The asymptomatic ocular surface reservoir of coronavirus disease-19 in infants and retinopathy of prematurity screening during the pandemic.

PURPOSE: This study aimed to evaluate the incidence of conjunctival and pharyngeal swab sample positivity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in asymptomatic preterm and term infants. We aimed to detect asymptomatic carrier potential in infants. MATERIALS AND METHODS: Patients screened for retinopathy of prematurity (ROP) at our clinic between January and June 2021 were recruited for this study. For all study cases, the previous history of possible exposure or contact with SARS-CoV-2, previous history of coronavirus disease-19 (COVID-19), or contact with any COVID-19 case was excluded. None of the patients showed signs of COVID-19 during sample collection. Pharyngeal and conjunctival swab samples were collected before the ophthalmic examination. Nucleic acid isolation from the samples was performed using an automated system. The presence of SARS-CoV-2 RNA in the samples was screened using a real-time polymerase chain reaction kit, and the positive samples were re-evaluated for the variant virus. RESULTS: Among the 127 patients with a median age of 40 weeks (range: 34-86) of postmenstrual age, positivity for SARS-CoV-2 RNA in the pharyngeal and conjunctival samples was 5/127 (3.93%) and 3/127 (2.36%), respectively. Isolated conjunctival positivity was not observed in any of the patients, and all three patients were positive for both conjunctiva and pharynx. CONCLUSION: Asymptomatic infants may be a reservoir for SARS-CoV-2, and conjunctival infection in infants may be a source of virus transmission. Since ROP screening cannot be postponed during the pandemic, caution should be exercised to prevent the spread of the disease.

[P323L Mutation in a Case with Prolonged SARS-CoV-2 PCR Positivity]. / Uzamis SARS-CoV-2 PCR Pozitifligi Olan Olguda P323L Mutasyonu.

Coronavirus disease-2019 (COVID-19) emerged in the last months of 2019 and caused a pandemic effecting the whole world. Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the causative agent of COVID-19 has changed by various mutations since the day it was first identified, causing the pandemic to continue. Age, male gender, obesity, and comorbidity, which are general risk factors for COVID-19, can also cause prolonged PCR positivity. In this report, a case of 37-year-old male who is working in the hospital's COVID-19 molecular diagnostics laboratory was presented. He was vaccinated with three doses of inactivated vaccine, CoronaVac (Sinovac Biotech, Beijing-China), within the context of the vaccination program carried out in Türkiye. His first SARS-CoV-2 positivity was detected on 12.01.2021, four months after the last vaccination, and he continued to be detected positive for SARS-CoV-2 throughout a period of 39 days by quantitative reverse transcription polymerase chain reaction (qRT-PCR) tests performed with 2-3-day intervals. The patient has a 20-pack/year smoking history and his body mass index (BMI) was 29.8 kg/m2 at the time of his COVID-19. The case, which was clinically defined as mild COVID-19 with symptoms including back and headache, cough, fever (38.5°C), and loss of taste-smell, and without any additional complications or respiratory distress during the disease process. In the radiological examination, the lung was found within normal ranges. Prophylactic enoxaparin sodium anti-xa IU/0.6 ml was administered to the patient due to his cardiovascular risk, and no additional treatment was given. Whole genome sequencing was performed from nasopharyngeal swab samples of the patient at the beginning and 16th day of the infection to investigate the the specific genomic features and mutation pattern of the virus in the host over time, due to the prolonged SARS-CoV-2 PCR positivity. Library preparation for the whole next-generation sequencing (NGS) was performed by the SARS-CoV-2 Panel, Paragon CleanPlex kit (Paragon Genomics, USA), and indexing of the library was done by Clean-Plex Dual-Indexed PCR Primers for Illumina Set B kit (Paragon Genomics, USA). NGS analysis was performed on the Illumina Miniseq (Illumina, USA) platform. As a result of the bioinformatics evaluation, both samples were determined as SARS-CoV-2 Delta variant (Nextclade; 21J-Delta variant, Pango lineage; AY.43). Remarkably, the SARSCoV-2 sequences in the two samples taken 15 days apart; several identical mutations; such as D614G in the S gene, P323L in the ORF 1b gene region, and P1228L in the Nsp3 gene region, were detected. Besides that, when compared to the first sample, three additional mutations (P383L, P539S, L838I) were observed in the sequence of the second sample, which led to three amino acid changes, the clinical significance of which has not yet been determined in the literature. It is thought that; these mutations that change amino acid expression, as well as the other three mutations detected, may contribute to the improvement of the fitness of the virus and may be one of the factors responsible for the prolonged SARS-CoV-2 PCR positivity. Additional data to be obtained by further epidemiological sequencing studies will shed light on this issue.

