Silent Infection of B and CD8+ T Lymphocytes by Influenza A Virus in Children with Tonsillar Hypertrophy.

Influenza A viruses (IAVs) cause more than 2 million annual episodes of seasonal acute respiratory infections (ARI) and approximately 500,000 deaths worldwide. Depending on virus strain and host immune status, acute infections by IAV may reach sites other than the respiratory tract. In the present study, IAV RNA and antigens were searched for in tissues of palatine tonsils and adenoids removed from patients without ARI symptoms. A real-time reverse transcriptase PCR (RT-PCR) screening revealed that 8 tissue samples from 7 patients out of 103 were positive for IAV. Positive samples were subjected to next-generation sequencing (NGS) and 3 of 8 tissues yielded complete IAV pH1N1 genomes, whereas in 5 samples, the PB1 gene was not fully assembled. Phylogenetic analysis placed tonsil-derived IAV in clusters clearly segregated from contemporaneous Brazilian viruses. Flow cytometry of dispersed tissue fragments and serial immunohistochemistry of paraffin-embedded sections of naturally infected biopsies indicated that CD20+ B lymphocytes, CD8+ T lymphocytes, and CD11c+ cells are susceptible to IAV infection. We sought to investigate whether these lymphoid tissues could be sites of viral replication and sources of viable virus particles. MDCK cells were inoculated with tissue lysates, enabling recovery of one IAV isolate confirmed by immunofluorescence, reverse transcriptase quantitative PCR (RT-qPCR), and NGS. The data indicate that lymphoid tissues not only harbor expression of IAV proteins but also contain infectious virus. Asymptomatic long-term infection raises the possibility of IAV shedding from tonsils, which may have an impact on host-to-host transmission.IMPORTANCE Influenza A virus (IAV) infections are important threats to human health worldwide. Although extensively studied, some aspects of virus pathogenesis and tissue tropism remain unclear. Here, by different strategies, we describe the asymptomatic infection of human lymphoid organs by IAV in children. Our results indicate that IAV was not only detected and isolated from human tonsils but displayed unique genetic features in comparison with those of contemporaneous IAVs circulating in Brazil and detected in swabs and nasal washes. Inside the tissue microenvironment, immune cells were shown to be carrying IAV antigens, especially B and T CD8+ lymphocytes. Taken together, these results suggest that human lymphoid tissues can be sites of silent IAV infections with possible impact on virus shedding to the population.

Prevalence of oropharyngeal high-risk human papillomavirus in tumor-free tonsil tissue in adults.

PURPOSE: To determine the prevalence of oropharyngeal high-risk human papillomavirus (HPV) in patients undergoing tonsillectomy by detection of high-risk HPV in tonsil tissues using the in situ hybridization (ISH) technique. MATERIALS AND METHODS: The patients who underwent tonsillectomy between 2014 and 2018 were examined retrospectively. The pediatric cases and patients who underwent tonsillectomy due to malignancy were excluded. The study included 270 adult cases selected by age and gender randomization. The tonsillar tissue of each case was re-examined by the pathology department, and the presence of high-risk HPV was investigated via the ISH technique. Multiple logistic regression models were used for predictions of different factors. RESULTS: The prevalence of high-risk HPV in the 270 patients (male: 154 [57%]; female: 116 [43%]; mean age: 36.44 ± 12.87 years) was found to be 6.7% (n = 18). The prevalence was found 8.4% in men and 4.3% in women; 8.9% in cases under the age of 40 and 2.9% in cases over the age of 40; and 10.9% in patients who underwent tonsillectomy for infectious indications and 2.3% for non-infectious indications. Multivariate analysis identified that the infectious indications for tonsillectomy were significantly associated with high-risk HPV positivity (OR 5.328; p = 0.009). CONCLUSIONS: The prevalence of oropharyngeal high-risk HPV was found to be 6.7% and higher in younger people and men. Additionally, the HPV positivity was found to be higher in patients who underwent tonsillectomy for infectious indications. To our knowledge, this is the first study that reports the correlation between recurrent tonsil infections and HPV positivity in tonsil tissue.

