Immunity to SARS-CoV-2 in a Dialyzed Patient Who Developed COVID-19 Twenty Days After the Second Dose of BNT162b2 Vaccine: a Case Report

IntroductionEnd stage kidney disease (ESKD) and cancer have been identified as risk factors for severe and fatal cases of COVID-19, making vaccination in these patients a priority. Patients suffering from ESKD have a significantly weaker response to common vaccines than general population. However, humoral and cellular immune responses after two doses of RNA-based vaccine BNT162b2 (Pfizer–BioNTech) have been poorly explored in this vulnerable population.Case presentationA 69-year-old male patient was followed for ESKD and myeloma. He developed a severe SARS-CoV-2 pneumonia twenty days after two doses of BNT162b2 vaccine. Whole genome sequencing found that the virus belonged to the 20I/501Y.V1 clade. A serology draws eight days after the 2nd vaccine dose showed positive RBD IgG without neutralizing activity. A serum specimen sampled thirty days after the onset of SARS-CoV-2 infection showed seroconversion against both RBD and N antigens. This specimen was shown to exhibit a frank neutralizing activity. The QuantiFERON® SARS-CoV-2 (Qiagen) showed a positive specific cellular response although the QuantiFERON monitor displayed a weak cellular response. ConclusionsImpaired immunity due to renal failure probably explain the severe pneumonia despite vaccination. The fact that the patient developpe a neutralizing activity and a cellular response after a third stimulation by infection may suggest to systemically administrate a third dose of vaccine in ESKD patients.

Combination of rRT-PCR and Anti-Nucleocapsid/Anti-Spike Antibodies to Characterize Specimens with Very Low Viral SARs-CoV-2 Load: A Real-Life Experience

The objective of the present study was to evaluate the true positivity among people, whose results of initial testing of nasopharyngeal swabs (NPS) showed a very low viral load of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Seventy-seven people detected with low viral loads of SARs-CoV-2 in nasopharyngeal samples (Ct ≥ 35) were enrolled in the study. For this purpose, a second NPS was collected for rRT-PCR (real-time reverse transcription polymerase chain reaction) combined with a pair of serum samples for detection of anti-nucleocapsid (anti-N) and anti-spike (anti-S) antibodies. In 8 people, subsequent examinations indicated an increase in viral loads, thereafter, followed by an increase of anti-N and anti-S antibodies, findings compatible with an early stage of COVID-19 infection. In 9 people, who already had increased anti-N antibodies, subsequent examination showed a decrease or absence of viral load and an increase in antibodies, indicative of a late stage of COVID-19 infection. In 60 people, subsequent examination showed absence of infection (as indicated by absence of viral load and antibodies). We propose that the combination of a second NPS and one serum-specimen, both taken three days after the first NPS, helps significantly to avoid false-positive results.

Identification of Dengue Serotypes using a Single Serum Specimen Algorithm in a Tertiary Care Hospital, Alappuzha, Kerala, India

Introduction: The geographic location of Alappuzha, a district in the South Indian state of Kerala, the distinct weather conditions and frequent natural calamities present a unique ecology that contributes to the prevalence of vector-borne diseases like dengue. Early dengue virus infection can be detected by using a combination of tests on a single serum specimen. Aim: To identify the dengue virus serotypes among hospitalised patients in a South Indian teaching hospital in Alappuzha, Kerala, India. Materials and Methods: Patient samples that tested positive for dengue non-structural protein-1 (NS1) antigen by ELISA were further evaluated for dengue virus RNA by real-time, multiplex reverse transcriptase Polymerase Chain Reaction (PCR) and the serotype was determined. Anonymised patient data was collected using a questionnaire as a data collection tool. The data was analysed for statistical significance. Results: Among 422 non-duplicate patient serum samples received in the Department of Microbiology, in the year 2019, 30 were positive for dengue NS1 antigen by ELISA. Dengue viral RNA was detected in 50% of the samples (15/30). DENV-3 serotype was the most prevalent (nine) followed by DENV-1 (five) and DENV-2 (one). Common presentations of the patients were fever, headache, and myalgia. No statistically significant association was found between a PCR positive result and the presence of warning signs and thrombocytopenia. Conclusion: DENV-3 was the most common serotype in the study population. Early dengue virus infection is associated with varied symptoms.