Evaluation of the Diagnostic Performance of a SARS-CoV-2 and Influenza A/B Combo Rapid Antigen Test in Respiratory Samples.

This study aimed to evaluate the performance characteristics of a rapid antigen test developed to detect SARS-CoV-2 (COVID-19), influenza A virus (IAV), and influenza B virus (IBV) (flu) compared with those of the real-time reverse transcription-polymerase chain reaction (rRT-PCR) method. One hundred SARS-CoV-2, one hundred IAV, and twenty-four IBV patients whose diagnoses were confirmed by clinical and laboratory methods were included in the patient group. Seventy-six patients, who were negative for all respiratory tract viruses, were included as the control group. The Panbio™ COVID-19/Flu A&B Rapid Panel test kit was used in the assays. The sensitivity values of the kit were 97.5%, 97.9%, and 33.33% for SARS-CoV-2, IAV, and IBV, respectively, in samples with a viral load below 20 Ct values. The sensitivity values of the kit were 16.7%, 36.5%, and 11.11% for SARS-CoV-2, IAV, and IBV, respectively, in samples with a viral load above 20 Ct. The kit's specificity was 100%. In conclusion, this kit demonstrated high sensitivity to SARS-CoV-2 and IAV for viral loads below 20 Ct values, but the sensitivity values were not compatible with PCR positivity for lower viral loads over 20 Ct values. Rapid antigen tests may be preferred as a routine screening tool in communal environments, especially in symptomatic individuals, when diagnosing SARS-CoV-2, IAV, and IBV with high caution.

One-Year Post-Vaccination Longitudinal Follow-Up of Quantitative SARS-CoV-2 Anti-Spike Total Antibodies in Health Care Professionals and Evaluation of Correlation with Surrogate Neutralization Test.

Numerous vaccines have been generated to decrease the morbidity and mortality of COVID-19. This study aims to evaluate the immunogenicity of the heterologous boosts by BioNTech against homologous boosts by CoronaVac at three-month intervals in two health care worker (HCW) cohorts, with or without prior COVID-19, for one year post-vaccination. This is a prospective cohort study in which the humoral responses of 386 HCWs were followed-up longitudinally in six main groups according to their previous COVID-19 exposure and vaccination status. Anti-SARS-CoV-2 spike-RBD total antibody levels were measured and SARS-CoV-2 neutralization antibody (NAbs) responses against the ancestral Wuhan and the Omicron variant were evaluated comparatively using international standard serum for Wuhan and Omicron, as well as with the aid of a conversion tool. The anti-SARS-CoV-2 spike-RBD total Ab and Nab difference between with and without prior COVID-19, three months after two-dose primary vaccination with CoronaVac, was statistically significant (p = 0.001). In the subsequent follow-ups, this difference was not observed between the groups. Those previously infected (PI) and non-previously infected (NPI) groups receiving BioNTech as the third dose had higher anti-SARS-CoV-2 spike total Ab levels (14.2-fold and 17.4-fold, respectively, p = 0.001) and Nab responses (against Wuhan and Omicron) than those receiving CoronaVac. Ab responses after booster vaccination decreased significantly in all groups at the ninth-month follow-up (p < 0.05); however, Abs were still higher in all booster received groups than that in the primary vaccination. Abs were above the protective level at the twelfth-month measurement in the entire of the second BioNTech received group as the fourth dose of vaccination. In the one-year follow-up period, the increased incidence of COVID-19 in the groups vaccinated with two or three doses of CoronaVac compared with the groups vaccinated with BioNTech as a booster suggested that continuing the heterologous CoronaVac/BioNTech vaccination, revised according to current SARS-CoV-2 variants and with at least a six-month interval booster would be an effective and safe strategy for protection against COVID-19, particularly in health care workers.

