Pathology, Tissue Distribution, and Phylogenetic Characterization of Largemouth Bass Virus Isolated from a Wild Smallmouth Bass (Micropterus dolomieu)

Abstract

We performed a diagnostic disease investigation on a wild smallmouth bass (Micropterus dolomieu) with skin ulcers that was collected from Lake Oahe, South Dakota, following reports from anglers of multiple fish with similar lesions. Gross and histologic lesions of ulcerative dermatitis, myositis, and lymphocytolysis within the spleen and kidneys were consistent with largemouth bass virus (LMBV) infection. LMBV was detected by conventional PCR in samples of a skin ulcer, and the complete genome sequence of the LMBV (99,184 bp) was determined from a virus isolate obtained from a homogenized skin sample. A maximum likelihood (ML) phylogenetic analysis based on the major capsid protein (MCP) gene alignment supported the LMBV isolate (LMBV-SD-2023) as a member of the species Ranavirus micropterus1, branching within the subclade of LMBV isolates recovered from North American largemouth (Micropterus salmoides) and smallmouth bass. This is the first detection of LMBV in wild smallmouth bass from South Dakota. The ultrastructure of the LMBV isolate exhibited the expected icosahedral shape of virions budding from cellular membranes. Viral nucleic acid in infected cells was visualized via in situ hybridization (ISH) within dermal granulomas, localized predominantly at the margin of epithelioid macrophages and central necrosis. Further sampling is needed to determine the geographic distribution, affected populations, and evolutionary relationship between isolates of LMBV.

Keywords: iridovirus; largemouth bass virus; ranavirus; smallmouth bass.

Figure 1

Gross lesions in the smallmouth bass (Micropterus dolomieu). (A) Cutaneous ulcers and erythema on the left flank and ventrum. (B) Liver with pale tan mottling, ovary, spleen, and coelomic adipose tissue. (C) Cutaneous ulcers on the left flank. (D) Large cutaneous ulcer on the left flank, measuring 7.0 cm × 4.0 cm, with central necrotic tissue and exposure of the underlying muscle.

Figure 2

Matching H&E histopathology and tissue distribution of largemouth bass virus (LMBV) in a section of skin ulcer by in situ hybridization (ISH). (A) Granuloma within the skin, with central necrotic debris (asterisk) rimmed by epithelioid macrophages (arrow), H&E. (B) Multifocal positive reactivity for LMBV localized predominantly at the margin of epithelioid macrophages and central necrosis (asterisk).

Figure 3

Maximum likelihood cladogram depicting the relationship of the largemouth bass virus isolate (SD-2023) (red) to 53 ranaviruses, based on the nucleotide sequence alignment of the major capsid protein gene. Numbers at each node represent bootstrap support of the ML analysis (values ≥ 80% shown). See Table S1 for viral taxa abbreviations.

Figure 4

Largemouth bass virus in fathead minnow (Pimephales promelas) cell line 48 h after infection. (A–C) Non-enveloped virions in the cell cytoplasm with an icosahedral capsid containing an electron-dense core. Notice viral budding in (A,C). (D) Non-enveloped virions budding from the cell membrane, with few virions enveloped by the cellular plasma membrane (arrows). (E,F) Viral assembly regions within the cell cytoplasm demarcated by an area of electron dense (E) or electron lucent (F) cytoplasm containing cross and longitudinal sections of tubular elements.