An Outbreak of SARS in a Diabetes Room of a General Hospital without Infected Medical Staff

Objective To investigate the epidemiologic features of an outbreak of SARS that occurred in a single diabetes room of a general hospital in Beijing in late March 2003. Methods Field investigation was carried out in the ward, the nursing log and the hospitalization medical record of correlative patients were consulted. SARS-CoV in serum specimen from SARS patient was detected by PCR. Results The room where SARS outbreak occurred was on the 13th floor of the 16-story main ward building. There were 6 beds in the room, living with 6 female patients (aged 45-67) who were all hospitalized due to type 2 diabetes. On March 24, 2003, Patient 1 began to have a fever and cough, chest X-ray showed pneumonia. Five and six days later, Patient 2 and Patient 3 began to have a fever, respectively. Finally, all of these 3 patients died. Their beds were all at the same side of the room, and the other 3 patients at the opposite side were not infected. Serum SARS CoV-RNA of the Patient 3 was positive by nest-PCR. The daughter-in-law of Patient 1 who accompanied Patient 1 by the bedside several days, mainly near the window, upwind of Patient 1, was not infected. Medical staff, family members and visitors of the 6 patients were not infected. Conclusions This outbreak was not transmitted by aerosol. The distance droplets travels could be up to 3.43 meters. Droplet spread has direction, and the droplets direction of propagation is closely related with the wind direction and speed. Those at the downwind position of SARS patients were susceptible to be infected. Medical staff wore face masks and good natural ventilation of this ward building may be important reasons for the prevention of infection.

USFDA-GUIDELINE BASED VALIDATION OF TESTING METHOD FOR RIFAMPICIN IN INDONESIAN SERUM SPECIMEN

Regarding a new regulation from Indonesia FDA (Badan POM-RI), all new non patent drugs should show bioequivalence with the originator drug prior to registration. Bioequivalence testing (BE-testing) has to be performed to the people that represented of population to which the drug to be administrated. BE testing need a valid bio-analytical method for certain drug target and group of population. This research report specific validation of bio-analysis of Rifampicin in Indonesian serum specimen in order to be used for BE testing. The extraction was performed using acetonitrile while the chromatographic separation was accomplished on a RP 18 column (250 × 4.6 mm i.d., 5 µm), with a mobile phase composed of KH2PO4 10 mM-Acetonitrile (40:60, v/v) and UV detection was set at 333 nm. The method shown specificity compared to blank serum specimen with retention time of rifampicin at 2.1 min. Lower limit of quantification (LLOQ) was 0.06 µg/mL with dynamic range up to 20 µg/mL (R>0.990). Precision of the method was very good with coefficient of variance (CV) 0.58; 7.40 and 5.56% for concentration at 0.06, 5, 15 µg/mL, respectively. Accuracies of the method were 3.22; 1.94; 1.90% for concentration 0.06, 5 and 15 µg/mL respectively. The average recoveries were 97.82, 95.50 and 97.31% for concentration of rifampicin 1, 5 and 5 µg/mL, respectively. The method was also shown reliable result on stability test on freezing-thawing, short-term and long-term stability as well as post preparation stability. Validation result shown that the method was ready to be used for Rifampicin BE testing with Indonesian subject. Keywords: Rifampicin, Validation, USFDA-Guideline

A Decline in C6 Antibody Titer Occurs in Successfully Treated Patients with Culture-Confirmed Early Localized or Early Disseminated Lyme Borreliosis

ABSTRACT C6, a Borrelia burgdorferi-derived peptide, is used as the antigen in the C6-Lyme disease diagnostic test. We assessed retrospectively whether a fourfold decrease or a decrease to a negative value in anti-C6 antibody titer is positively correlated with a positive response to treatment in a sample of culture-confirmed patients with either early localized (single erythema migrans [EM]; n = 93) or early disseminated (multiple EM; n = 27) disease. All of these patients had been treated with antibiotics and were free of disease within 6 to 12 months of follow-up. Results show that a serum specimen taken at this time was either C6 negative or had a ≥4-fold decrease in C6 antibody titer with respect to a specimen taken at baseline (or during the early convalescent period if the baseline specimen was C6 negative) for all of the multiple-EM patients (P < 0.0001) and in 89% of the single-EM patients (P < 0.0001). These results indicate that a decline in anti-C6 antibody titer coincides with effective antimicrobial therapy in patients with early localized or early disseminated Lyme borreliosis.