Molecular Diagnosis of COVID-19; Biosafety and Pre-analytical Recommendations.

From the beginning of the COVID-19 epidemic, clinical laboratories around the world have been involved with tests for detection of SARS-CoV-2. At present, RT-PCR (real-time reverse transcription polymerase chain reaction assay) is seen as the gold standard for identifying the virus. Many factors are involved in achieving the highest accuracy in this test, including parameters related to the pre-analysis stage. Having instructions on the type of sample, how to take the sample, and its storage and transportation help control the interfering factors at this stage. Studies have shown that pre-analytical factors might be the cause of the high SARS-CoV-2 test false-negative rates. Also, the safety of personnel in molecular laboratories is of utmost importance, and it requires strict guidelines to ensure the safety of exposed individuals and prevent the virus from spreading. Since the onset of the outbreak, various instructions and guidelines have been developed in this field by the institutions and the Ministry of Health of each country; these guidelines are seriously in need of integration and operation. In this study, we try to collect all the information and research done from the beginning of this pandemic in December 2019 - August 2022 concerning biosafety and protective measures, sample types, sampling methods, container, and storage solutions, sampling equipment, and sample storage and transportation for molecular testing of SARS-CoV-2.

Detection of Human Boca Virus in Gastric Adenocarcinoma.

Background: Gastric cancer is one of the most common cancers worldwide. Human bocavirus (HBoV), a recently isolated virus, has been investigated for its role in many respiratory and enteric diseases. Few studies have reported its presence in solid tumors, such as lung and colon cancers. The aim of this study was to detect the presence of the HBoV1 genome in gastric adenocarcinoma, which has not yet been evaluated. Methods: Formalin-fixed paraffin-embedded (FFPE) blocks of 189 gastric tumors and 50 blocks of non-tumor gastric tissue products from elective weight reduction operations were collected. DNA extraction and real-time polymerase chain reaction (PCR) were performed for HBoV1 detection. DNA sequencing was performed using ABI Genetic Analyzer series 3500. Results: The mean age of the patients was 60±13.33 years. Tumors were more common in males than females (2.5/1). HBoV1 PCR was positive in 34 (18%) cases of GC and 10 (20%) cases of chronic gastritis (P>0.05). There was no association between age, sex, stage, and histologic subtype of the tumor and HBoV1 positivity (P>0.05) in tumor samples. The rate of intestinal metaplasia and presence of lymphoid stroma were also not more frequent in HBoV1-positive tumors (P>0.05). Conclusion: The HBoV1 can be detected in a relatively high proportion of Iranian patients with gastric cancer (18%) with no predilection for specific subtypes and no association with the degree of lymphocytic infiltration. HBoV1 can also be observed in approximately 20% of chronic gastritis cases. Further comprehensive studies are needed to elucidate the role of HBoV1 in gastric cancer development.

Perinatal Presentation of a Congenital Intramedullary Capillary Hemangioma in a Neonate Born with Hydrocephalus and Paraplegia.

Spinal intramedullary capillary hemangioma is a rare condition. Although most spinal/spinal cord hemangiomas are inborn in origin, perinatal presentation is uncommon. We present a neonate with an intramedullary hemangioma, born with communicating hydrocephalus and complete paraplegia. Spinal imagining showed an intradural mass with hemorrhagic foci, a low-lying conus medullaris, and scalloping of dorsal elements. Ventriculoperitoneal shunting was performed in an emergent setting, with cerebrospinal fluid showing a brownish appearance with high viscosity which implied a possible old hemorrhage. Tissue sample was taken from the spinal lesion in an elective setting. Histopathological examination revealed capillary lobules and extralobular large vessels resembling abnormal veins. The mentioned clues suggested that this congenital hemangioma had become symptomatic from the prenatal period. Though it is a are event, it should be kept in mind while evaluating a neonate with communicating hydrocephalus and weakness of extremities with or without cutaneous hemangiomatous stigmata.

Chediak Higashi Syndrome with Hemophagocytic Lymphohistiocytosis.

