Clinical and immunological responses in sheep after inoculation with Himar1-transformed Anaplasma phagocytophilum and subsequent challenge with a virulent strain of the bacterium

Eskeland, Sveinung; Stuen, Snorre; Munderloh, Ulrike G.; Barbet, Anthony F.; Crosby, Liliana; Lybeck, Kari; Wilhelmsson, Peter; Lindgren, Per-Eric; Makvandi‐Nejad, Shokouh; Tollefsen, Stig; Granquist, Erik Georg

doi:10.1016/j.vetimm.2020.110165

Cited by 3 publications

(13 citation statements)

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Section: Resultsmentioning

confidence: 99%

“…For transposon mutagenesis using the Himar 1 system, we sought to propagate these variants in cell culture [ 2 ]. This resulted in the isolation of three mutant populations (referred to as CL1A2, CL2B5, and CL3D3) in the Ixodes scapularis ISE6 tick cell line [ 2 ]. PCR analysis to determine whether the isolated Himar1-mutants belonged to the major or minor population showed that mutants CL1A2 and CL2B5 represented only the minor genotype while mutant CL3D3 included both genotypes, suggesting a mixed population ( Figure 3 ).…”

Section: Resultsmentioning

confidence: 99%

“…Recently, it has also been identified as the causative agent of an emerging disease in humans, known as human granulocytic anaplasmosis (HGA), with > 5000 cases identified in the U.S. by the CDC in 2019 ( accessed on 24 March 2022). In Norway, an estimated ~300,000 sheep are infected annually, with severe economic and animal welfare consequences [ 1 , 2 ]. This has stimulated the search for a vaccine against the disease in sheep.…”

Section: Introductionmentioning

confidence: 99%

“…Culture of sheep A. phagocytophilum strains has proven more difficult because they do not grow in HL-60 cells. Nevertheless, ruminant strains of A. phagocytophilum can be isolated using tick cell lines [ 3 , 4 , 5 ], and we recently obtained cultures of two strains from experimentally-infected Norwegian sheep, known as ApSheep_NorV1 and ApSheep_NorV2, in the tick cell line ISE6 [ 2 ]. Of these two strains, ApSheep_NorV1 is of particular interest for vaccine development because of its virulence [ 6 ].…”

Section: Introductionmentioning

confidence: 99%

“…Of these two strains, ApSheep_NorV1 is of particular interest for vaccine development because of its virulence [ 6 ]. However, it was recently shown that ApSheep_NorV1, derived from ISE6 cells, was not as immunoprotective in lambs as the original field strain derived from infected sheep blood [ 2 ]. Here, we identify a possible reason for this discrepancy, and provide a genomic analysis of the cultured ApSheep_NorV1 strain in comparison with other A. phagocytophilum strains of diverse animal and geographic origins.…”

Section: Introductionmentioning

confidence: 99%

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Comparative Whole Genome Analysis of an Anaplasma phagocytophilum Strain Isolated from Norwegian Sheep

et al. 2022

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Anaplasma phagocytophilum is a Gram-negative obligate intracellular tick-borne alphaproteobacteria (family Anaplasmatacea, order Rickettsiales) with a worldwide distribution. In Norway, tick borne fever (TBF), caused by A. phagocytophilum, presents a major challenge in sheep farming. Despite the abundance of its tick vector, Ixodes ricinus, and A. phagocytophilum infections in wild and domestic animals, reports of infections in humans are low compared with cases in the U.S. Although A. phagocytophilum is genetically diverse and complex infections (co-infection and superinfection) in ruminants and other animals are common, the underlying genetic basis of intra-species interactions and host-specificity remains unexplored. Here, we performed whole genome comparative analysis of a newly cultured Norwegian A. phagocytophilum isolate from sheep (ApSheep_NorV1) with 27 other A. phagocytophilum genome sequences derived from human and animal infections worldwide. Although the compared strains are syntenic, there is remarkable genetic diversity between different genomic loci including the pfam01617 superfamily that encodes the major, neutralization-sensitive, surface antigen Msp2/p44. Blast comparisons between the msp2/p44 pseudogene repertoires from all the strains showed high divergence between U. S. and European strains and even between two Norwegian strains. Based on these comparisons, we concluded that in ruminants, complex infections can be attributed to infection with strains that differ in their msp2/p44 repertoires, which has important implications for pathogen evolution and vaccine development. We also present evidence for integration of rickettsial DNA into the genome of ISE6 tick cells.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Comparative Whole Genome Analysis of an Anaplasma phagocytophilum Strain Isolated from Norwegian Sheep

et al. 2022

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Temporal patterns of gene expression in response to inoculation with a virulent Anaplasma phagocytophilum strain in sheep

