A competitive enzyme-linked immunosorbent assay using recombinant major surface protein 5 (rMSP5-cELISA) of Anaplasma marginale was validated in a naturally infected cattle herd in an area of eastern Oregon where A. marginale is endemic. The true positive and negative A. marginale infection status of 235 randomly selected cattle was determined by using a nested PCR (nPCR) coupled with msp5 sequence analysis and hybridization. Judgment of the reliability of the nPCR and hybridization for detection of persistent infections was based on three observations. First, the nPCR was able to detect as few as 30 infected erythrocytes per ml. Second, the nPCR was able to consistently detect low levels of rickettsemia in seven carrier cattle experimentally infected with A. marginale. Third, msp5 sequence analysis showed >95% identity among 30 nPCR amplicons from cattle naturally infected with field strains of A. marginale. The nPCR and hybridization identified 151 infected and 84 uninfected cattle among the 235 animals tested. With a cutoff point of 28%, the rMSP5-cELISA showed a sensitivity of 96% and a specificity of 95%. These results indicate that the rMSP5-cELISA can sensitively and specifically detect cattle with naturally acquired persistent A. marginale infections and suggest that it is an excellent assay for epidemiological studies, eradication programs, and regulation of international cattle movement.

Detection of Cattle Naturally Infected with Anaplasma marginale in a Region of Endemicity by Nested PCR and a Competitive Enzyme-Linked Immunosorbent Assay Using Recombinant Major Surface Protein 5

Fifty years ago, bacteria in the genus Brucella were known to cause infertility and reproductive losses. At that time, the genus was considered to contain only 3 species: Brucella abortus, Brucella melitensis, and Brucella suis. Since the early 1960s, at least 7 new species have been identified as belonging to the Brucella genus (Brucella canis, Brucella ceti, Brucella inopinata, Brucella microti, Brucella neotomae, Brucella ovis, and Brucella pinnipedialis) with several additional new species under consideration for inclusion. Although molecular studies have found such high homology that some authors have proposed that all Brucella are actually 1 species, the epidemiologic and diagnostic benefits for separating the genus based on phenotypic characteristics are more compelling. Although pathogenic Brucella spp have preferred reservoir hosts, their ability to infect numerous mammalian hosts has been increasingly documented. The maintenance of infection in new reservoir hosts, such as wildlife, has become an issue for both public health and animal health regulatory personnel. Since the 1960s, new information on how Brucella enters host cells and modifies their intracellular environment has been gained. Although the pathogenesis and histologic lesions of B. abortus, B. melitensis, and B. suis in their preferred hosts have not changed, additional knowledge on the pathology of these brucellae in new hosts, or of new species of Brucella in their preferred hosts, has been obtained. To this day, brucellosis remains a significant human zoonosis that is emerging or reemerging in many parts of the world.

https://journals.sagepub.com/doi/pdf/10.1177/0300985814540545

Advancement of knowledge of Brucella over the past 50 years.

The performance and characteristics of Roche COBAS AMPLICOR HIV-1 MONITOR version 1.5 (CA MONITOR 1.5) UltraSensitive (usCA MONITOR 1.5) and Standard (stCA MONITOR 1.5) procedures, Organon Teknika NucliSens HIV-1 RNA QT with Extractor (NucliSens), and Bayer Quantiplex HIV RNA version 3.0 (bDNA 3.0) were compared in a multicenter trial. Samples used in this study included 460 plasma specimens from human immunodeficiency virus (HIV) type 1 (HIV-1)-infected persons, 100 plasma specimens from HIV antibody (anti-HIV)-negative persons, and culture supernatants of HIV-1 subtype A to E isolates diluted in anti-HIV-negative plasma. Overall, bDNA 3.0 showed the least variation in RNA measures upon repeat testing. For the Roche assays, usCA MONITOR 1.5 displayed less variation in RNA measures than stCA MONITOR 1.5. NucliSens, at an input volume of 2 ml, showed the best sensitivity. Deming regression analysis indicated that the results of all three assays were significantly correlated (P < 0.0001). However, the mean difference in values between CA MONITOR 1.5 and bDNA 3.0 (0.274 log10 RNA copies/ml; 95% confidence interval, 0.192 to 0.356) was significantly different from 0, indicating that CA MONITOR 1.5 values were regularly higher than bDNA 3.0 values. Upon testing of 100 anti-HIV-negative plasma specimens, usCA MONITOR 1.5 and NucliSens displayed 100% specificity, while bDNA 3.0 showed 98% specificity. NucliSens quantified 2 of 10 non-subtype B viral isolates at 1 log10 lower than both CA MONITOR 1.5 and bDNA 3.0. For NucliSens, testing of specimens with greater than 1,000 RNA copies/ml at input volumes of 0.1, 0.2, and 2.0 ml did not affect the quality of results. Additional factors differing between assays included specimen throughput and volume requirements, limit of detection, ease of execution, instrument work space, and costs of disposal. These characteristics, along with assay performance, should be considered when one is selecting a viral load assay.

