Short beak and dwarfism syndrome among Pekin ducks: First detection, full genome sequencing, and immunohistochemical signals of novel goose parvovirus in tongue tissue.

Novel goose parvovirus (NGPV) is continuously threatening the global duck industry, as it causes short beak and dwarfism syndrome among different duck breeds. In this study, we investigated the viral pathogenesis in the tongue of affected ducks, as a new approach for deeper understanding of the syndrome. Seventy-three, 14- to 60-day-old commercial Pekin ducks were clinically examined. Thirty tissue pools of intestine and tongue (15 per tissue) were submitted for molecular identification. Clinical signs in the examined ducks were suggestive of parvovirus infection. All examined ducks had short beaks. Necrotic, swollen, and congested protruding tongues were recorded in adult ducks (37/73, 51%). Tongue protrusion without any marked congestion or swelling was observed in 20-day-old ducklings (13/73, 18%), and no tongue protrusion was observed in 15-day-old ducklings (23/73, 32%). Microscopically, the protruding tongues of adult ducks showed necrosis of the superficial epithelial layer with vacuolar degeneration. Glossitis was present in the nonprotruding tongues of young ducks, which was characterized by multifocal lymphoplasmacytic aggregates and edema in the propria submucosa. Immunohistochemical examination displayed parvovirus immunolabeling, mainly in the tongue propria submucosa. Based on polymerase chain reaction, goose parvovirus was detected in 9 out of 15 tongue sample pools (60%). Next-generation sequencing confirmed the presence of a variant goose parvovirus that is globally named NGPV and closely related to Chinese NGPV isolates. Novel insights are being gained from the study of NGPV pathogenesis in the tongue based on molecular and immunohistochemical identification.

Natural Canine Distemper Virus Infection in Linnaeus's 2-Toed Sloths (Choloepus didactylus).

An outbreak of canine distemper virus in a private zoo in eastern Tennessee in July 2016 led to fatal clinical disease in 5 adult, wild-caught Linnaeus's 2-toed sloths (Choloepus didactylus). Clinical signs included hyporexia, lethargy, mucopurulent nasal discharge, and oral and facial ulcers. At necropsy, affected animals had crusts and ulcers on the lips, nose, tongue, and oral cavity. Microscopically, all sloths had widespread, random, hepatic necrosis; lymphoid depletion; and bronchointerstitial pneumonia. The central nervous system did not contain gross or histopathologic lesions in any of the 5 sloths, although immunoreactivity for viral antigen was present within vessel walls. Epithelial cells and histiocytes within numerous organs contained intranuclear and intracytoplasmic inclusions and occasional syncytial cells. Canine distemper virus was confirmed with immunohistochemistry and virus isolation. Viral sequencing identified the novel American-4 strain prevalent in eastern Tennessee wildlife. This is the first pathologic characterization of canine distemper virus infection in sloths (family Choloepodidae, order Pilosa) and emphasizes the significant morbidity and mortality in this species.

Coinfection of a lingual lesion with bovine papular stomatitis virus and bovine papillomavirus.

To date, there have been no reports of coinfection with bovine papular stomatitis virus (BPSV) and bovine papillomavirus (BPV) in the same lesion. In the present study, one lingual papilloma-like sample was collected at an abattoir from the tongue of a 31-month-old Japanese black cow. Coinfection with BPSV and BPV was confirmed by histopathology, immunohistochemistry, PCR and RT-PCR. The evidence for coinfection with BPSV and BPV in the same lesion and an association of BPV with lingual papillomatosis will contribute to future epidemiological studies of these two viruses.

A pathological study of the tongues of rabid dogs in the Philippines.