Eskeland,

Bø-Granquist,

Stuen

et al. 2023

Sci Rep

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The aim of this study was to characterize the gene expression of host immune- and cellular responses to a Norwegian virulent strain of Anaplasma phagocytophilum, the cause of tick-borne fever in sheep. Ten sheep were intravenously inoculated with a live virulent strain of A. phagocytophilum. Clinical-, observational-, hematological data as well as bacterial load, flow cytometric cell count data from peripheral blood mononuclear cells and host’s gene expression post infection was analysed. The transcriptomic data were assessed for pre-set time points over the course of 22 days following the inoculation. Briefly, all inoculated sheep responded with clinical signs of infection 3 days post inoculation and onwards with maximum bacterial load observed on day 6, consistent with tick-borne fever. On days, 3–8, the innate immune responses and effector processes such as IFN1 signaling pathways and cytokine mediated signaling pathways were observed. Several pathways associated with the adaptive immune responses, namely T-cell activation, humoral immune responses, B-cell activation, and T- and B-cell differentiation dominated on the days of 8, 10 and 14. Flow-cytometric analysis of the PBMCs showed a reduction in CD4+CD25+ cells on day 10 and 14 post-inoculation and a skewed CD4:CD8 ratio indicating a reduced activation and proliferation of CD4-T-cells. The genes of important co-stimulatory molecules such as CD28 and CD40LG, important in T- and B-cell activation and proliferation, did not significantly change or experienced downregulation throughout the study. The absence of upregulation of several co-stimulatory molecules might be one possible explanation for the low activation and proliferation of CD4-T-cells during A. phagocytophilum infection, indicating a suboptimal CD4-T-cell response. The upregulation of T-BET, EOMES and IFN-γ on days 8–14 post inoculation, indicates a favoured CD4 Th1- and CD8-response. The dynamics and interaction between CD4+CD25+ and co-stimulatory molecules such as CD28, CD80, CD40 and CD40LG during infection with A. phagocytophilum in sheep needs further investigation in the future.

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Recent advances in genetic systems in obligate intracellular human-pathogenic bacteria

Fisher

Beare²

2023

Front. Cell. Infect. Microbiol.

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The ability to genetically manipulate a pathogen is fundamental to discovering factors governing host–pathogen interactions at the molecular level and is critical for devising treatment and prevention strategies. While the genetic “toolbox” for many important bacterial pathogens is extensive, approaches for modifying obligate intracellular bacterial pathogens were classically limited due in part to the uniqueness of their obligatory lifestyles. Many researchers have confronted these challenges over the past two and a half decades leading to the development of multiple approaches to construct plasmid-bearing recombinant strains and chromosomal gene inactivation and deletion mutants, along with gene-silencing methods enabling the study of essential genes. This review will highlight seminal genetic achievements and recent developments (past 5 years) for Anaplasma spp., Rickettsia spp., Chlamydia spp., and Coxiella burnetii including progress being made for the still intractable Orientia tsutsugamushi. Alongside commentary of the strengths and weaknesses of the various approaches, future research directions will be discussed to include methods for C. burnetii that should have utility in the other obligate intracellular bacteria. Collectively, the future appears bright for unraveling the molecular pathogenic mechanisms of these significant pathogens.

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Clinical and immunological responses in sheep after inoculation with Himar1-transformed Anaplasma phagocytophilum and subsequent challenge with a virulent strain of the bacterium

Cited by 3 publications

References 55 publications

Comparative Whole Genome Analysis of an Anaplasma phagocytophilum Strain Isolated from Norwegian Sheep

Comparative Whole Genome Analysis of an Anaplasma phagocytophilum Strain Isolated from Norwegian Sheep

Temporal patterns of gene expression in response to inoculation with a virulent Anaplasma phagocytophilum strain in sheep

Recent advances in genetic systems in obligate intracellular human-pathogenic bacteria

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