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Persistent Foot-and-Mouth Disease Virus Infection in the Nasopharynx of Cattle; Tissue-Specific Distribution and Local Cytokine Expression.

Pacheco JM, Smoliga GR, O'Donnell V, Brito BP, Stenfeldt C, Rodriguez LL, Arzt J - PLoS ONE (2015)

Bottom Line: Immunomicroscopy indicated that within persistently infected mucosal tissues, FMDV antigens were rarely detectable within few epithelial cells in regions of mucosa-associated lymphoid tissue (MALT).Transcriptome analysis of persistently infected pharyngeal tissues by qRT-PCR for 14 cytokine genes indicated a general trend of decreased mRNA levels compared to uninfected control animals.Overall, this data demonstrates that during the FMDV carrier state in cattle, viral persistence is associated with epithelial cells of the nasopharynx in the upper respiratory tract and decreased levels of mRNA for several immunoregulatory cytokines in the infected tissues.

View Article: PubMed Central - PubMed

Affiliation: Plum Island Animal Disease Center, Foreign Animal Disease Research Unit, Agricultural Research Service, United States Department of Agriculture, Plum Island, NY, United States of America.

ABSTRACT
Tissues obtained post-mortem from cattle persistently infected with foot-and-mouth disease virus (FMDV) were analyzed to characterize the tissue-specific localization of FMDV and partial transcriptome profiles for selected immunoregulatory cytokines. Analysis of 28 distinct anatomic sites from 21 steers infected with FMDV serotype A, O or SAT2, had the highest prevalence of overall viral detection in the dorsal nasopharynx (80.95%) and dorsal soft palate (71.43%). FMDV was less frequently detected in laryngeal mucosal tissues, oropharyngeal mucosal sites, and lymph nodes draining the pharynx. Immunomicroscopy indicated that within persistently infected mucosal tissues, FMDV antigens were rarely detectable within few epithelial cells in regions of mucosa-associated lymphoid tissue (MALT). Transcriptome analysis of persistently infected pharyngeal tissues by qRT-PCR for 14 cytokine genes indicated a general trend of decreased mRNA levels compared to uninfected control animals. Although, statistically significant differences were not observed, greatest suppression of relative expression (RE) was identified for IP-10 (RE = 0.198), IFN-β (RE = 0.269), IL-12 (RE = 0.275), and IL-2 (RE = 0.312). Increased relative expression was detected for IL-6 (RE = 2.065). Overall, this data demonstrates that during the FMDV carrier state in cattle, viral persistence is associated with epithelial cells of the nasopharynx in the upper respiratory tract and decreased levels of mRNA for several immunoregulatory cytokines in the infected tissues.

No MeSH data available.


Related in: MedlinePlus

Immunohistochemical detection of persistent FMDV in nasopharyngeal mucosa.A) Dorsal soft palate (caudal), steer #626, 37 dpe, FMDV O1 Manisa. FMDV non-structural protein localized to multiple cells within basal layers of crypt epithelium (arrows) as well as a single cell within associated lymphoid follicle (arrow head). 4x magnification, scale bar 200μm. Anti-FMDV 3D monoclonal antibody. Micropolymer alkaline phosphatase. Gill s hematoxylin counterstain. Inset; 20x magnification of region indicated in dashed rectangle in A, scale bar 50μm. B), 40x magnification of region within dashed rectangle in Fig 3A inset. Large, polygonal, immunopositive cells are within basal epithelium, scale bar 25 μm. C) Dorsal nasopharynx (rostral) steer #625, 37 dpi, FMDV O1 Manisa. FMDV structural protein localized to scarce individual cells within superficial and deeper layers of MALT-associated surface epithelium (arrows). 10x magnification, scale bar 100 μm. Anti-FMDV VP1 monoclonal antibody. Micropolymer alkaline phosphatase. Gill’s hematoxylin counterstain. D) 40x magnification of region within dashed rectangle in Fig 3C, superficial cells containing FMDV antigen are squamous, whereas deeper immunopositive cell is polygonal, scale bar 25 μm.
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pone.0125698.g003: Immunohistochemical detection of persistent FMDV in nasopharyngeal mucosa.A) Dorsal soft palate (caudal), steer #626, 37 dpe, FMDV O1 Manisa. FMDV non-structural protein localized to multiple cells within basal layers of crypt epithelium (arrows) as well as a single cell within associated lymphoid follicle (arrow head). 4x magnification, scale bar 200μm. Anti-FMDV 3D monoclonal antibody. Micropolymer alkaline phosphatase. Gill s hematoxylin counterstain. Inset; 20x magnification of region indicated in dashed rectangle in A, scale bar 50μm. B), 40x magnification of region within dashed rectangle in Fig 3A inset. Large, polygonal, immunopositive cells are within basal epithelium, scale bar 25 μm. C) Dorsal nasopharynx (rostral) steer #625, 37 dpi, FMDV O1 Manisa. FMDV structural protein localized to scarce individual cells within superficial and deeper layers of MALT-associated surface epithelium (arrows). 10x magnification, scale bar 100 μm. Anti-FMDV VP1 monoclonal antibody. Micropolymer alkaline phosphatase. Gill’s hematoxylin counterstain. D) 40x magnification of region within dashed rectangle in Fig 3C, superficial cells containing FMDV antigen are squamous, whereas deeper immunopositive cell is polygonal, scale bar 25 μm.