Association of SARS-CoV-2 IgG Antibody Levels with Clinical/Laboratory Characteristics in COVID-19 Patients: Data from a Turkish Study.

BACKGROUND: COVID-19 causes clinical manifestations ranging from asymptomatic infection to multi-organ failure. It is reported that those with severe disease have higher anti-SARS-CoV-2 antibody titers compared to asymptomatic or mild cases. We evaluated the correlation of antibody responses with laboratory and clinical indicators in COVID-19 patients. METHODS: Seventy-nine male and 66 female patients (mean age: 39) with at least one positive SARS-CoV-2 RT-PCR test and SARS-CoV-2 IgG antibody result after acute infection were included. RESULTS: Seventy-six (52%), 45 (31%), and 24 (17%) patients had mild, moderate, and severe clinical findings, respectively. Patients with high body mass index and advanced age had significantly more severe disease (p < 0.001). A significant correlation was found between the increase in lymphopenia, C-reactive protein, ferritin, D-dimer, and lactate dehydrogenase and the severity of clinical findings (p = 0.0001). SARS-CoV-2 IgG antibody test was positive in 128 (88.3%) patients. A significant correlation was found between disease severity and antibody levels in the comparison of all groups (p < 0.001). CONCLUSIONS: Long-term monitoring of immune responses will be required to determine the appropriate time for the administration of new vaccines.

Higher antibody responses after mRNA-based vaccine compared to inactivated vaccine against SARS-CoV-2 in Behcet's syndrome.

There are limited data about humoral response to vaccine in Behçet's syndrome (BS). We compared SARS-CoV-2 antibody response after two doses of inactivated (Sinovac/CoronaVac) or mRNA (Pfizer/BioNTech) vaccines in patients with BS and healthy controls (HCs). We studied 166 (92M/74F) patients with BS (mean age: 42.9 ± 9.6 years) and 165 (75M/90F) healthy controls (mean age: 42.4 ± 10.4 years), in a single-center cross-sectional design between April 2021 and October 2021. A total of 80 patients with BS and 89 HCs received two doses of CoronaVac, while 86 patients with BS and 76 HCs were vaccinated with BioNTech. All study subjects had a negative history for COVID-19. Serum samples were collected at least 21 days after the second dose of the vaccine. Anti-spike IgG antibody titers were measured quantitatively using a commercially available immunoassay method. We found that the great majority in both patient and HC groups had detectable antibodies after either CoronaVac (96.3% vs 100%) or BioNTech (98.8% vs 100%). Among those vaccinated with CoronaVac, BS patients had significantly lower median (IQR) titers compared to HCs [36.5 (12.5-128.5) vs 102 (59-180), p < 0.001]. On the other hand, antibody titers did not differ among patients with BS and HCs who were vaccinated with BioNTech [1648.5 (527.0-3693.8) vs 1516.0 (836.3-2599.5), p = 0.512). Among different treatment regimen subgroups in both vaccine groups, those who were using anti-TNF-based treatment had the lowest antibody titers. However, the difference was statistically significant only among those vaccinated with CoronaVac. Among patients vaccinated with BioNTech, there was no statistically significant difference between different treatment regimen groups. Compared to inactivated COVID-19 vaccine, mRNA-based vaccine elicited higher antibody titers among BS patients. Only in the CoronaVac group, patients especially those using anti-TNF agents were found to have low titers compared to healthy subjects. BS patients vaccinated with BioNTech were found to have similar seroconversion rates and antibody levels compared to healthy controls. Further studies should assess whether the low antibody titers are associated with diminished protection against COVID-19 in both vaccine groups.

Development of novel spectroscopic and machine learning methods for the measurement of periodic changes in COVID-19 antibody level.

