HR, RR and MAP. Time points used for comparison were T1 (after induction of anaesthesia before surgery), T2 (after lidocaine infiltration of the mesovarium) and T3 (surgical manipulation of the ovaries). Data were analysed using a mixed model for repeated measurement anova and the Tukey adjustment. Results are presented as mean ± SD; p < 0.05 was considered significant.ResultsIn both groups, HR and fr remained stable at the three time points. Mean values ranged from 84 to 94 beats minute⁻¹ and from 10 to 14 breaths minute⁻¹. The Fe′_iso was significantly lower at T3 compared to T1 and mean values ranged from 0.95% to 1.24%. The mean arterial blood pressure was significantly higher at T3 compared to T1 and mean values ranged from 58 to 96 mm Hg. At none of the time points were there significant differences between the two groups for HR, fr, MAP or Fe′_iso.ConclusionNeither an isoflurane sparing effect nor a difference in autonomic response to surgery was demonstrated following local anaesthesia of the mesovarium.Clinical relevanceThere appeared to be minimal benefit from local anaesthesia of the mesovarium during this study.