Mentions: Tissues within which FMDV or FMDV RNA were detected were further examined by conventional microscopy, immunohistochemistry (Fig 3) and MIF microscopy (Fig 4) with anti-FMDV-VP1 (capsid) and anti-FMDV-3D (non-structural) primary antibodies. In all cases, epithelia were consistently intact with no evidence of erosion, ulceration or microvesiculation. Lymph nodes and MALT were populated by appropriate quantity of cells with normal tissue architecture. FMDV antigen was microscopically localized within a subset of analyzed tissues. Within nasopharyngeal epithelia, FMDV structural and non-structural proteins were identified by immunohistochemistry as scarce individual cells or as small clusters of up to10 immunopositive cells within basal and/or superficial layers of epithelium (Figs 3 and 4). Epithelial segments containing FMDV antigen were typically closely associated with MALT follicles and were often lining the opening or lumens of crypts. Additionally, rare single cells containing FMDV protein were localized to subepithelial lymphoid regions. Simultaneous immunostaining for host cell antigens indicated that FMDV-positive cells were morphologically and phenotypically consistent with epithelial cells (cytokeratin+/MHCII-/CD11c-; Fig 4). In some tissues, cytokeratin-/MHCII+ intra-epithelial cells (presumptive DC) were in close proximity, but were never observed to contain FMDV antigen.


Persistent Foot-and-Mouth Disease Virus Infection in the Nasopharynx of Cattle; Tissue-Specific Distribution and Local Cytokine Expression.

Pacheco JM, Smoliga GR, O'Donnell V, Brito BP, Stenfeldt C, Rodriguez LL, Arzt J - PLoS ONE (2015)

Immunohistochemical detection of persistent FMDV in nasopharyngeal mucosa.A) Dorsal soft palate (caudal), steer #626, 37 dpe, FMDV O1 Manisa. FMDV non-structural protein localized to multiple cells within basal layers of crypt epithelium (arrows) as well as a single cell within associated lymphoid follicle (arrow head). 4x magnification, scale bar 200μm. Anti-FMDV 3D monoclonal antibody. Micropolymer alkaline phosphatase. Gill s hematoxylin counterstain. Inset; 20x magnification of region indicated in dashed rectangle in A, scale bar 50μm. B), 40x magnification of region within dashed rectangle in Fig 3A inset. Large, polygonal, immunopositive cells are within basal epithelium, scale bar 25 μm. C) Dorsal nasopharynx (rostral) steer #625, 37 dpi, FMDV O1 Manisa. FMDV structural protein localized to scarce individual cells within superficial and deeper layers of MALT-associated surface epithelium (arrows). 10x magnification, scale bar 100 μm. Anti-FMDV VP1 monoclonal antibody. Micropolymer alkaline phosphatase. Gill’s hematoxylin counterstain. D) 40x magnification of region within dashed rectangle in Fig 3C, superficial cells containing FMDV antigen are squamous, whereas deeper immunopositive cell is polygonal, scale bar 25 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4440813&req=5