During rabies virus infections, the minor salivary glands are one of the important organs for virus replication and excretion into the oral cavity. However, details of pathological findings and viral antigen distribution in the minor salivary glands remain poorly understood. In this study, we conducted pathological tests on the tongues of 71 rabid dogs in the Philippines; the minor salivary glands (von Ebner's glands, lingual glands), circumvallate papilla, autonomic ganglia, and skeletal muscles were evaluated. Inflammatory changes were observed in the von Ebner's glands of 20/71 dogs, in the circumvallate papilla of 10/71, and in the tongue muscle of 1/71. Conversely, no morphological changes were observed in the lingual glands and autonomic ganglia. Viral antigens were detected via immunohistochemistry-based methods in the cytoplasm of the acinar epithelium in the von Ebner's glands of all 71 dogs. Virus particles were confirmed in the intercellular canaliculi and acinar lumen via electron microscopy. In the autonomic ganglia, viral antigens were detected in 67/71 rabid dogs. Viral antigens were detected in the taste buds of all 71 dogs, and were distributed mainly in type II and III taste bud cells. In tongue muscle fibers, viral antigens were detected in 11/71 dogs. No virus antigens were detected in lingual glands. These findings suggest that rabies virus descends in the tongue along the glossopharyngeal nerve after proliferation in the brain, and von Ebner's glands and taste buds are one of the portals of virus excretion into the saliva in rabid dogs.

Thermal inactivation of foot and mouth disease virus in extruded pet food.

The risk of importing foot and mouth disease, a highly contagious viral disease of livestock, severely restricts trade and investment opportunities in many developing countries where the virus is present. This study was designed to investigate the inactivation of foot and mouth disease virus (FMDV) by heat treatments used in extruded commercial pet food manufacture. If extrusion could be shown to reliably inactivate the virus, this could potentially facilitate trade for FMDV-endemic countries. The authors found that there was no detectable virus following: i) treatment of FMDVspiked meat slurry at 68°C for 300 s; ii) treatment of FMDV-spiked slurry and meal mix at 79°C for 10 or 30 s, or iii) treatment of homogenised bovine tongue epithelium, taken from an FMDV-infected animal, at 79°C for 10 s. This corresponds to an estimated 8 log10 reduction in titre (95% credible interval: 6 log10 -13 log10). Furthermore, the authors found that the pH of the slurry and meal mix was sufficient to inactivate FMDV in the absence of heat treatment. This demonstrates that heat treatments used in commercial pet food manufacture are able to substantially reduce the titre of FMDV in infected raw materials.

En raison du risque d'introduction de la fièvre aphteuse, une maladie virale très contagieuse affectant le bétail, les échanges internationaux et les possibilités d'investissement de nombreux pays en développement sont soumis à des restrictions majeures. La présente étude vise à déterminer si les traitements thermiques appliqués lors de la fabrication industrielle d'aliments extrudés destinés aux animaux de compagnie sont efficaces pour inactiver le virus de la fièvre aphteuse dans ces produits. Si la procédure d'extrusion avait pour propriété démontrée d'inactiver le virus de manière fiable, les pays où le virus de la fièvre aphteuse est présent à l'état endémique pourraient accéder plus facilement aux échanges internationaux. Les auteurs n'ont pas trouvé de virus détectable à l'issue des opérations suivantes : i) traitement à 68 °C pendant 300 s d'une bouillie de viandes à laquelle avait été ajoutée une quantité connue de virus de la fièvre aphteuse ; ii) traitement à 79 °C pendant 10 ou 30 s d'un mélange de bouillie et de pâté auquel avait été ajoutée une quantité connue de virus de la fièvre aphteuse ; iii) traitement à 79 °C pendant 10 s d'un échantillon homogénéisé d'épithélium lingual prélevé d'un bovin atteint de fièvre aphteuse. Ces résultats correspondent à une réduction estimée du titre de 8 log10 (intervalle de confiance à 95 % : 6 log10 ­13 log10). En outre, les auteurs ont constaté que le pH du mélange de bouillie et de pâté était de nature à inactiver le virus de la fièvre aphteuse, en l'absence de traitement thermique. Ces résultats démontrent que les traitements thermiques utilisés lors de la fabrication industrielle d'aliments pour animaux de compagnie permettent de réduire de manière importante les titres du virus de la fièvre aphteuse dans les matières premières infectées.