INTRODUCTION: Chediak-Higashi syndrome (CHS) is caused by dysfunction of lysosomal trafficking and presents with hypopigmentation, bleeding tendencies, neurological symptoms, and NK cell dysfunction. Hemophagocytic lymphohistiocytosis (HLH) can complicate CHS due to the abnormal function of NK cells. CASE PRESENTATION: This 1.5-year-old light-skinned gray-haired girl microscopically had abnormal hair pigment clumps and lilac inclusions in the myeloid series, characteristic of CHS. She presented with HLH, requiring treatment with etoposide and dexamethasone followed by cyclosporine and dexamethasone. CONCLUSION: CHS is one of the underlying primary causes of HLH.

Ameloblastic Fibro-Odontoma.

Introduction: Ameloblastic fibro-odontoma (AFO) is a benign odontogentic tumor without an aggressive behavior, unlike the similar ameloblastic fibroma. Case Presentation: A 9-year-old boy, with tooth eruption failure, underwent enucleation and curettage of a well-defined variable radiolucent and radio-opaque right mandible lesion. There was odontogenic epithelium with peripheral palisading in a loose myxoid stroma as well as a disorganized component of dentin, enamel, and cementum, features of an AFO. Conclusion: AFO is an odontogenic mixed tumor of epithelium and mesenchyme.

CNS tumor with BCOR internal tandem duplication.

CNS tumor with BCOR internal tandem duplication is a recently proposed malignant tumor. The patient was an 18-month-old boy with torticollis and vomiting due to cerebellar hemispheric mass with extension to cerebellopontine angle and foramen magnum. Histopathologic examination of the tumor showed a moderately cellular tumor with microcystic formation, myxoid change, and atypical rosettes resembling Homer Wright rosettes. Illumina TruSight RNA Pan-Cancer NGS of the tumor genome revealed BCOR gene exon 15 internal tandem duplications.

Alteration of Plasma Amino Acid Concentrations in Iranian Children with COVID-19.

COVID-19 is an acute viral disease that has so far infected more than 200 million and killed more than four million worldwide. It affects the immune system and other organs. Here, we investigated the level of free plasma amino acids in COVID-19 patients and compared them with non-COVID-19 subjects. We also compared amino acids levels in critically ill patients admitted to the intensive care unit (ICU) with non-ICU patients and expired and recovered patients. Twenty-six COVID-19 patients and 32 non-COVID-19 subjects were included in the study. The mean of glutamic acid, serine, glycine, threonine, phenylalanine, leucine, lysine, alanine, arginine, aspartic acid, and ornithine was significantly higher in cases than controls. In addition, the mean of glutamine was significantly lower in patients than controls (443.89 ± 254.31 vs. 651.73 ± 107.38, PV < 0.001). Low level of glutamine and isoleucine was seen in the majority of ICU and expired patients, respectively. Logistic regression analysis showed low level of isoleucine as a predictor variable in mortality (P = 0.02, EXP (B) = 16.5, and CI 95% = (1.48, -183.07)). There was a positive and significant relationship between some amino acids levels, serum liver enzymes, and sodium concentrations. There was also a significant but negative correlation between histidine levels, ESR, and ferritin. Phenylalanine had a highly positive relationship with serum procalcitonin in patients (R 2 = 0.534, PV = 0.015). Our studies have shown the alteration of plasma amino acids concentration in COVID-19 patients. These changes are more evident in critically ill and at-risk patients.

Evaluation of human bocavirus as well as other well-known viral etiologic agents by PCR in the cerebrospinal fluid of children with clinical impression of viral meningoencephalitis referred to the Children's Medical Center in Tehran, Iran, from 2019 to 2020.

Viral meningoencephalitis is one of the most important diseases that most commonly affect children. In many cases of viral meningoencephalitis, the underlying cause of the disease is not identified, raising the possibility of a variety of pathogens that are not routinely tested. Bocaviruses belong to a newly identified class of viruses that have been reported in some studies to be associated with viral encephalitis. In the present study, we investigated the prevalence of bocaviruses and other viruses in the patients suspected of having viral encephalitis and their associations with various demographic and clinical variables. Two hundred patients with suspected viral meningoencephalitis referred to Children's Medical Center were studied from 2019 to 2020. Age, sex, length of hospitalization, and course of the disease were gathered. Cerebrospinal fluid (CSF) samples were taken from the patients and subjected to biochemical examinations and PCR to identify the underlying cause. Bocaviruses were detected in none of the DNA samples extracted from the CSF specimens. The most identified organisms were mumps and enteroviruses. In 92% of cases, the underlying cause was not identified. PCR-based identification of the underlying causes of viral meningoencephalitis in CSF specimens was not successful in most cases. Bocavirus was not found in any of the collected CSF samples. Further studies are required for drawing more accurate conclusions.