pone.0125698.g003: Immunohistochemical detection of persistent FMDV in nasopharyngeal mucosa.A) Dorsal soft palate (caudal), steer #626, 37 dpe, FMDV O1 Manisa. FMDV non-structural protein localized to multiple cells within basal layers of crypt epithelium (arrows) as well as a single cell within associated lymphoid follicle (arrow head). 4x magnification, scale bar 200μm. Anti-FMDV 3D monoclonal antibody. Micropolymer alkaline phosphatase. Gill s hematoxylin counterstain. Inset; 20x magnification of region indicated in dashed rectangle in A, scale bar 50μm. B), 40x magnification of region within dashed rectangle in Fig 3A inset. Large, polygonal, immunopositive cells are within basal epithelium, scale bar 25 μm. C) Dorsal nasopharynx (rostral) steer #625, 37 dpi, FMDV O1 Manisa. FMDV structural protein localized to scarce individual cells within superficial and deeper layers of MALT-associated surface epithelium (arrows). 10x magnification, scale bar 100 μm. Anti-FMDV VP1 monoclonal antibody. Micropolymer alkaline phosphatase. Gill’s hematoxylin counterstain. D) 40x magnification of region within dashed rectangle in Fig 3C, superficial cells containing FMDV antigen are squamous, whereas deeper immunopositive cell is polygonal, scale bar 25 μm.
Mentions: Tissues within which FMDV or FMDV RNA were detected were further examined by conventional microscopy, immunohistochemistry (Fig 3) and MIF microscopy (Fig 4) with anti-FMDV-VP1 (capsid) and anti-FMDV-3D (non-structural) primary antibodies. In all cases, epithelia were consistently intact with no evidence of erosion, ulceration or microvesiculation. Lymph nodes and MALT were populated by appropriate quantity of cells with normal tissue architecture. FMDV antigen was microscopically localized within a subset of analyzed tissues. Within nasopharyngeal epithelia, FMDV structural and non-structural proteins were identified by immunohistochemistry as scarce individual cells or as small clusters of up to10 immunopositive cells within basal and/or superficial layers of epithelium (Figs 3 and 4). Epithelial segments containing FMDV antigen were typically closely associated with MALT follicles and were often lining the opening or lumens of crypts. Additionally, rare single cells containing FMDV protein were localized to subepithelial lymphoid regions. Simultaneous immunostaining for host cell antigens indicated that FMDV-positive cells were morphologically and phenotypically consistent with epithelial cells (cytokeratin+/MHCII-/CD11c-; Fig 4). In some tissues, cytokeratin-/MHCII+ intra-epithelial cells (presumptive DC) were in close proximity, but were never observed to contain FMDV antigen.

Bottom Line: Immunomicroscopy indicated that within persistently infected mucosal tissues, FMDV antigens were rarely detectable within few epithelial cells in regions of mucosa-associated lymphoid tissue (MALT).Transcriptome analysis of persistently infected pharyngeal tissues by qRT-PCR for 14 cytokine genes indicated a general trend of decreased mRNA levels compared to uninfected control animals.Overall, this data demonstrates that during the FMDV carrier state in cattle, viral persistence is associated with epithelial cells of the nasopharynx in the upper respiratory tract and decreased levels of mRNA for several immunoregulatory cytokines in the infected tissues.

View Article: PubMed Central - PubMed

Affiliation: Plum Island Animal Disease Center, Foreign Animal Disease Research Unit, Agricultural Research Service, United States Department of Agriculture, Plum Island, NY, United States of America.

ABSTRACT
Tissues obtained post-mortem from cattle persistently infected with foot-and-mouth disease virus (FMDV) were analyzed to characterize the tissue-specific localization of FMDV and partial transcriptome profiles for selected immunoregulatory cytokines. Analysis of 28 distinct anatomic sites from 21 steers infected with FMDV serotype A, O or SAT2, had the highest prevalence of overall viral detection in the dorsal nasopharynx (80.95%) and dorsal soft palate (71.43%). FMDV was less frequently detected in laryngeal mucosal tissues, oropharyngeal mucosal sites, and lymph nodes draining the pharynx. Immunomicroscopy indicated that within persistently infected mucosal tissues, FMDV antigens were rarely detectable within few epithelial cells in regions of mucosa-associated lymphoid tissue (MALT). Transcriptome analysis of persistently infected pharyngeal tissues by qRT-PCR for 14 cytokine genes indicated a general trend of decreased mRNA levels compared to uninfected control animals. Although, statistically significant differences were not observed, greatest suppression of relative expression (RE) was identified for IP-10 (RE = 0.198), IFN-β (RE = 0.269), IL-12 (RE = 0.275), and IL-2 (RE = 0.312). Increased relative expression was detected for IL-6 (RE = 2.065). Overall, this data demonstrates that during the FMDV carrier state in cattle, viral persistence is associated with epithelial cells of the nasopharynx in the upper respiratory tract and decreased levels of mRNA for several immunoregulatory cytokines in the infected tissues.

No MeSH data available.


Related in: MedlinePlus