El riesgo de importar fiebre aftosa, enfermedad vírica muy contagiosa que afecta al ganado vacuno, restringe sobremanera el comercio y las oportunidades de inversión en muchos países en desarrollo en los que el virus está presente. Los autores describen un estudio encaminado a analizar el método de inactivación del virus por calor que se emplea en la fabricación industrial de piensos extrusionados para animales de compañía. Si quedaba demostrado que la extrusión es un método fiable para inactivar el virus de la fiebre aftosa, ello podría abrir perspectivas comerciales a todos aquellos países donde dicho virus es endémico. Los autores observaron que no había presencia detectable de virus después de: i) mantener a 68°C durante 300 segundos una emulsión cárnica enriquecida con virus de la fiebre aftosa; ii) mantener a 79°C durante 10 o 30 segundos una emulsión y una mezcla de harinas enriquecidas con el virus; o iii) mantener a 79°C durante 10 segundos un homogenado de epitelio lingual bovino procedente de un animal infectado por el virus. Ello corresponde a una reducción estimada de la titulación de log10 8 (intervalo de confianza al 95%: log10 6 ­ log10 13). Además, los autores observaron que, en ausencia de tratamiento térmico, el pH de la emulsión y la mezcla de harinas bastaba para inactivar el virus de la fiebre aftosa, lo que demuestra que los tratamientos térmicos utilizados en la fabricación industrial de piensos pueden reducir sustancialmente el título de virus de la fiebre aftosa en materias primas infectadas.

Association between human papilloma virus/Epstein-Barr virus coinfection and oral carcinogenesis.

BACKGROUND: The recent epidemic of head and neck squamous cell carcinomas associated with human papilloma virus (HPV) has not addressed its association with lymphoid tissue in the oropharynx or the potential role of Epstein-Barr virus (EBV)/HPV coinfection. METHODS: The prevalence of HPV and EBV infection/coinfection and CD21 mRNA expression were determined in normal and cancerous tissues from the oropharynx using in situ hybridization (ISH), p16, and quantitative reverse transcriptase PCR (qRT-PCR). The effects of coinfection on tumorigenicity were evaluated using proliferation and invasion assays. RESULTS: Normal oropharynx, tonsil, non-cancer base of tongue (BOT), and BOT from sleep apnea patients demonstrated EBV positivity ranging from 7% to 36% depending on the site and methods of detection used (qRT-PCR or ISH). Among non-malignant BOT samples, HPV positivity was noted only in 20%. The percent of tonsil and BOT cancers positive for HPV (up to 63% and 80%, respectively) or coinfected with HPV/EBV (up to 25% and 70%, respectively) were both significantly associated with cancer status. Notably, HPV/EBV coinfection was observed only in malignant tissue originating in lymphoid-rich oropharynx sites (tonsil, BOT). CD21 mRNA (the major EBV attachment receptor) was detected in tonsil and BOT epithelium, but not in soft-palate epithelium. Coinfected cell lines showed a significant increase in invasiveness (P < 0.01). CONCLUSIONS: There is a high prevalence of HPV/EBV infection and coinfection in BOT and tonsil cancers, possibly reflecting their origins in lymphoid-rich tissue. In vitro, cells modeling coinfection have an increased invasive potential.

Spectrum of VP1 region genetic variants in the foot-and-mouth disease virus serotype O populations derived from infected cattle tongue epithelium.

