Patient 1 was a 26 years old female with no known risk factor. She presented on April 7, 2020 an isolated anosmia-agueusia. Three days later she felt a deep asthenia. She tested positive for SARS-CoV-2 by reverse transcriptase-polymerase chain reaction (RT-PCR) on April 12, 2020. She continued to experience a profound asthenia for 15 days, and completely healed except for the dysnosmia, which was still partially present at day 100.
Patient 2 was a male, 51 with no risk factor besides age. He worked with the patient 1 on April 8. He started to slightly cough on April 11, 2020. The following day, he felt tired, sub-febrile with an increasing cough. He consulted for a suspicion of COVID-19 at a hospital emergency department on April 12, 2020. At the initial examination, patient 2 had a polypnea at 32 respirations/minutes. The blood gas showed a hypoxemia with a PaO2 at 72 %, while a lymphopenia at 680 lymphocytes/mm3 was noted on the blood cell count. The chest computed-tomography scanner was normal, and the nasopharyngeal RT-PCR was positive for SARS-CoV-2. Patient 2 was discharged from the emergency room with a diagnosis of a mild form of COVID-19. He recovered at home within 15 days without major clinical complication, besides a month-long asthenia.
The SARS-CoV-2 strains from both patients were sequenced from naso-pharyngeal samples by MinION technology, following Artic protocol by PCR tiling . Data were analyzed according to the bio-informatic protocol developed by the Artic consortium. Both patients were infected by the same strain, its sequence harboring 7 SNPs compared to the reference genome Wuhan/Hu-1/2019 (NCBI Nucleotide – NC_045512, GenBank –MN908947) and belonging to the G3b phylum , thus carrying the recently identified D614G mutation . On August 1st and 2nd, 2020, the two sequences were deposited on the GISAID platform with accession ID EPI_ISL_505003 and EPI_ISL_506041 for patient 1 and 2 respectively.
The humoral immune response of both patients was followed serially for up to 100 days. An in-house enzyme-linked immuno-sorbent assay (ELISA) was developed for detecting IgG against SARS-CoV-2 adapted from a work made by Florian Krammer . The ELISA detection was based on the receptor-binding domain (RBD) recognition of the SARS-CoV-2 spike (S)-glycoprotein. ELISA results are presented as optical density (OD) ratio obtained by dividing the average OD of duplicate wells from that of the corresponding blank non-coated wells.
For each time point, the presence of nAbs was also sought by a seroneutralisation assay performed on Vero cells using the Institut Pasteur SARS-CoV-2 reference strain in a BSL3 facility.
Both patients rapidly developed an IgG immune response against RBD as they were positive within 12 days, then marked a steep increase followed by a plateau and a slow decrease (Figure 1). Patient 1 had a stronger IgG anti-RBD response while presenting a pauci-symptomatic infection. Patient 2 had also a robust anti-RBD response, while presenting mild clinical symptoms, that included blood desaturation as measured initially. Strikingly, patient 1 did only develop a very moderate neutralizing immune response with low nAb titers that turned negative by day 100, suggesting that virus clearance and the clinical recovery occurred independently of the nAb response.