Human adenovirus replication and persistence in hypertrophic adenoids and palatine tonsils in children.

The role of human adenovirus (HAdV) infection in different acute diseases, such as febrile exudative tonsillitis, conjunctivitis, and pharyngoconjunctival fever is well established. However, the relationships, if any, of HAdV persistence and reactivation in the development of the chronic adenotonsillar disease is not fully understood. The present paper reports a 3-year cross-sectional hospital-based study aimed at detecting and quantifying HAdV DNA and mRNA of the HAdV hexon gene in adenoid and palatine tonsil tissues and nasopharyngeal secretions (NPS) from patients with adenotonsillar hypertrophy or recurrent adenotonsillitis. HAdV C, B, and E were detectable in nearly 50% of the patients, with no association with the severity of airway obstruction, nor with the presence of recurrent tonsillitis, sleep apnea or otitis media with effusion (OME). Despite the higher rates of respiratory viral coinfections in patients with HAdV, the presence of other viruses, including DNA and RNA viruses, had no association with HAdV replication or shedding in secretions. Higher HAdV loads in adenoids showed a significant positive correlation with the presence of sleep apnea and the absence of OME. Although this study indicates that a significant proportion (~85%) of individuals with chronic adenotonsillar diseases have persistent nonproductive HAdV infection, including those by HAdV C, B, and E, epithelial and subepithelial cells in tonsils seem to be critical for HAdV C production and shedding in NPS in some patients, since viral antigen was detected in these regions by immunohistochemistry in four patients, all of which were also positive for HAdV mRNA detection.

Unusual manifestation of disseminated herpes simplex virus type 2 infection associated with pharyngotonsilitis, esophagitis, and hemophagocytic lymphohisitocytosis without genital involvement.

BACKGROUND: Herpes simplex virus (HSV) has various presentations, depending on the patient's immune status, age, and the route of transmission. In adults, HSV type 1 is found predominantly in the oral area, and HSV type 2 (HSV-2) is commonly found in the genital area. HSV-2 infection without genital lesions is uncommon. Herein we report a unique case of pharyngotonsillitis as an initial manifestation of disseminated HSV-2 infection without genital involvement. CASE PRESENTATION: A 46-year-old male was admitted to our hospital with a 1-week history of fever and sore throat. His past medical history included hypereosinophilic syndrome diagnosed at age 45 years. Physical examination revealed throat congestion, bilaterally enlarged tonsils with exudates, tender cervical lymphadenopathy in the left posterior triangle, and mild epigastric tenderness. The laboratory data demonstrated bicytopenia, elevated liver enzyme levels, and hyperferritinemia. A bone marrow smear showed hypocellular marrow with histiocytes and hemophagocytosis. The diagnosis of HLH was confirmed, and the patient was treated with methylprednisolone pulse therapy on days 1-3. On day 5, despite initial improvement of the fever and sore throat, multiple, new, small bullae developed on the patient's face, trunk, and extremities. Additional testing showed that he was positive for HSV-specific immunoglobulin M and immunoglobulin G. Disseminated HSV infection was suspected, and intravenous acyclovir (10 mg/kg every 8 h) was begun. A subsequent direct antigen test of a bulla sample was positive for HSV-2. Moreover, tonsillar and esophageal biopsies revealed viral inclusion bodies. Immunohistochemical staining and a quantitative real-time polymerase chain reaction (PCR) assay confirmed the presence of HSV-2. Disseminated HSV-2 infection with multiple bullae, tonsillitis, esophagitis, and suspected hepatic involvement was diagnosed. After a 2-week course of intravenous acyclovir, his hematological status and liver function normalized, and his cutaneous skin lesions resolved. He was discharged on day 22 in good general health and continued taking oral valacyclovir for viral suppression due to his immunosuppressed status. CONCLUSION: Disseminated HSV-2 infection should be considered as one of the differential diagnoses in patients with pharyngotonsillitis and impaired liver function of unknown etiology even if there are no genital lesions.

