Experimental Inoculation with Human Granulocytic Ehrlichia Agent Derived from High- and Low-Passage Cell Culture in Horses

Pusterla, Nicola; Madigan, John E.; Asanovich, K.; Chae, Joon; DeRock, E.; Leutenegger, Christian M.; Pusterla, Jeannine Berger; Lutz, Hans; Dumler, J. Stephen

doi:10.1128/jcm.38.3.1276-1278.2000

Cited by 22 publications

(10 citation statements)

References 21 publications

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“…This implies that A. phagocytophilum cells contain components that nonspecifically stimulate lymphocyte proliferation and cytokine production, and this activity is quantitatively greater in low passage bacteria and facilitates stronger total cellular immune responses. This observation supports prior differences observed in vivo among horses infected by low and high passage A. phagocytophilum (Pusterla et al, 2000), and supports an important role for innate immunity in the induction of immunopathologic injury with infection (Scorpio et al, 2006).…”

Section: Discussionsupporting

confidence: 87%

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Msp2 variation inAnaplasma phagocytophilum in vivodoes not stimulate T cell immune responses or interferon-Î³ production

Choi

Scorpio

Barat

et al. 2007

FEMS Immunology &Amp; Medical Microbiology

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Msp2 is Anaplasma phagocytophilum's immunodominant protein. Antigenic variability with msp2 gene conversion may drive differential immunopathology with infection by bacteria of different in vitro passage intervals. We examined msp2 transcript variation and its relationship to histopathology, T-cell and antibody responses in mice infected with differentially passaged A. phagocytophilum. Hepatic inflammation peaked on day 2-4 with low passage bacteria and on day 4-7 with high passage bacteria infection. Nineteen msp2 variant transcripts were identified. The low and high passage inocula shared four, but differed in one and two msp2 transcript variants, respectively. After infection, three and two msp2 variants were only identified in low or high passage infected mice. However, per mouse, msp2 variant profiles were unique with no evident expression program. In low and high passage bacteria-infected mice, splenocytes proliferated to whole A. phagocytophilum at day 7-10, diminishing thereafter. Weak mitogenic responses to whole bacteria were detected in mock and infected mice at d0 and sporadically thereafter. Essentially no lymphoproliferation or IFN-gamma production resulted from stimulation by six Msp2 hypervariable region proteins, although antibodies were detected to all, including cross-reactions. Differential A. phagocytophilum Msp2 expression is unrelated to T-cell response and unlikely to induce the cellular immunopathology underlying disease manifestations.

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Section: Discussionsupporting

confidence: 87%

“…Scorpio, manuscript in preparation). These findings occur despite the confirmation of dichotomous clinical manifestations with high and low passage A. phagocytophilum infection in horses (Pusterla et al, 2000), and with differential histopathologic responses in mice, as demonstrated here.…”

Section: Discussioncontrasting

confidence: 60%

Msp2 variation inAnaplasma phagocytophilum in vivodoes not stimulate T cell immune responses or interferon-Î³ production

Choi

Scorpio

Barat

et al. 2007

FEMS Immunology &Amp; Medical Microbiology

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“…3,18 In contrast, a number of studies have been performed where horses have been inoculated with the humanderived HGE agent, which results in clinical changes largely indistinguishable from EGE disease. 5,[19][20][21][22][23] EGE is commonly diagnosed in Sweden. This paper is part of a longer-term study addressing whether persistence of infection with A phagocytophilum can occur in horses.…”

mentioning

confidence: 99%

Acute Clinical, Hematologic, Serologic, and Polymerase Chain Reaction Findings in Horses Experimentally Infected with a European Strain of Anaplasma phagocytophilum

et al. 2005

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Six horses were experimentally infected by administration of horse blood containing a Swedish strain of Anaplasma phagocytophilum. The polymerase chain reaction (PCR) signal was consistently detected 2-3 days before appearance of clinical signs and persisted 4-9 days beyond abatement of clinical signs, whereas diagnostic inclusion bodies were 1st noted on average 2.6 +/- 1.5 (SD) days after onset of fever. Clinical signs and hematologic changes were largely indistinguishable from those previously reported for diseases caused by A phagocytophilum (formerly Ehrlichia equi--"Californian agent") and the human-derived human granulocytic ehrlichiosis agent. Horses 1st demonstrated antibody response 12-16 days after inoculation, 2 cases of which were still febrile, and serotiters rapidly peaked within 3-7 days of clinical illness. One horse died during the acute stage of disease, but initial clinical signs and hematologic changes were similar to those of other infected horses. This report shows that, despite minor genetic differences, a European equine-derived strain of A. phagocytophilum may be similar in pathogenicity to the Californian agent. The PCR used holds promise to widen the diagnostic window and would also be diagnostic during the initial days of clinical disease when inclusions in neutrophils in blood smears are not yet apparent.