In this research, blood samples of 47 patients infected by COVID were analyzed. The samples were taken on the 1st, 3rd and 6th month after the detection of COVID infection. Total antibody levels were measured against the SARS-CoV-2 N antigen and surrogate virus neutralization by serological methods. To differentiate COVID patients with different antibody levels, Fourier Transform InfraRed (FTIR) and Raman spectroscopy methods were used. The spectroscopy data were analyzed by multivariate analysis, machine learning and neural network methods. It was shown, that analysis of serum using the above-mentioned spectroscopy methods allows to differentiate antibody levels between 1 and 6 months via spectral biomarkers of amides II and I. Moreover, multivariate analysis showed, that using Raman spectroscopy in the range between 1317 cm-1 and 1432 cm-1, 2840 cm-1 and 2956 cm-1 it is possible to distinguish patients after 1, 3, and 6 months from COVID with a sensitivity close to 100%.

Comparison of the Clinical and Laboratory Findings and Outcomes of Hospitalized COVID-19 Patients Who Were Either Fully Vaccinated with Coronavac or Not: An Analytical, Cross Sectional Study.

COVID-19 vaccines are highly protective against severe disease; however, vaccine breakthrough infections resulting in hospitalization may still occur in a small percentage of vaccinated individuals. We investigated whether the clinical and microbiological features and outcomes were different between hospitalized COVID-19 patients who were either fully vaccinated with Coronovac or not. All hospitalized COVID-19 patients who had at least one dose of Coronavac were included in the study. The oldest unvaccinated patients with comorbidities, who were hospitalized during the same period, were chosen as controls. All epidemiologic, clinical and laboratory data of the patients were recorded and compared between the fully vaccinated and unvaccinated individuals. There were 69 and 217 patients who had been either fully vaccinated with Coronavac or not, respectively. All breakthrough infections occurred in the first 3 months of vaccination. Fully vaccinated patients were older and had more comorbidities than unvaccinated patients. There were minor differences between the groups in symptoms, physical and laboratory findings, anti-spike IgG positivity rate and level, the severity of COVID-19, complications, and clinical improvement rate. The mortality rate of fully vaccinated patients was higher than the mortality rate in unvaccinated patients in univariate analysis, which was attributed to the fact that vaccinated patients were older and had more comorbidities. The severity and clinical outcomes of hospitalized patients with breakthrough COVID-19 after Coronavac vaccination were similar to those of unvaccinated patients. Our findings suggest that the immune response elicited by Coronovac could be insufficient to prevent COVID-19-related severe disease and death within 3 months of vaccination among elderly people with comorbidities.

Anti-SARS-CoV-2 IgG and Neutralizing Antibody Levels in Patients with Past COVID-19 Infection: A Longitudinal Study

Background: Monitoring the longevity of immunoglobulin G (IgG) responses following severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infections is vital to understanding the role of antibodies in preventing infection. Aims: To determine the quantitative IgG responses specific to the Spike-S1 (S1) receptor-binding domain (S1/RBD) region of the virus in serum samples taken between 4 weeks and 7 months after polymerase chain reaction (PCR) positivity in patients who are diagnosed with coronavirus disease-2019 (COVID-19). Study Design: A longitudinal study. Methods: This study included 113 patients with a clinical and molecular diagnosis of COVID-19. The first and second serum samples were taken 1 and 7 months, respectively, after the PCR positivity. S1/RBD-specific IgG antibody response was assayed using anti-SARS-CoV- 2 QuantiVac ELISA (IgG) kit (Euroimmun, Lübeck, Germany). The neutralizing antibodies were investigated in 57 patients whose IgG test results were above the cut-off value. Results: In 57 patients with SARS-CoV-2 IgG, the anti-SARS-CoV-2 IgG quantitative antibody levels significantly decreased after 7 months (Z = −2.197, p = 0.028). A correlation was detected between the anti-SARS-CoV-2 IgG and nAb percent inhibition (IH%) levels detected in 1 month (rs = 0.496, p < 0.001), but without significant correlation in serum samples taken on 7 months. The nAb IH% levels of the first and second were compared for COVID-19 severity and revealed no statistical difference (p = 0.256). In the second serum sample, the nAb IH%s of patients with moderate COVID-19 showed a statistically significant difference from patients with mild COVID-19 (p = 0.018), but without significant differences between severe and moderate or mild COVID-19. Conclusion: SARS-CoV-2 quantitative IgG antibody titers are significantly reduced at long-term follow-up (> 6 months). Due to the limited information on seroconversion, comprehensive studies should be conducted for long-term follow-up of the immune response against SARS-CoV-2.