Detection of high-risk human papillomavirus in the tonsils of galilee region adults and young adults undergoing tonsillectomy.

INTRODUCTION: The presence of high-risk HPV in non-malignant tonsil tissues from patients who underwent tonsillectomy in the Galilee area might explain the low incidence of HPV-related oropharyngeal malignancy in the country. The aim of this retrospective study was to study the prevalence of human papillomavirus (HPV) in non-malignant tonsillectomy specimens of adults in the Galilee area. MATERIALS AND METHODS: We conducted a retrospective analysis of all tonsil samples in our medical center. Tonsils from patients over 20 years of age who underwent tonsillectomy for the indication of recurrent tonsillitis and sleep apneas were eligible. Samples of formalin-fixed paraffin-embedded (FFPE) tonsillar tissue were tested for the presence of HPV DNA using polymerase chain reaction and by p16 immunohistochemistry. RESULTS: Of the 71 tonsil samples, age range 20-65, none were positive for HPV DNA. Fifty-two FFPE specimens of tonsil with HPV-positive cancer tested positive by the same method served as positive controls. CONCLUSIONS: HPV DNA is rare in non-malignant tonsil tissues of young adults and adults who underwent tonsillectomy in the Galilee area. Further research should be done in larger cohorts.

[CLINICAL AND IMMUNOLOGICAL CRITERIA FOR THE ADVERSE COURSE OF INFECTIOUS MONONUCLEOSIS IN CHILDREN].

The article presents the results of our own studies to determine the criteria for the adverse variants of the course of infectious mononucleosis (IM) in children. The study was conducted in the regional children's infectious clinical hospital in Kharkov. 161 children aged three to fifteen years were under observation with diagnosis of infectious moninucleosis. Out of 161 ill children, 140 (86.9%) had moderate severity of disease, and 21 (13.1%) had severe forms. All children were prescribed standard clinical and laboratory-instrumental examinations. The diagnosis of IM was verified by PCR (detection of VEB DNA in the blood) and ELISA (anti-VEB Ig M and Ig G). In 140 children (86.9%) IM proceeded sharply, smoothly (the first group), in 21 (13.1%) - unfavorably (wave and / or prolonged course) - the second group. The groups were comparable according to age, the severity of the disease and other parameters. All children received therapy according to approved protocols (Order of the Ministry of Health of Ukraine No. 354 of 09.07.2004). Immune status of children was assessed by determining the relative contents of CD3 +, CD4 +, CD8 +, CD16 +, CD19 + blood cells with appropriate monoclonal antibodies, serum IgA, IgM, IgG concentration by Mancini and interleukin (IL) -1ß cytokine response and - 4, tumor necrosis factor (TNF α) is a solid-phase enzyme-linked immunosorbent assay. Based on the results of observations, it was established that the prognostically unfavorable criteria of IМ at the stages of manifestation of disease include: generalized lymphadenopathy involving 5-6 groups of lymph nodes and a significant increasing of them, purulent tonsillitis, marked increasing of size of liver and spleen on the background of anemia, thrombocytopenia, neutropenia and the absence of atypical mononuclears in the complete blood count. There is a depression of the cellular link and an increase in the humoral mechanisms of immune responses in case of development of adverse course of IM.

Multiplex detection in tonsillar tissue of all known human polyomaviruses.

