Repeatedly negative PCR results in patients with COVID-19 symptoms: Do they have SARS-CoV-2 infection or not?

OBJECTIVE: To point out possible infection with SARS-CoV-2 in symptomatic patients despite repeated negative nasopharyngeal swab tests for SARS-CoV-2. MATERIAL AND METHODS: A retrospective observational study carried out at the Na Bulovce Hospital from the beginning of the pandemic until November 2020 included patients (1) who had symptoms compatible with COVID-19; (2) whose nasopharyngeal swab PCR tests in the presence of acute respiratory infection symptoms yielded two consecutive negative results; (3) in whom SARS-CoV-2 infection was subsequently confirmed by serology. Basic demographic and epidemiological data, symptoms, laboratory test results, X-ray findings and timing of virological tests were analysed for these patients. RESULTS: Seventeen patients met the inclusion criteria, 14 men and three women, aged 19-84 years with a median of 59 years, of whom 14 were hospitalized and three were treated as outpatients. Only seven patients were aware of the previous contact with an infected person. The main symptoms were fever, cough, headache, weakness, fatigue and shortness of breath. Pneumonia was found in 12 patients, four of whom developed respiratory insufficiency requiring ventilatory support. Most patients showed a uniform combination of haematological, biochemical and radio-logical findings: absence of eosinophils and increased polymorphonuclear/lymphocyte ratio; elevation of serum lactate dehydrogenase; elevation of CRP without rise of procalcitonin; typical chest CT or X-ray findings. All patients recovered. Coronavirus antigen test was performed in six patients, with all of them testing negative. SARS-CoV-2 infection was confirmed serologically by the detection of specific IgG and IgA in all 17 patients and also IgM in six patients, not before day 8 of the onset of symptoms. CONCLUSIONS: Our study showed that some patients with acute COVID-19 may test repeatedly negative by nasopharyngeal swab PCR. These cases should be interpreted as a low viral load in the upper respiratory tract rather than false negativity of PCR. Such alternative is not envisaged in the algorithms used. Considering our results, the following recommendation can be made: If, despite negative PCR tests, COVID-19 is still suspected based on clinical symptoms and epidemiological evidence, preliminary diagnosis can be made on the basis of comprehensive assessment of the laboratory test and X-ray findings. Final confirmation of the aetiology relies on serological tests performed two weeks after the onset of symptoms.

Bacterial proteases: production, isolation and utilization in animal nutrition.

Under conditions of submerged fermentation of Bacillus licheniformis strain L-3 in 15-L MBR-Schulzer bioreactor, the maximum production of proteolytic enzymes was achieved in the nutrient medium which contained 1% milk powder, 0.3% yeast autolysate, 0.5% corn starch and malt (20 ml per 100 ml of the medium). The preparation obtained of extracellular alkaline bacterial Ser-protease of the subtilisin type is characterized by optimum pH 9.8-10.2 good up to a temperature stability (65 degrees C) and has molecular weight cca of 26 kDa. The use of chemical mutagens (HNO2, 5-bromouracil) has enabled to select new strains (L-3N and L-3U) whose protease activity is 1.8-2.2 times higher as compared to the original Bacillus licheniformis strain L-3. The addition of these enzymes to the fodder has a positive effect on the retention of nitrogen substances.