RNA virus population exists as a complex distribution of non-identical but closely related sequences known as viral quasispecies. Variant strains are selected from this quasispecies population in response to changing environment. The quasispecies dynamics of a virus existing within an infected host differs from that in a cell culture-adapted population. This study was carried out to explore the genetic variations present in the VP1 coding region of the foot-and-mouth disease (FMD) virus serotype O derived directly from infected cattle tongue epithelium. Molecular clonal populations of two serotype O strains belonging to lineages Ind2001 (IND 30/2011) and PanAsia2 (IND 5/2011) were sequenced at VP1 coding region. For IND 30/2011, 19 clones were sequenced and analysis showed variations at 12 nucleotide positions (nt) resulting in 8 amino acid (aa) replacements. Similarly, for IND 5/2011 virus, 18 clones were sequenced, of which six showed nt variations leading to 3 aa replacements. Most of the variable positions mapped to the surface-exposed loops and some of them were found in the neutralizing antigenic sites (position 81, 149, 169, 186 and 202 of IND 30/2011 and 141 of IND 5/2011), which potentially could be beneficial in rapid adaptive evolution of the virus by giving rise to antigenic variants to overcome neutralizing antibodies. These findings encourage further research into the landscape of the viral quasispecies population in vivo and its implication for viral ecology.

Use of quantitative diffusion-weighted magnetic resonance imaging to predict human papilloma virus status in patients with oropharyngeal squamous cell carcinoma.

Although identification of human papilloma virus (HPV) status in oropharyngeal squamous cell carcinoma (OPSCC) is essential in predicting treatment response, no imaging modality can currently determine whether a tumor is HPV-related. In this retrospective study, 26 patients with OPSCC confined to the lateral wall or the base of tongue underwent neck magnetic resonance imaging, using T1-, T2- and diffusion-weighted imaging (DWI). Apparent diffusion coefficients (ADCs) in a region of interest covering the largest available primary tumor area of OPSCC on a single slice of the ADC map were calculated using two b values (0 and 1,000 s/mm(2)). Mean and minimum ADCs were compared with HPV status, using p16 immunohistochemistry as a surrogate marker for HPV infection. Mean and minimum ADCs for HPV(+) OPSCC were significantly lower than those for HPV(-) OPSCC. A cut-off value of mean ADC for HPV(+) OPSCC of 1.027 × 10(-3) mm(2)/s yielded sensitivity and specificity of 83.33 and 78.57%, respectively. In conclusion, the present study indicates that ADC could be used to predict HPV status in patients with OPSCC.

Impaired metal perception and regulation of associated human foliate papillae tongue transcriptome in long-COVID-19.

Chemosensory impairment is an outstanding symptom of SARS-CoV-2 infections. We hypothesized that measured sensory impairments are accompanied by transcriptomic changes in the foliate papillae area of the tongue. Hospital personnel with known SARS-CoV-2 immunoglobulin G (IgG) status completed questionnaires on sensory perception (n = 158). A subcohort of n = 141 participated in forced choice taste tests, and n = 43 participants consented to donate tongue swabs of the foliate papillae area for whole transcriptome analysis. The study included four groups of participants differing in IgG levels (≥ 10 AU/mL = IgG+; < 10 AU/mL = IgG-) and self-reported sensory impairment (SSI±). IgG+ subjects not detecting metallic taste had higher IgG+ levels than IgG+ participants detecting iron gluconate (p = 0.03). Smell perception was the most impaired biological process in the transcriptome data from IgG+/SSI+ participants subjected to gene ontology enrichment. IgG+/SSI+ subjects demonstrated lower expression levels of 166 olfactory receptors (OR) and 9 taste associated receptors (TAS) of which OR1A2, OR2J2, OR1A1, OR5K1 and OR1G1, as well as TAS2R7 are linked to metallic perception. The question raised by this study is whether odorant receptors on the tongue (i) might play a role in metal sensation, and (ii) are potential targets for virus-initiated sensory impairments, which needs to be investigated in future functional studies.

Severe acute respiratory syndrome coronavirus 2 pathology and cell tropism in tongue tissues of COVID-19 autopsies.