Juvenile Polyps with Osseous Metaplasia: Report of Two Pediatric Cases and Review of the Literature.

BACKGROUND: Osseous metaplasia is considered as a response to insults such as chronic inflammation or mucin extravasation in colorectal polyps. CASE REPORT: we report two cases of osseous metaplasia in juvenile (retention) polyps as incidental histopathological findings in a case of rectal juvenile polyposis and a patient with solitary rectal juvenile polyp. CONCLUSION: Osseous metaplasia can occur in colorectal juvenile polyps and is considered a probable response to mucin extravasation and/or chronic inflammation. The clinical and prognostic significance of osseous metaplasia in these polyps is unknown.

Jordans' Anomaly as a Red Flag for Neutral Lipid Storage Diseases.

Background: Jordans' anomaly is characterized by lipid vacuoles in granulocytes which are observed in neutral lipid storage diseases like Chanarin-Dorfman syndrome. Case report: This six-year-old boy had skin ichthyosis, elevated liver enzymes, and prominent vacuoles in neutrophils, eosinophils, monocytes, and basophils (Jordans' anomaly), leading to the diagnosis of Chanarin-Dorfman Syndrome, which was successfully treated with medium-chain triglyceride oil. Conclusion: Jordans' anomaly is a red flag for the diagnosis of neutral lipid storage diseases especially in patients with ichthyosis and elevated liver enzymes.

Large intrathoracic lipoblastoma.

Lipoblastoma is a rare benign tumor of primitive white fat in children, which usually involves the trunk and extremities. It rarely involves retroperitoneum, head, neck, and mediastinum. Herein, the authors introduce a rare presentation of lipoblastoma as a large intrathoracic mass.

Henoch-Schönlein Purpura (IgA Vasculitis) in Association with Thyrotoxicosis.

Graves' disease is the most common cause of hyperthyroidism, which is characterized by thyroid antibodies and the following clinical manifestations: goiter, ophthalmopathy, and pretibial myxedema. On the other hand, Henoch-Schönlein purpura is an IgA-mediated small-vessel vasculitis. Review of the literature showed a relationship between propylthiouracil overdose and the following Henoch-Schönlein purpura (IgA vasculitis) as a side effect. The patient was a 31-year-old woman with a chief complaint of tremor and significant weight loss who contracted pruritic palpable purpura during her disease course. Then, she underwent the treatment of hyperthyroidism by methimazole which intensified her cutaneous lesions. The diagnosis of Henoch-Schönlein purpura (IgA vasculitis) was confirmed after skin biopsy. Finally, she was treated with colchicine, prednisolone, and radioiodine ablation, which caused her lesions to disappear. The temporal priority of pruritic palpable skin lesions to hyperthyroidism treatment with methimazole suggested that Henoch-Schönlein purpura (IgA vasculitis) was related to hyperthyroidism and was intensified by antithyroid agents in this patient.

JAK2, CALR, and MPL Mutation Profiles in BCR-ABL Negative Myeloproliferative Neoplasms, a Referral Center Experience in the Middle East.

BACKGROUND & OBJECTIVE: JAK2, CALR, and MPL genes play pivotal roles in the pathogenesis of BCR-ABL negative myeloproliferative neoplasms. This study was conducted to evaluate the frequency of JAK2, CALR, and MPL mutations in BCR-ABL negative myeloproliferative neoplasms and their association with demographic data and hematologic parameters in a referral center, in the Middle East. METHODS: Seventy-one patients with BCR-ABL negative myeloproliferative neoplasms were evaluated for JAK2 V617F, CALR type 1, CALR type 2, and MPL by allele-specific PCR and conventional PCR from 2018 to 2019. RESULTS: Twenty-three patients were categorized as polycythemia vera, JAK2 V617F was observed in 91.3% of these cases. Thirty-eight patients were classified as essential thrombocythemia of which 52.6% showed JAK2 V617F, 18.4% demonstrated CALR type 1, 7.9% denoted CALR type 2 and there was no mutation reported in 21.1%. Seven patients were recognized as primary myelofibrosis and exhibited JAK2 V617F mutation in 57.1%, CALR type 1 in 14.3 %, CALR type 2 in 14.3% and no mutation in 14.3%. Three patients were diagnosed as MPN, unclassifiable and 33.3% revealed JAK2 V617F mutation, and no mutation was found in 66.6%. The age (59.15±13.10) and neutrophil percent (70.78±10.14) were higher in patients with JAK2 V617 mutation compared to other mutations (P=0.000, and P=0.03). Platelet count was significantly higher in patients with CALR type 1 mutation (1240400± 402053) (P=0.000). CONCLUSION: JAK2 V617F was associated with patients' higher age and higher neutrophil count in CBC. CALR mutation had an association with higher platelet count. No MPL mutation was found in this study and it seems that its frequency is lower than what is expected in this region.

