The role of phagocytic cells in enhanced susceptibility of broilers to colibacillosis after Infectious Bronchitis Virus infection

Colibacillosis results from infection with avian pathogenic Escherichia coli bacteria. Healthy broilers are resistant to inhaled E. coli, but previous infection with vaccine or virulent strains of Infectious Bronchitis Virus (IBV) predisposes birds for severe colibacillosis. We investigated whether IBV affects recruitment and function of phagocytic cells and examined NO production, phagocytic and bactericidal activity, and kinetics of peripheral blood mononuclear cells (PBMC) and splenocytes. Moreover, we measured cytokine mRNA expression in lung and spleen samples. Broilers were inoculated with IBV H120 vaccine or virulent M41 and challenged 5 days later with E. coli 506. A PBS control and E. coli group without previous virus inoculation were also included. Birds were sacrificed at various time points after inoculation (h/dpi). Inoculation with IBV induced extended and more severe colibacillosis than with E. coli alone. At 4dpi, the number of KUL-01(+) PBMC in all E. coli-inoculated groups was significantly higher than in PBS-inoculated birds, which correlated with lesion scores. From 1 to 4dpi, NO production by PBMC from all E. coli-inoculated animals was elevated compared to PBS birds. Bactericidal activity of PBMC in IBV-inoculated animals at 7dpi was lower than in PBS- and E. coli-inoculated birds, but phagocytic capacity and recruitment were not severely impaired. In spleen samples of IBV-infected animals reduced expression of IL-1beta, IL-6, IL-8, IL-10, IL-18 and IFN-gamma mRNA was found 1dpi. Our results suggest that enhanced colibacillosis after IBV infection or vaccination is caused at least by altered innate immunity and less by impairment of phagocytic cell function.     

Development of a multiplex PCR test for identification of Actinobacillus pleuropneumoniae serovars 1, 7, and 12

A PCR assay for simultaneous species identification and separation of Actinobacillus pleuropneumoniae serovars 1, 7 and 12 was developed. Primers specific for genes involved in biosynthesis of the capsular polysaccharides (cps genes) of serovars 1, 7, and 12 were combined with a species-specific PCR test based on the omlA gene. The PCR test was evaluated with the serovar reference strains of A. pleuropneumoniae as well as 183 Danish field isolates. For all typable strains, a complete correspondence was found between results obtained with the multiplex PCR test and results from the traditional serotyping methods. Among eight serologically cross-reacting strains designated K1:O7, seven isolates produced amplicons of similar sizes as serovar 1 and one isolate produced amplicons of similar sizes as serovar 7. The species specificity of the assay was evaluated using a collection of 126 strains representing 25 different species within the family Pasteurellaceae including 45 field strains of the phylogenetically affiliated species Actinobacillus lignieresii. All these isolates tested negative for the cps genes by the multiplex PCR test except for 6 isolates of A. lignieresii. Five of these isolates produced an amplicon identical to the cps gene of serovar 7, whereas one isolate produced an amplicon identical to the cps gene of serovar 1. In addition, four isolates of Actinobacillus genomospecies 1 tested positive for the omlA gene but negative for the cps genes. The test represents a convenient and specific method for serotyping A. pleuropneumoniae in diagnostic laboratories.     

Electron microscopic identification of viruses associated with poult enteritis in turkeys grown in California 1993-2003

Poult enteritis (PE) is one of the most common diseases seen in young turkey flocks. Since 1993, more than 1800 cases of suspected PE have been submitted for examination by negative stain electron microscopy; this has involved more than 2400 individual results, because in many cases more than one virus was identified; at least 1500 individual results were positive for viruses. Viruses have been identified in poults as young as 3 days and up to 9 wk of age. The most commonly found viruses are rotavirus-like viruses and small round viruses ranging from 15 nm to 30 nm, either alone or in combination. Reovirus, birnavirus, and adenovirus have also been detected. There has been no evidence to suggest the presence of coronaviruses. This report summarizes our findings.     