Since 2019, Coronavirus Disease 2019(COVID-19) has affected millions of people worldwide. Except for acute respiratory distress syndrome, dysgeusis is also a common symptom of COVID-19 that burdens patients for weeks or permanently. However, the mechanisms underlying taste dysfunctions remain unclear. Here, we performed complete autopsies of five patients who died of COVID-19. Integrated tongue samples, including numerous taste buds, salivary glands, vessels, and nerves were collected to map the pathology, distribution, cell tropism, and receptor distribution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the tongue. Our results revealed that all patients had moderate lymphocyte infiltration around the salivary glands and in the lamina propria adjacent to the mucosa, and pyknosis in the epithelia of taste buds and salivary glands. This may be because the serous acini, salivary gland ducts, and taste buds are the primary sites of SARS-CoV-2 infection. Multicolor immunofluorescence showed that SARS-CoV-2 readily infects Keratin (KRT)7+ taste receptor cells in taste buds, secretory cells in serous acini, and inner epithelial cells in the ducts. The major receptors, angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine subtype 2 (TMPRSS2), were both abundantly expressed in these cells. Viral antigens and receptor were both rarely detected in vessels and nerves. This indicates that SARS-CoV-2 infection triggers pathological injury in the tongue, and that dysgeusis may be directly related to viral infection and cellular damage.

Striated muscle involvement in experimental oral infection by herpes simplex virus type 1.

Herpes simplex virus type 1 is one of the most frequent causes of oral infection in humans, especially during early childhood. Several experimental models have been developed to study the pathogenesis of this virus but all of them employed adult animals. In this work, we developed an experimental model that uses mice younger than 4 days old, to more closely resemble human infection. Mice were infected subcutaneously with the prototype strain McIntyre of Herpes simplex-1, and the progression of infection was studied by immunoperoxidase. All animals died within 24-72 h post-infection, while viral antigens were found in the oral epithelium, nerves and brain. The most striking result was the finding of viral antigens in the nucleus and cytoplasm of cells belonging to striated muscles. Organotypic cultures of striated muscles were performed, and viral replication was observed in them by immunocytochemistry, electron microscopy and viral isolation. We conclude that the infection of striated muscles is present from the onset of oral infection and, eventually, could explain some clinical observations in humans.

Morphological and Immunopathological Aspects of Lingual Tissues in COVID-19.

COVID-19, a recently emerged disease caused by SARS-CoV-2 infection, can present with different degrees of severity and a large variety of signs and symptoms. The oral manifestations of COVID-19 often involve the tongue, with loss of taste being one of the most common symptoms of the disease. This study aimed to detect SARS-CoV-2 RNA and assess possible morphological and immunopathological alterations in the lingual tissue of patients who died with a history of SARS-CoV-2 infection. Sixteen cadavers from 8 SARS-CoV-2 positive (COVID-19+) and 8 negative (COVID-19-) subjects provided 16 tongues, that were biopsied. Samples underwent molecular analysis through Real-Time RT-PCR for the detection of SARS-CoV-2 RNA. Lingual papillae were harvested and processed for histological analysis and for immunohistochemical evaluation for ACE2, IFN-γ and factor VIII. Real-Time RT-PCR revealed the presence of SARS-CoV-2 RNA in filiform, foliate, and circumvallate papillae in 6 out of 8 COVID-19+ subjects while all COVID-19- samples resulted negative. Histology showed a severe inflammation of COVID-19+ papillae with destruction of the taste buds. ACE2 and IFN-γ resulted downregulated in COVID-19+ and no differences were evidenced for factor VIII between the two groups. The virus was detectable in most COVID-19+ tongues. An inflammatory damage to the lingual papillae, putatively mediated by ACE2 and IFN-γ in tongues from COVID-19+ cadavers, was observed. Further investigations are needed to confirm these findings and deepen the association between taste disorders and inflammation in SARS-CoV-2 infection.