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“…Real-time PCR has been used for many years to study several aspects of the epidemiology and pathophysiology of EGE. 38,50 For clinical purposes, the material of choice is whole blood. Real-time PCR has been shown to be a very sensitive and specific tool, helping with diagnosis especially during early and late stages, when the number of organisms is too small for diagnosis by microscopy (Fig 4).…”

Section: Miscellaneous Pathogensmentioning

confidence: 99%

Real‐Time Polymerase Chain Reaction: A Novel Molecular Diagnostic Tool for Equine Infectious Diseases

Pusterla¹,

Madigan²,

Leutenegger³

2006

Veterinary Internal Medicne

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The focus of rapid diagnosis of infectious disease of horses in the last decade has shifted from the conventional laboratory techniques of antigen detection, microscopy, and culture to molecular diagnosis of infectious agents. Equine practitioners must be able to interpret the use, limitations, and results of molecular diagnostic techniques, as they are increasingly integrated into routine microbiology laboratory protocols. Polymerase chain reaction (PCR) is the best-known and most successfully implemented diagnostic molecular technology to date. It can detect slow-growing, difficult-to-cultivate, or uncultivatable microorganisms and can be used in situations in which clinical microbiology diagnostic procedures are inadequate, time-consuming, difficult, expensive, or hazardous to laboratory staff. Inherent technical limitations of PCR are present, but they are reduced in laboratories that use standardized protocols, conduct rigid validation protocols, and adhere to appropriate quality-control procedures. Improvements in PCR, especially probe-based real-time PCR, have broadened its diagnostic capabilities in clinical infectious diseases to complement and even surpass traditional methods in some situations. Furthermore, real-time PCR is capable of quantitation, allowing discrimination of clinically relevant infections characterized by pathogen replication and high pathogen loads from chronic latent infections. Automation of all components of PCR is now possible, which will decrease the risk of generating false-positive results due to contamination. The novel real-time PCR strategy and clinical applications in equine infectious diseases will be the subject of this review.Key words: Horse; Pathogen detection; TaqMan polymerase chain reaction. Equine practitioners have always considered accurate and rapid identification of infectious pathogens a priority. The ready availability of a correct etiologic diagnosis, particularly in contagious infections, enables the veterinarian to make early decisions on the patient's care and management, address appropriate treatment, and allow timely notification and discussion of management issues regarding prevention of disease spread. The last 2 decades have seen a revolution in the understanding, management, diagnosis, control, and prevention of infectious diseases.1,2 This period has encompassed the discovery of emerging equine agents, antimicrobials, vaccines, as well as a wealth of improved diagnostic tests for equine practitioners. Despite these advances, infectious diseases remain a leading cause of equine morbidity and mortality, with resurgence of certain infections (eg, West Nile virus), an increasing population of elderly, more susceptible horses, and an increasing international equine commerce, expanding the geographic distribution of pathogens. 3,4The focus of rapid diagnosis of infectious diseases also has shifted during this time. The most obvious change has been the appearance and increasing importance of nucleic acid (NA) amplification-based techniques, primarily the...

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Experimental Inoculation with Human Granulocytic Ehrlichia Agent Derived from High- and Low-Passage Cell Culture in Horses

Cited by 22 publications

References 21 publications

Msp2 variation inAnaplasma phagocytophilum in vivodoes not stimulate T cell immune responses or interferon-Î³ production

Msp2 variation inAnaplasma phagocytophilum in vivodoes not stimulate T cell immune responses or interferon-Î³ production

Acute Clinical, Hematologic, Serologic, and Polymerase Chain Reaction Findings in Horses Experimentally Infected with a European Strain of Anaplasma phagocytophilum

Real‐Time Polymerase Chain Reaction: A Novel Molecular Diagnostic Tool for Equine Infectious Diseases

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