TB diagnosis in non-human primates: comparison of two interferon-gamma assays and the skin test for identification of Mycobacterium tuberculosis infection

To examine the effect of recombinant bovine interferon-gamma (rbIFN-gamma) on cattle persistently infected with bovine leukemia virus (BLV), BLV-infected cattle were inoculated intraperitoneally with IFN-gamma. All cattle were febrile after inoculation with IFN-gamma and then recovered within 48 h. Flow cytometric analysis showed that the numbers of CD4+ and CD8+ T cells were decreased for 2-3 days and then their numbers were recovered. The number of gammadelta T cells increased after the fever. In contrast, the number of IgM+ lymphocytes remained low for about 1 week. Moreover, the numbers of syncytia produced by peripheral blood lymphocytes decreased and remained low compared to that before IFN-gamma administration. These results suggest that IFN-gamma induces the up-regulation of gammadelta T cells, decreases the number of IgM+ lymphocytes and suppresses the growth of BLV in BLV-infected cattle in vivo.     

Analysis of world strains of Anaplasma marginale using major surface protein 1a repeat sequences

Anaplasma marginale is a tick-borne pathogen of cattle that causes the disease bovine anaplasmosis worldwide. Major surface proteins (MSPs) are involved in host-pathogen and tick-pathogen interactions and have been used as markers for the genetic characterization of A. marginale strains and phylogenetic studies. MSP1a is involved in the adhesion and transmission of A. marginale by ticks and varies among geographic strains in the number and sequence of amino-terminal tandem repeats. The aim of this study was to characterize the genetic diversity of A. marginale strains collected from countries in North and South America, Europe, Asia, Africa and Australia, inclusive of all continents. In this study, we characterized 131 strains of A. marginale using 79 MSP1a repeat sequences. These results corroborated the genetic heterogeneity of A. marginale strains in endemic regions worldwide. The phylogenetic analyses of MSP1a repeat sequences did not result in clusters according to the geographic origin of A. marginale strains but provided phylogeographic information. Seventy-eight percent of the MSP1a repeat sequences were present in strains from a single geographic region. Strong (> or =80%) support was found for clusters containing sequences from Italian, Spanish, Chinese, Argentinean and South American strains. The phylogenetic analyses of MSP1a repeat sequences suggested tick-pathogen co-evolution and provided evidence of multiple introductions of A. marginale strains from various geographic locations worldwide. These results contribute to the understanding of the genetic diversity and evolution of A. marginale and tick-pathogen interactions.     

A ten-year review of commercial vaccine performance for control of tick infestations on cattle

Ticks are important ectoparasites of domestic and wild animals, and tick infestations economically impact cattle production worldwide. Control of cattle tick infestations has been primarily by application of acaricides which has resulted in selection of resistant ticks and environmental pollution. Herein we discuss data from tick vaccine application in Australia, Cuba, Mexico and other Latin American countries. Commercial tick vaccines for cattle based on the Boophilus microplus Bm86 gut antigen have proven to be a feasible tick control method that offers a cost-effective, environmentally friendly alternative to the use of acaricides. Commercial tick vaccines reduced tick infestations on cattle and the intensity of acaricide usage, as well as increasing animal production and reducing transmission of some tick-borne pathogens. Although commercialization of tick vaccines has been difficult owing to previous constraints of antigen discovery, the expense of testing vaccines in cattle, and company restructuring, the success of these vaccines over the past decade has clearly demonstrated their potential as an improved method of tick control for cattle. Development of improved vaccines in the future will be greatly enhanced by new and efficient molecular technologies for antigen discovery and the urgent need for a tick control method to reduce or replace the use of acaricides, especially in regions where extensive tick resistance has occurred.     

Molecular and serologic characterization, pathogenicity, and protection studies with infectious bronchitis virus field isolates from California

In this study, we characterized three variant infectious bronchitis virus (IBV) strains isolated in 2003 and 2004 from broiler chickens in California and compared them to previously isolated California variant viruses and to common vaccine serotypes used in the United States. We conducted genetic, serologic, and pathogenicity studies on all three isolates, then tested different vaccines against one of the viruses. Genetically the three variant IBV strains, designated CA557/03, CA706/03, and CA1737/04, were not related to each other. GenBank BLAST database search and phylogenetic analysis of the hypervariable region of the S1 subunit of the spike gene to determine the most closely related viruses to the three variants showed the CA557/03 variant to be 81.8% similar to the CAV/CA56b/91 whereas the CA706/03 and CA1737/04 variant viruses were only distantly related to Dutch/D1466/81 (72.2%), a vaccine strain used in Europe, and Korea/K142/02 (72.7%), a Korean field isolate, respectively. Cross virus-neutralization testing showed that none of the 2003-04 California IBV variant viruses were serologically related to each other or to Ark, Conn, or Mass vaccine strains. In addition the CA1737/04 isolate was also tested against DE072 and found not to be serologically related. All three variant viruses were pathogenic in 1-wk-old broilers and vaccination with Mass/Conn followed by Holland/Conn provided 80% protection against the CA1737/04 virus. The 2003-04 California variant viruses were not compared with variants isolated in California during 1970s and 1980s because, to our knowledge, no genetic information is available and those viruses are no longer obtainable. This study shows that the CA557/03 virus was distantly related to the CAV-type viruses isolated in California in the early 1990s, but that none of the 2003-04 viruses were similar genetically or serologically to the CAL99-type viruses, indicating that new IBV variants continue to emerge and cause disease in commercial chickens in California.     