Epstein-Barr virus infection of polarized tongue and nasopharyngeal epithelial cells.

Epstein-Barr virus (EBV) initially enters the body through the oropharyngeal mucosa and subsequently infects B lymphocytes through their CD21 (CR2) complement receptor. Mechanisms of EBV entry into and release from epithelial cells are poorly understood. To study EBV infection in mucosal oropharyngeal epithelial cells, we established human polarized tongue and pharyngeal epithelial cells in culture. We show that EBV enters these cells through three CD21-independent pathways: (i) by direct cell-to-cell contact of apical cell membranes with EBV-infected lymphocytes; (ii) by entry of cell-free virions through basolateral membranes, mediated in part through an interaction between beta1 or alpha5beta1 integrins and the EBV BMRF-2 protein; and (iii) after initial infection, by virus spread directly across lateral membranes to adjacent epithelial cells. Release of progeny virions from polarized cells occurs from both their apical and basolateral membranes. These data indicate that multiple approaches to prevention of epithelial infection with EBV will be necessary.

Mixed HPV-related Neuroendocrine Carcinoma and HPV-related Squamous Cell Carcinoma of the Base of Tongue in a Patient With Incidental Identification of Synchronous Metastatic Papillary Thyroid Carcinoma.

BACKGROUND: While it is not uncommon in patients with head and neck cancer to present with multiple metachronous primary neoplasms, rarely do these present as a singular mass composed of intertwined, histologically distinct malignant tumors. Sometimes referred to as collision tumors, these entities are poorly understood and only appear in a handful of case studies in the literature. CASE REPORT: Here we present a 58-year-old male diagnosed with a human papillomavirus-related collision tumor consisting of oropharyngeal squamous cell carcinoma and small-cell neuroendocrine carcinoma, as well as an incidentally discovered metastatic thyroid papillary carcinoma, despite an unremarkable thyroid gland. The patient underwent transoral robotic base-of-tongue resection and partial pharyngectomy with selective neck dissection followed by chemoradiotherapy. At the 18-month follow-up the patient was doing well. His thyroid was normal and no recurrent or metastatic carcinoma was identified on the computed tomography and positron-emission tomography/computed tomography imaging findings. CONCLUSION: To the best of our knowledge, this is the first such case in English literature. This case demonstrates the importance of tumor morphology and immunohistochemical testing in HPV-related oropharyngeal carcinomas, despite the overall good prognosis of such tumors, due to the possibility of synchronous or colliding primary neoplasms.

Detection of an emerging novel sublineage Ind2001BD1 and lineage PanAsia of foot-and-mouth disease virus serotype O in cattle in Manikgonj district of Bangladesh, 2018.

Background: Foot-and-mouth disease (FMD) is an endemic disease of cloven-hoofed animals in Bangladesh and multiple outbreaks occur every year because of the FMD virus (FMDV). Aim: The aim of the present investigation was to determine the molecular characterization of the VP1 coding region of FMDV serotype O outbreak in cattle. Methods: A total of four tongue epithelial specimens were collected from clinically FMD-positive cattle during June 2018 in Manikgonj district of Bangladesh. Results: All four isolates were recorded positive for FMDV serotype O. The phylogenetic analysis showed that two isolates were clustered within an emerging novel sublineage Ind2001BD1 under lineage Ind2001 of FMDV serotype O, which was identified during 2012-2016 in Bangladesh. One isolate was clustered within the lineage PanAsia of FMDV serotype O and was closely related to an isolate identified in Nepal in 2009. The phylogenetic reconstruction revealed that all the four isolates belong to the Middle East-South Asia topotype. Conclusion: Therefore, multiple lineages of the FMDV serotype O are circulating among the cattle in the outbreak area, which make it more complex for the FMD control program in Bangladesh. A comprehensive study on the genetic characteristics of FMDV across the country is required for effective FMD prevention and control strategy.