An atlas of gross and histologic lesions and immunohistochemical immunoreactivity during the temporal progression of aerosolized Lassa virus induced hemorrhagic fever in cynomolgus macaques

Abstract

Lassa virus (LASV) causes an acute multisystemic hemorrhagic fever in humans known as Lassa fever, which is endemic in several African countries. This manuscript focuses on the progression of disease in cynomolgus macaques challenged with aerosolized LASV and serially sampled for the development and progression of gross and histopathologic lesions. Gross lesions were first noted in tissues on day 6 and persisted throughout day 12. Viremia and histologic lesions were first noted on day 6 commencing with the pulmonary system and hemolymphatic system and progressing at later time points to include all systems. Immunoreactivity to LASV antigen was first observed in the lungs of one macaque on day 3 and appeared localized to macrophages with an increase at later time points to include immunoreactivity in all organ systems. Additionally, this manuscript will serve as a detailed atlas of histopathologic lesions and disease progression for comparison to other animal models of aerosolized Arenaviral disease.

Keywords: Lassa fever; Lassa virus; cynomolgus macaques; histologic lesion; immunohistochemistry; non-human primate.

Figure 1

Tracheobronchial LN, Monkey 9, Day 6 PE. (A) Note the inner cortex and medulla is obscured by fibrin, edema and mononuclear cells (*) with a thin rim of outer cortex composed of lymphoid follicles (➔) H&E, 20x. (B) Immunoreactive mononuclear cells throughout the inner cortex and medulla, LASV IHC, 20x. (C) Higher magnification demonstrating immunoreactive mononuclear cells (dendritic cells, macrophages) throughout the subcapsular sinus and parafollicular cortex, LASV IHC, 400x. Tracheobronchial LN, Monkey 11, Day 10 PE. (D) Necrosis, hemorrhage, fibrin, and edema obscure the inner cortex and medulla, H&E, 20x. (E) There are a few clusters of immunoreactive cells present, LASV IHC, 20x. (F) Higher magnification demonstrating paucity of immunoreactive cells, LASV IHC, 200x. Tracheobronchial LN, Monkey 16. Day 11 PE. (G) Immunoreactive cells in center of follicle and perinodal adipocytes. LASV IHC, 100x. Tracheobronchial LN, Monkey 18, Day 12 PE. (H) Focus of necrosis at periphery of node, H&E, 100x. (I) Immunoreactivity in adjacent focus of necrosis and absent in the rest of the tissue, LASV IHC, 100x.

Figure 2

Mediastinal LN, Monkey 8, Day 6 PE. (A) 20x, (B) 100x, (C) 400x. Immunoreactive mononuclear cells throughout the sinuses. LASV IHC. Mediastinal LN, Monkey 11, Day 10 PE. LASV IHC. (D) 20x, (E) 200x, (F) 400x. Note the marked reduction in immunoreactive cells by day 10 PE with only a few clusters of immunoreactive cells present.

Figure 3

Presence of Lassa viral genomes in serum detected by qRT-PCR.

Figure 4

Gross pathologic findings, (A) Axillary rash. Monkey 18 Day 12 PE. (B) Enlarged pale liver. Monkey 12 Day 10 PE. (C) Congested and cloudy meninges. Monkey 14 Day 11 PE. (D) Hyperemic lungs with edema at the level of the tracheobronchial lymph nodes (Arrow). Monkey 19 Day 12 PE; (E) Mucosal hemorrhages and edema of the ileocecal junction (Arrow). Monkey 19 Day 12 PE.

Figure 5

(A) Lung, Monkey 6, Day 3 PE. Immunoreactive mononuclear cells within a focus of inflammation. LASV IHC, 200x. (B) Lung, Monkey 8, Day 6 PE. Immunoreactive mononuclear cells and pneumocytes within a focus of slight interstitial pneumonia. LASV IHC, 200x. (C) Lung, Monkey 12, Day 10 PE. Immunoreactive mononuclear cells and pneumocytes within a focus of pleural inflammation and interstitial pneumonia. LASV IHC, 200x.

Figure 6

Lung, Monkey 16. Day 11 PE. (A) Alveolitis and interstitial pneumonia characterized by thickened alveolar septa, sub-acute inflammation, alveolar fibrin, and edema (*). H&E, 100X. (B) Numerous immunoreactive mononuclear cells presumed to be alveolar macrophages and occasional pneumocytes; LASV IHC, 100X. (C) Higher magnification, demonstrating alveolar macrophages, fibrin, and mildly thickened alveolar septa. H&E, 400X. (D) Higher magnification, demonstrating immunoreactive alveolar macrophages and pneumocytes (➔). LASV IHC, 400X.

Figure 7

Spleen, Monkey 17 Day 12 PE. (A) The red pulp is expanded by numerous histiocytes. H&E, 40X. (B) Numerous immunoreactive mononuclear cells presumed to be venous sinus endothelial cells and FRCs. LASV IHC, 40X. (C) Higher magnification demonstrating numerous plump mononuclear cells throughout the red pulp. H&E, 200X. (D) Higher magnification, demonstrating immunoreactive venous sinus endothelial cells and FRCs. LASV IHC, 400X.

Figure 8

Liver, Monkey 18. Day 12 PE. (A) Multiple foci of hepatocellular necrosis with mild lipid-type vacuolar degeneration. H&E, 400X. (B) Multifocal immunoreactive hepatocytes and mononuclear cells presumed to be Kupffer cells demonstrating predominantly membrane associated immunoreactivity. LASV IHC, 400X.

Figure 9

Brain, Monkey 15. Day 11 PE. (A) Mononuclear inflammation surrounding blood vessels and scattered locally throughout the section. H&E, 100X. (B) Numerous immunoreactive blood vessels and immunoreactive debris within the adjacent tissue. LASV IHC, 100X. (C) Higher magnification, demonstrating small focus of perivascular mononuclear (lymphocytes and macrophages) inflammation and adjacent neuronal satellitosis. H&E, 400X. (D) Higher magnification, demonstrating immunoreactivity associated with the vessel and surrounding inflammatory cells. LASV IHC, 400X.

Figure 10

Temporal detection of viral antigen.