BACKGROUND: In the past few years, eleven new human viruses have joined the two previously known members JCPyV and BKPyV of the Polyomaviridae family, by virtue of molecular methods. Serology data suggest that infections with human polyomaviruses (HPyVs) occur since childhood and the viruses are widespread in the general population. However, the viral persistence sites and transmission routes are by and large unknown. Our previous studies demonstrated that the four new HPyVs - KIPyV, WUPyV, MCPyV and TSPyV - were present in the tonsils, and suggested lymphoid tissue as a persistent site of these emerging human viruses. We developed a Luminex-based multiplex assay for simultaneous detection of all 13 HPyVs known, and explored their occurrence in tonsillar tissues of children and adults mostly with tonsillitis or tonsillar hypertrophy. METHODS: We set up and validated a new Luminex-based multiplex assay by using primer pairs and probes targeting the respective HPyV viral protein 1 (VP1) genes. With this assay we tested 78 tonsillar tissues for DNAs of 13 HPyVs. RESULTS: The multiplex assay allowed for simultaneous detection of 13 HPyVs with high analytical sensitivity and specificity, with detection limits of 100-102 copies per microliter, and identified correctly all 13 target sequences with no cross reactions. HPyV DNA altogether was found in 14 (17.9%) of 78 tonsils. The most prevalent HPyVs were HPyV6 (7.7%), TSPyV (3.8%) and WUPyV (3.8%). Mixed infection of two HPyVs occurred in one sample. CONCLUSIONS: The Luminex-based HPyV multiplex assay appears highly suitable for clinical diagnostic purposes and large-scale epidemiological studies. Additional evidence was acquired that the lymphoid system plays a role in HPyV infection and persistence. Thereby, shedding from this site during reactivation might take part in transmission of the newly found HPyVs.

Isolated Unilateral Soft Palate Palsy Following Tonsillopharyngitis Caused by Epstein-Barr Virus Infection.

Soft palate palsy with or without pharyngeal or laryngeal paralysis or facial palsy has been described in the literature fewer than 40 times. Of these cases, isolated unilateral soft palate palsy (occurring without any other neurologic manifestation) has been rare. Most cases of isolated soft palate palsy were termed idiopathic in the past. In a few cases, a concomitant viral infection such as varicella zoster, herpes simplex, measles, and coxsackie A9 was diagnosed. We describe a case of isolated soft palate palsy, following a pharyngotonsillitis caused by Epstein-Barr virus. This viral infection has been linked in the past with bilateral facial nerve palsy and radial nerve palsy. Our patient recovered spontaneously over a period of 1 month.

Detection of human bocavirus in children with upper respiratory tract infection by polymerase chain reaction.

BACKGROUND: The aim of this study was to investigate whether in children with middle ear effusions (MEE), adenoid and tonsil tissues are associated with human bocavirus (HBoV). MATERIALS AND METHODS: A total of 124 patients (56 females (45.2%) and 68 males (54.8%)) with chronic adenotonsillitis and serous otitis media under the age of 15 were recruited. Two hundered four samples (113 adenoid (55.4%), 68 tonsil (33.3%), and 23 middle ear effusion (11.3%)) were analyzed for the presence of HBoV using polymerase chain reaction (PCR). RESULTS: HBoV was detected in only 6 (4.8%) adenoid tissue samples each belonging to a different patient. CONCLUSIONS: Our findings are consistent with the results of other studies, reporting approximately 5 - 10% of the samples being positive for HBoV. To understand the detailed role of HBoV in the etiology of RTI in children, further studies would be needed.

Occurrence of human bocaviruses and parvovirus 4 in solid tissues.

Human bocaviruses 1-4 (HBoV1-4) and parvovirus 4 (PARV4) are recently discovered human parvoviruses. HBoV1 is associated with respiratory infections of young children, while HBoV2-4 are enteric viruses. The clinical manifestations of PARV4 remain unknown. The objective of this study was to determine whether the DNAs of HBoV1-4 and PARV4 persist in human tissues long after primary infection. Biopsies of tonsillar tissue, skin, and synovia were examined for HBoV1-4 DNA and PARV4 DNA by PCR. Serum samples from the tissue donors were assayed for HBoV1 and PARV4 IgG and IgM antibodies. To obtain species-specific seroprevalences for HBoV1 and for HBoV2/3 combined, the sera were analyzed after virus-like particle (VLP) competition. While HBoV1 DNA was detected exclusively in the tonsillar tissues of 16/438 individuals (3.7%), all of them ≤8 years of age. HBoV2-4 and PARV4 DNAs were absent from all tissue types. HBoV1 IgG seroprevalence was 94.9%. No subject had HBoV1 or PARV4 IgM, nor did they have PARV4 IgG. The results indicate that HBoV1 DNA occurred in a small proportion of tonsils of young children after recent primary HBoV1 infection, but did not persist long in the other tissue types studied, unlike parvovirus B19 DNA. The results obtained by the PARV4 assays are in line with previous results on PARV4 epidemiology.