Clinical Misdiagnosis of COVID-19 Infection with Confusing Clinical Course.

BACKGROUND: Similarities in the febrile course and other manifestations of some diseases may lead to clinical misdiagnosis of COVID-19 infection. Here, we report a case in a young child with a potentially confusing clinical course. Case Presentation. A 29-month-old boy presented with a 2-month history of fever. His PCR test for COVID-19 was positive, and there was pleural effusion plus positive findings in the lower left lobe of the lung on computed tomography scan. Mid-sized splenomegaly was found on abdominal ultrasound, and laboratory tests disclosed pancytopenia. In light of the atypical lymphocyte counts in laboratory tests, he underwent bone marrow aspiration. The suggested diagnosis was hemophagocytic lymphohistiocytosis, and prednisolone was initiated. Subsequently, Leishman-Donovan bodies were seen in the bone marrow aspirate, and treatment was started with amphotericin, which led to clinical improvement. CONCLUSION: In cases with vague clinical symptoms in tropical countries where other infectious diseases occur, possible simultaneous infection should be considered even during a pandemic. Familiarity with the possible differential diagnoses and appropriate, step-by-step consideration to rule out other possible causes are needed in all situations, and the coexistence of infectious disease should be considered in evaluating the clinical conditions of patients in tropical countries.

Isolated primary spinal epidural hydatid cyst in a child with progressive paraparesis.

BACKGROUND: Primary spinal hydatid cyst is a rare but serious condition which occurs in about 1% of patients with hydatid disease. This disease may result in severe spinal cord compression presenting with various neurological deficits. CASE REPORT: A 3-year-old boy was referred to our center with progressive weakness of lower limbs, frequency, and urinary incontinence. His parents did not report back pain during child's illness. Lumbar magnetic resonance imaging (MRI) revealed an extradural well-defined thin-walled cystic lesion at L4 to S3 vertebral levels without enhancement. The cyst had compressed the thecal sac associated with bone scalloping of vertebral bodies and posterior elements. Osteoplastic laminectomy of L4-L5 and laminectomy of S1-S3 was done. Intraoperative and histopathological findings indicated an epidural hydatid cyst. CONCLUSION: Although spinal hydatid cysts are rare but might be considered in the differential diagnosis of any patient with signs and symptoms of spinal cord compression. The pathophysiology, clinical manifestations, epidemiology, diagnosis, and surgical treatment of extradural hydatid cyst are discussed.

Case Report: Encephalitis Caused by Balamuthia mandrillaris in a 3-Year-Old Iranian Girl.

It is about half a century since free-living amoebae were recognized as pathogenic organisms, but there is still much we should learn about these rare fatal human infectious agents. A recently introduced causative agent of granulomatous amoebic encephalitis, Balamuthia mandrillaris, has been reported in a limited number of countries around the world. A 3-year-old girl was referred to our tertiary hospital because of inability to establish a proper diagnosis. She had been experiencing neurologic complaints including ataxia, altered level of consciousness, dizziness, seizure, and left-sided hemiparesis. The patient's history, physical examination results, and laboratory investigations had led to a wide differential diagnosis. Computed tomography (CT) scan and magnetic resonance imaging analyses revealed multiple mass lesions. As a result, the patient underwent an intraoperative frozen section biopsy of the brain lesion. The frozen section study showed numerous cells with amoeba-like appearances in the background of mixed inflammatory cells. Medications for free-living amoebic meningoencephalitis were administered. PCR assay demonstrated B. mandrillaris as the pathogenic amoeba. Unfortunately, the patient died 14 days after her admission. To our knowledge, this is the first report of B. mandrillaris meningoencephalitis in the Middle East and the first time we have captured the organism during a frozen-section study.