Study design and setting
The current prospective clinical trial study included 134 patients with COVID-19, who were randomly selected from the Quarantine Department of Qena University Hospitals, Faculty of Medicine, South Valley University, Qena, Egypt, during the period from May 2020 to August 2020. Prior to start in the study, an institutional ethical committee approval was taken (Ethical approval code: SVU-MED-MBC004-2020-04). A written informed consent was obtained from all participants in this study. Diagnosis of SARS-CoV-2 was based on history of epidemiologic exposure. Clinical manifestations include: (1) respiratory symptoms and/or fever; (2) imaging features of SARS-CoV-2infection; (3) total leucocytic counts showing normal, or reduced lymphocyte count in early stage [30], imaging features of COVID- 19, also diagnosed by real-time reverse transcription-polymerase chain reaction (rRT-PCR) in samples from respiratory tract swabs which were performed at Central Laboratories, Ministry of Health and Population, Cairo, Egypt.
Patients with history of nasal surgery, sinusitis, nasal polyposis allergic rhinitis, history of head injury, or chronic nasal disease were excluded. Also, patients with anosmia and or hyposmia before the diagnosis of COVID-19 were excluded.
Data collections
Demographic data were recorded for all patients including age, sex, BMI, co morbidities and smoking. Full history was taken from all patients with special stress about presence or absence of anosmia (loss of smell) or hyposmia (decrease sense of smell). In addition to subjective smell tests (using coffee or chocolate), formal smell tests may be required which give more accurate level of loss of smell, in that a minimum concentration of a chemical at which the patient can detect, can be given and compared to the average threshold for that patient's age group e.g. chemosensory test and butanol threshold test [31]. In addition, proper examination of nasal cavity and paranasal sinuses was performed.
COVID-19 was categorized into mild, common, severe, and extremely severe in accordance with the 6th edition for Diagnostic Standards for COVID-19 [32]. Consequently, mild COVID-19 was considered to be associated with mild clinical symptoms, with no pneumonia manifestations on imaging. Patients with common COVID-19 had fever, respiratory tract, or other symptoms, and imaging that showed pneumonia. Severe COVID-19 was considered to meet one of the following conditions: (1) Shortness of breath, 30 beats/ min; (2) in the resting state, pulse oxygen saturation< 93%, arterial blood oxygen pressure (PaO2)/oxygen concentration (FiO2) < 300mmHg; or (3) pulmonary imaging showed lesion progression > 50% within 24-48 hours. Extremely severe COVID-19 needed meet one of the following conditions: (1) development of respiratory failure requiring mechanical ventilation; (2) shock; (3) combined organ failure requiring ICU monitoring [33].
The patients with anosmia and / or hyposmia were divided randomly into two groups; the first group included 49 patients with olfactory dysfunction, who received zinc therapy (220mg zinc sulfate equivocal to 50 mg elemental zinc twice daily [34]) plus the Egyptian protocol of treatment of COVID-19) and the second group included 56 patients with olfactory dysfunction, who received the Egyptian protocol of COVID-19 treatment without zinc therapy. Follow up of all included patients till complete recovery of COVID-19 (pharyngeal swap becomes negative) and complete recovery of olfactory symptoms and the recovery durations for the included patients were recorded in days.
Biochemical and molecular assays
- Diagnostic kit (PCR-Fluorescence probing) of Nucleic acid was used for the qualitative estimation of the ORF1ab and a specific conserved sequence of coding nucleocapsid protein N genes of novel corona virus (2019-nCOV). This kit was supplied by Sansure Biotech Inc., Hunan Province, China with catalog No. S3102E, using a 7500 real-time fluorescence quantitative RT-PCR system (Applied Biosystems, Foster City, CA, USA) to detect RNA through fluorescent signal changes [35].
- Serum levels of zinc were measured for all patients, 3 ml of peripheral venous blood samples was collected in plain collection tubes for serum recovery. Samples were left to clot for 30 minutes at 37 °C before centrifugation, and sera obtained were aliquoted into 1-mL cryotubes and stored at -20 °C until biochemical assays of zinc. Zinc level was measured colorimetrically (Spectrum Diagnostics, Cairo, Egypt, Catalog No. 330001) [36-40].
Statistical analysis
Analysis of data was performed using Statistical Program for Social Science (SPSS) version 24. Kolmogorov–Smirnov test was used to check for data normality. Qualitative data was expressed as frequency; numbers and percentages. Parametric quantitative data was expressed as mean ± standard deviation, while median and inter-quartile range used for non-parametric data. For comparison between two groups, the Chi-square test (χ2) was used for qualitative variables, while independent-samples t-test of significance: was used for normally distributed parametric data. For abnormally distributed quantitative variables (non-parametric data), the Mann-Whitney U test was used. Probability (P-value) P-value > 0.05 was considered insignificant, P-value < 0.05 was considered significant; P-value < 0.001 was considered as highly significant.