Multicenter Comparison of Roche COBAS AMPLICOR MONITOR Version 1.5, Organon Teknika NucliSens QT with Extractor, and Bayer Quantiplex Version 3.0 for Quantification of Human Immunodeficiency Virus Type 1 RNA in Plasma

Sheep infected with Brucella ovis produce antibody responses to the rough lipopolysaccharide and to proteins present in hot saline (HS) extracts of B. ovis (J. I. Riezu-Boj, I. Moriyon, J. M. Blasco, C. M. Marin, and R. Diaz, J. Clin. Microbiol. 23:938-942, 1986). The distribution and antigenic relatedness of proteins in HS extracts and in outer membrane blebs were established by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting for 41 strains of B. ovis and 26 strains of Brucella melitensis of diverse geographic origin. Five major groups of proteins were identified in HS extracts of B. ovis that had been freed of rough lipopolysaccharide: proteins of 43 kilodaltons (kDa), group A (25.5 to 32.0 kDa), group B (21.5 to 22.5 kDa); group C (18.0 to 19.5 kDa), and group D (13.0 to 15.5 kDa). Group A, B, C, and D proteins were also present in blebs. The profiles of proteins in HS extracts or blebs from strains of both Brucella species were very similar. Cross-reactions were demonstrated among HS extracts and blebs of all strains tested in immunoblots performed with an antiserum against the HS extract of a reference strain of B. ovis. Evidence was also provided of an antigenic relationship between group 3 proteins of the outer membrane and some of the proteins in groups A, B, and C. The conservation of these antigens and their immunogenicity in infected animals provide promise that they may serve as components of an effective subcellular vaccine for ovine brucellosis.

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Comparative analyses of proteins extracted by hot saline or released spontaneously into outer membrane blebs from field strains of Brucella ovis and Brucella melitensis.

In a lo-year survey started in 1980, specimens from 8,995 bovine abortions and stillbirths were submitted to the South Dakota Animal Disease Research and Diagnostic Laboratory. Of these, 8,962 were suitable for some type of examination. Viruses were associated with 948 (10.58%). Bovine herpesvirus-1 (IBR) was detected in 485 (5.41%), and bovine viral diarrhea virus (BVDV) was detected in 407 (4.54%). In 1 year of the survey, BVDV was detected in 8/32 fetuses that had lesions of passive congestion. Bovine herpesvirus- was isolated from 47 specimens (0.52%), parvovirus and enterovirus were each isolated from 2, and adenovirus, Parainfluenza virus, and pseudorabies virus were each isolated from 1. Malignant lymphoid neoplasia was present in 2 fetuses, and their abortion was assumed to have been caused by the bovine leukosis virus. The materials and methods and a summary of the the cows were susceptible to the virus. In some in- combined results of the diagnoses from all other causes stances, IBR abortions occurred when calves vacci- of bovine abortion and stillbirth during this lo-year nated with the modified live vaccine were turned back survey have been published recently. 13 Of the speci- in with their pregnant dams. However, vaccinated mens received from 8,995 bovine abortions and still- calves were never proven to be the source of infection births, 33 were unsuitable for any type of examination. in these cases. 19 In this article, the viral agents identified in the re- IBR abortion outbreaks occurred frequently in herds maining 8,962 aborted and stillborn bovine fetuses are isolated from other cattle and with no history of the discussed along with the associated lesions and syn- introduction of new animals or the use of live vaccines. dromes. The IBR virus becomes latent in naturally infected Results

https://journals.sagepub.com/doi/pdf/10.1177/104063879200400402

Brinton L. Swift

Papers

Vertical transmission of Anaplasma marginale in cattle

Abortion in cows inoculated with Anaplasma marginale

Inoculation of pregnant beef heifers at midgestation with Anaplasma marginale

Bovine fetal anoxia observed in pregnant beef heifers experimentally inoculated with Anaplasma marginale

Ram epididymitis: A clinical report.