Clinical features of 167 children with the novel influenza A (H1N1) virus infection in Xi'an, China.

Since its first recognition, the 2009 pandemic influenza A (H1N1) virus rapidly spread worldwide. We observed the clinical characteristics of 167 hospitalized patients who were confirmed by testing pharyngeal or nasopharyngeal swabs with the use of a real-time reverse-transcriptase polymerase chain reaction (RT-PCR) assay. The mean age of the 167 hospitalized patients was 4.1 years, and 58.7% were male. The most common symptoms and signs were fever (91.6%), cough (82.6%), pharyngeal congestion (95.2%), and swollen tonsils (34.1%). The major complications were bronchitis (19.2%), bronchial pneumonia (10.8%), neutropenia (49.7%), and leukopenia (38.9%). The duration of hospitalization, fever and the course of disease in the patients who were treated with oseltamivir were shorter than in those who were treated with ribavirin. All of the patients fully recuperated from the 2009 epidemic influenza A (H1N1) infection with one exception.

Presence of herpesviruses in adenoid tissues of children with adenoid hypertrophy and chronic adenoiditis.

OBJECTIVES: The aim of study was to determine the presence of some of the herpesviruses including herpes simplex virus (HSV), Epstein-Barr virus (EBV), and cytomegalovirus (CMV) in adenoid tissues of children with adenoid hypertrophy (AH) and chronic adenoiditis (CA) and to investigate the potential role of the herpesviruses in patogenesis of AH and CA. PATIENTS AND METHODS: A total of 72 patients (41 boys, 31 girls; mean age 4 years and 2 months; range 2 to 9 years) who underwent adenoidectomy or adenotonsillectomy (with or without placement of a ventilation tube) in our clinic between October 2007 and May 2008, were included. The patients were divided into two groups, as AH group (n=42) and the CA group (n=30). Adenoid tissues collected from patients in both groups were analyzed by polymerase chain reaction (PCR) for the presence of HSV, EBV and CMV-DNA. RESULTS: The results of the PCR indicated that 33.3% in the AH group and 36.6% in the CA group were herpesvirus DNA positive. Among the herpesviruses studied, HSV-DNA was detected at the highest level (14.2% and 16.6%, respectively) in both groups, although the difference between the groups was not significant. EBV-DNA positiveness was 11.9% and CMV-DNA was 4.7% in the AH group, whereas, EBV-DNA positiveness was 13.3% and CMV-DNA was 6.6% in the CA group. CONCLUSION: Herpesviruses were determined at a high rate in adenoid tissue of children with AH and CA, suggesting that there may be a potential relationship between the presence of herpesviruses and occurrence of AH and CA in children. However, more extensive studies are required to elucidate the role of herpesviruses in the pathogenesis of AH or CA.

[Chromogenic in situ hybridization in diagnostics of Epstein-Barr-virus infection in chronic tonsillitis].

In the issue we demonstrate results of Epstein-Barr virus (EBV) detection by chromogenic in situ hybridization (CISH) in epithelial and lymphoid cells of the palatine tonsil in patients with chronic tonsillitis. Virus genome detections were performed using RNA-probes with digoxigenin-labeled oligonucleotides which target EBV RNA, notably RNA-transcripts of virus genomic DNA. The obtained data confirm the virus lymphotropism and also tropism to epithelial cells of both surface and cryptal epithelium of the palatine tonsil. CISH method in combination with immunohistochemical identification of virus protein products opens new possibilities for clinicopathological monitoring of the different clinical forms of the chronic tonsillitis, as well as new horizon for understanding intrinsic role of EBV in tonsillar pathology.

[Pathogenetic ground of including reamberin and cycloferon combination into the therapy program for patients with severe cases of acute tonsillitis of a mixed viral/bacterial etiology].