Effect of BTN162b2 and CoronaVac boosters on humoral and cellular immunity of individuals previously fully vaccinated with CoronaVac against SARS-CoV-2: A longitudinal study.

BACKGROUND: It is essential to know about immune response levels after booster doses of the two different types of vaccines, mRNA, and the inactivated, currently used against COVID-19. For this purpose, we aimed to determine the effects of BNT162b2 (BNT) and CoronaVac (CV) boosters on the humoral and cellular immunity of individuals who had two doses of CV vaccination. METHODS: The study was conducted in three centers (Koc University Hospital, Istanbul University Cerrahpasa Hospital, and Istanbul University, Istanbul Medical School Hospital) in Istanbul, Turkey. Individuals who had been previously immunized with two doses of CV and no history of COVID-19 were included. The baseline blood samples were collected 3-5 months after the second dose of CV. Follow-up blood samples were taken 1 and 3 months after administration of third doses of CV, or one dose of BNT boosters. Neutralizing antibody titers were measured by plaque reduction assay. The CD4+ T cell, CD8+ T cell, effector CD4+CD38+CD69+ T cell, and effector CD8+CD38+CD69+ T cell ratios were determined by flow cytometry. The intracellular IFN-γ and IL-2 responses were measured by ELISpot assay. RESULTS: We found a 3.38-fold increase in neutralizing antibody geometric mean titers (NA GMT, 78.69) 1 month after BNT booster and maintained at the third month (NA GMT, 80). Nevertheless, in the CV booster group, significantly lower NA GMT than BNT after 1 month and 3 months were observed (21.44 and 28.44, respectively) (p < .001). In the ELISpot assay, IL-2 levels after BNT were higher than baseline and CV booster (p < .001) while IFN-γ levels were significantly higher than baseline (p < .001). The CD8+CD38+CD69+ and CD4+CD38+CD69+ T cells were stimulated predominantly in the third month of the BNT boosters. CONCLUSION: The neutralizing antibody levels after 3 months of the BNT booster were higher than the antibody levels after CV in fully vaccinated individuals. On the contrary, ratio of the effector T cells increased along with greater IFN-γ activation after BNT booster. By considering the waning immunity, we suggest a new booster dose with BNT for the countries that already had two doses of primary CV regimens.

[Evaluation of the Diagnostic Performance of SARS-CoV-2 Rapid Antigen Tests in COVID-19 Patients]. / SARS-CoV-2 Hizli Antijen Testlerinin COVID-19 Hastalarindaki Tanisal Performanslarinin Degerlendirilmesi.

The gold standard in the definitive diagnosis of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is nucleic acid amplification tests (NAAT) due to their high sensitivity and specificity in detecting viral ribonucleic acid. However, while leaving two years behind in the pandemic, resources have come to the point of exhaustion in terms of both the economy and the manpower working in the field of health services. Therefore, the need for rapid, simple and accurate tests to diagnose SARS-CoV-2 infection continues. In this study, it was aimed to compare the performance characteristics of SARS-CoV-2 rapid antigen tests (RAgT) in the diagnosis of coronavirus disease 2019 (COVID-19) cases with the real-time reverse transcription-polymerase chain reaction (rRT-PCR) method. In Istanbul University-Cerrahpasa Faculty of Medicine COVID-19 Molecular Diagnosis Laboratory, SARS-CoV-2 RNA positive respiratory tract samples with viral loads of <25 Ct (cycle of treshold), 25-29 Ct, 30-35 Ct and 35

A pan-resistant Myroides odoratimimus catheter-related bacteremia in a COVID-19 patient and review of the literature.