Comparing the detection of exposure to Ehrlichia ruminantium infection on a heartwater-endemic farm by the pCS20 polymerase chain reaction assay and an indirect MAP1-B enzyme linked immunosorbent assay

Detection of heartwater is not always easy especially because all the serological assays so far available either have poor sensitivity or specificity. The indirect MAP-1B ELISA has been reported to be the most specific test for heartwater, although it does also detect antibodies to some closely related ehrlichial agents. This study was undertaken to compare two methods for the detection of heartwater infection caused by the ehrlichial agent Ehrlichia (Cowdria) ruminantium. Fifteen cattle on a heartwater-endemic farm infested with high numbers of Amblyomma hebraeum ticks, and hence exposure to E. ruminantium infection were monitored over an 8-week period by pCS20 PCR and an indirect MAP-1B ELISA. Infection was detected by pCS20 PCR in most animals with the highest number of positives (60%) in week 6 of the study. Similarly, exposure to E. ruminantium was detected by indirect MAP-1B ELISA in some animals, with the highest number of seropositives (27%) at weeks 2-6 of the study. The data demonstrated a fluctuating rickettsaemia in cattle in a heartwater-endemic area. Comparison of the two tests indicated that the pCS20 PCR assay was more reliable because it detected more infections than the indirect MAP-1B ELISA and would therefore be the method of choice for detection of E. ruminantium infection.     

Discovery of a new avian bornavirus genotype in estrildid finches (Estrildidae) in Germany

Avian bornaviruses (ABV) are known to be the causative agent of proventricular dilatation disease (PDD) in parrots and their relatives (Psittaciformes). A broad range of ABV genotypes has been detected not only in psittacine birds, but also in other avian species including canary birds (Serinus canaria forma domestica) and Bengalese finches (Lonchura striata f. dom.), which are both members of the order songbirds (Passeriformes).During this study 286 samples collected from captive and wild birds of various passerine species in different parts of Germany were screened for the presence of ABV. Interestingly, only three ABV-positive samples were identified by RT-PCR. They originated from one yellow-winged pytilia (Pytilia hypogrammica) and two black-rumped waxbills (Estrilda troglodytes) from a flock of captive estrildid finches in Saxony. The ABV isolates detected here were only distantly related to ABV isolates found in passerine species in Germany and Japan and form a new genotype tentatively called ABV-EF (for “estrildid finches”).     

Validating an empiric sulfadiazine–trimethoprim dosage regimen for treatment of Escherichia coli and Staphylococcus delphini infections in mink (Neovison vison)

Antimicrobial agents are used extensively off‐label in mink, as almost no agents are registered for this animal species. Pharmacokinetic (PK) and pharmacodynamic (PD) data are required to determine antimicrobial dosages specifically targeting mink bacterial pathogens. The aims of this study were to assess, in a PKPD framework, the empirical dosage regimen for a combination of trimethoprim (TMP) and sulfadiazine (SDZ) in mink, and secondarily to produce data for future setting of clinical breakpoints. TMP and SDZ PK parameters were obtained experimentally in 22 minks following IV or oral administration of TMP/SDZ (30 mg/kg, i.e. 5 mg/kg TMP and 25 mg/kg SDZ). fAUC/MIC with a target value of 24 hr was selected as the PKPD index predictive of TMP/SDZ efficacy. Using a modeling approach, PKPD cutoffs for TMP and SDZ were determined as 0.062 and 16 mg/L, respectively. By incorporating an anticipated potentiation effect of SDZ on TMP against Escherichia coli and Staphylococcus delphini, the PKPD cutoff of TMP was revised to 0.312 mg/L, which is above the tentative epidemiological cutoffs (TECOFF) for these species. The current empirical TMP/SDZ dosage regimen (30 mg/kg, PO, once daily) therefore appears adequate for treatment of wild‐type E. coli and S. delphini infections in mink.