The increase of severe cases of acute tonsillitis (AT) is presently marked. Severe cases of AT disturb immune and metabolic homoeostasis initiating the development of disease. Therapy optimization is required to select the best treatment. In patients with severe cases of AT of mixed viral/bacterial etiology before the treatment it is revealed the increase of general activity of lactatedehydrigenase (LDH) and increase of the level of cathode "anaerobic" factions LDH4+5 and the decline of concentration ATP in the blood. There was a compensatory rise of level of ADP and АМP. The substantial decline of serum interferon (CIF) activity and diminishing maintenance of α-interferon (α-IFN) and γ-interferon (γ-IFN) in the blood of the patients, that testified to oppressing of interferonogenesis. Treatment of severe cases of AT of mixed viral/bacterial etiology of modern detoxic preparation reamberin and immunoactive preparation cycloferon combination positively influences the studied laboratory indexes. The improvement of power metabolism is marked, that was characterized by normalization of level adenine nucleotides (ATP, АDP, АМP) and general activity of LDH and its izoenzimes spectrum. At the same time the increase of CIF level is set, maintenances α-IFN and γ-IFN in the blood, that testified to the improvement of interferonogenesis. The results demonstrate the therapeutic potential of reamberin and cycloferon combination for treatment of patients with AT of mixed viral/bacterial etiology.

The prevalence of human papillomavirus in paediatric tonsils in Southwestern Ontario.

OBJECTIVE: To determine the prevalence of human papillomavirus in paediatric tonsils in Southwestern Ontario, Canada. MATERIALS AND METHODS: Patients aged 0-18 years undergoing tonsillectomy were recruited. Two specimens (left and right tonsils) were collected from each participant. Tonsillar DNA was analysed using quantitative polymerase chain reaction to determine the presence of human papillomavirus subtypes 6, 11, 16 or 18. RESULTS: A total of 102 patients, aged 1-18 years (mean age of 5.7 years), were recruited. Ninety-nine surveys were returned. There were 44 females (44.4 per cent) and 55 males (55.6 per cent). Forty patients (40.4 per cent) were firstborn children and 73 (73.7 per cent) were delivered vaginally. Six mothers (6.1 per cent) and one father (1.0 per cent) had prior known human papillomavirus infection, and one mother (1.0 per cent) had a history of cervical cancer. All tonsil specimens were negative for human papillomavirus subtypes 6, 11, 16 and 18. CONCLUSION: No human papillomavirus subtypes 6, 11, 16 or 18 were found in paediatric tonsil specimens from Southwestern Ontario.

Towards evidence-based emergency medicine: best BETs from the Manchester Royal Infirmary. Can inflammatory markers distinguish streptococcal from viral tonsillitis?

A short-cut review was performed to evaluate whether inflammatory markers such as C reactive protein (CRP), erythrocyte sedimentation rate (ESR), white cell count (WCC) and procalcitonin (PCT) are able to discriminate between streptococcal and viral tonsillitis, enabling a reduction in the overuse of antibiotics. Eight studies with a total of 1031 participants were found. The author, date and country of publication, patient group studied, study type, relevant outcomes, results and study weaknesses of these papers are tabulated. The clinical bottom line is that WCC, CRP and PCT levels are higher in patients with streptococcal tonsillitis compared to patients with tonsillitis or pharyngitis without group A streptococcus isolated from a throat swab. Which of these markers has the best test performance characteristics requires further study.

[Efficacy of Ingavirin in the combined treatment of ARVI complicated by tonsillitis].

This paper was designed to report the results of an open randomized comparative study of therapeutic efficacy of Ingavirin used for the treatment of adult patients with acute respiratory viral infections (ARVI) complicated by lacunar tonsillitis. The study has demonstrated that the combined application of an antibacterial agent and Ingavirin showing antiviral and anti-inflammatory properties produces a significantly more pronounced therapeutic effect compared with the standard treatment of the disease in question. This finding gives reason to recommend Ingavirin for the wide clinical application as an agent for combined therapy of acute respiratory viral infections.