Myroides spp. are opportunistic environmental Gram-negative bacteria. These affect mostly immunocompromised hosts and generally lead to soft tissue, and urinary tract infections. Bacteremia most commonly develop secondary to soft tissue or catheter related infections and may lead rarely to mortality. Myroides spp. are generally suscetible to fluoroquinolones, piperacillin/tazobactam, trimethoprim/sulfamethoxazole, carbapenems or tetracyclines however, pan-resistant isolates and multiple resistance genes have been reported in clinical isolates of Myroides spp. We report a pan-resistant Myroides odoratimimus bacteremia in a patient with severe COVID-19 ending with fatality and in this context a review of reported Myroides bacteremias are also described. In this study, a 64-year old male patient with history of coronary artery bypass was admitted to ICU with severe COVID-19 pneumonia accompanied by pneumomediastinum and pneumopericardium. Continous renal replacement therapy and extracorporeal membraneous-oxygenation were initiated due to acute renal failure and persistent hypercarbia/hypoxia, respectively. Within four weeks of hospitalization various episodes of bacteremia developed and multiple antibiotics were used. On the 5th week of follow-up, acute phase reactants increased and empirical broad spectrum antibiotics were initiated. Blood culture revealed Gram-negative rods. The patient became hypotensive and despite maximum medical care he was lost due to cardiac arrest. M. odoratimimus was identified by MALDI-TOF and the bacterium was pan-resistant. According to Center for Genomic Epidemiology results the strain was identified as M. odoratimimus PR63039 and the genome analysis revealed antibiotic resistance genes associated with resistance to beta-lactams (bla OXA-347, bla MUS-1, bla EBR-1), tetracyclines (tetX), sulfonamides (sul2), macrolides (ereD), (ermF).

Inactive SARS-CoV-2 vaccine generates high antibody responses in healthcare workers with and without prior infection.

BACKGROUND AND OBJECTIVES: Healthcare workers (HCWs) were among the first groups to be vaccinated in Turkey. The data to be obtained by the vaccination of HCWs would guide wide spread vaccination programs. MATERIALS AND METHODS: The study included 330 HCWs working at Istanbul University-Cerrahpasa, Cerrahpasa Medical Faculty Hospital and vaccinated with inactive CoronaVac (Sinovac Life Sciences, China) SARS-CoV-2 vaccine in two doses (28 days apart). Anti-Spike /RBD IgG levels were measured 14 days after the first dose and 28 days after the second dose. Chemiluminescent microparticle immunoassay (CMIA) (ARCHITECT IgG II Quant test, Abbott, USA), which is 100% compatible with plaque reduction neutralization test (PRNT), was used. RESULTS: Of the participants, 211 (63.9%) were female, 119 (36.1%) were male, and mean age was 39.6 ± 7.7 years. In those without prior COVID-19 history; (n = 255) antibody positivity was detected as 48.2% (95% CI: 42.1-54.3) 14 days after the first dose of vaccine, and 99.2% (95% CI: 98.1-100) at day 28 after the second dose. Antibody titers were significantly lower in patients with hypertension (p = 0.011). In those with prior history of COVID-19 (n = 75); both the antibody positivity rates after the first vaccine (48.2% vs 100%, p = 0.000) and the anti-spike/RBD antibody levels after the second vaccine (with a ≥ 1050 AU/mL titer equivalent to PRNT 1/80 dilution) was significant than infection-naive group (25.9% vs. 54.7%, p = 0.000). Antibody positivity after two doses of vaccination for all study group was 99.4% (95% CI: 98.6-100). CONCLUSIONS: Two doses CoronaVac produce effective humoral immunity in HCWs. Antibody response is significantly higher in those with prior history of COVID-19 than infection-naive group. Given no significant benefit of the second dose, a single shot of vaccination may be sufficient for those with prior history of COVID-19. Monitoring humoral and cellular immune responses, considering new variants, is required to validate this approach.

Detection of SARS-CoV-2 RNA in Upper Respiratory Swap Samples by Pooling Method

Background: Widespread and effective use of molecular diagnostic tests is indispensable for protecting public health and containing the severe respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. More than 1 year into the pandemic, as resources have reached a point of depletion, grouping samples in pools of certain sizes appears to be a reasonable method to reduce both the costs and the processing time without necessitating additional training, equipment, or materials. Aims: To assess whether the pooling strategy that was used in past outbreaks and is used in blood tests prior to transfusion for screening large populations can also be used in SARS CoV-2 tests. Study Design: Diagnostic accuracy study. Methods: This prospective study was conducted with 2815 samples, sent to the coronavirus disease 2019 (COVID-19) Laboratory of our hospital between February 12 and 21, 2021, to be tested for the presence of SARS-CoV-2. The samples were examined individually and in pools of five 100 µl taken from each sequential sample, using 3 different SARS-CoV-2 reverse transcription-polymerase chain reaction (RT-PCR) kits, the Allplex™ 2019-nCoV Assay kit (Seegene, Republic of Korea), the GeneMAP™ 2019-nCoV detection V.3 kit (GenMark, Türkiye), and the Bio-Speedy™ SARS-CoV-2 Double Gene™ RT-qPCR kit (Bioeksen, Türkiye) on the BioRAD CFX96™ Touch (Bio-Rad Laboratories Inc., Hercules, CA, USA) platform available in our laboratory. Results: Following the extraction of serial dilutions prepared from the SARS-CoV-2 RNA positive (cycle of threshold: 20) sample, the standard curves of RT-PCR were analyzed. By evaluating the efficiency (E) values, all 3 kits showed high sensitivity and similar results; while the highest level was detected with the Allplex™ 2019-nCoV Assay kit in the nucleocapsid (N) gene (E: 124%), the lowest was detected with the Double Gene™ RT-qPCR kit in the N and ORF 1ab genes (E: 90%). Of the samples included in the study, only 1 positive sample with low viral load was found to be negative when studied by pooling. The total number of kits to be used in pooled tests and then to individually retest the 5 samples in positive pools was calculated as 827 and the savings rate as 69.91% (1968/2815). Conclusion: The pooling strategy is an effective approach to extend the impact of limited testing resources and reagents available in certain periods of the COVID-19 pandemic. Testing by pooling samples requires improvement of RNA extraction methods and careful monitoring of RT-PCR test sensitivity to avoid missing low-positive entities. Therefore, based on the prevalence of COVID-19 in their regions, laboratories should conduct their own validation of pooling studies for RNA extraction and amplification methods they use.

Real-life experience: sensitivity and specificity of nasal and saliva samples for COVID-19 diagnosis.

BACKGROUND: COVID-19 (coronavirus disease 2019) outbreak has spread rapidly around the world, continues to show its effect, and it is not clear how long it will continue. For the diagnosis of COVID-19, it is important to ensure the comfort of the patients and to protect the healthcare workers (HCWs) by reducing the use of protective equipment. AIMS: To evaluate or assess whether the samples taken by the patient for COVID-19 testing during this pandemic period can be used in real-life experience. METHODS: Three different samples (nasopharyngeal taken by the healthcare worker, nasopharyngeal, and saliva taken by the patient) from 132 patients were evaluated for the diagnosis of COVID-19. The sensitivity and specificity of the samples in the diagnosis of COVID-19 were compared with real-life experience. RESULTS: Paired analyzes were performed by comparing each sample taken by the healthcare worker with the sample taken by the patient. The sensitivity of the three samples (nasopharyngeal taken by the healthcare worker, nasopharyngeal, and saliva taken by the patient) in the diagnosis of the COVID-19 was (100%, 98.7%, and 96.1%, respectively) accepted to be accurate. CONCLUSIONS: The sample taken by the paramedic was compatible compared to the real-life experience for the samples taken by the patient in the COVID-19 pandemic period. During the pandemic that is unknown when it will end, this study demonstrated that taking the sample of the patient alone for the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) test is a beneficial approach to the protection of the healthcare worker, reducing the need for protective equipment, increasing the patient's comfort and rapid sampling.

Novel SARS-CoV-2 Omicron variants in Istanbul; Rapid Preponderance of BA.2 and BA.5.

Objective: In Turkey, the fourth wave of SARS-CoV-2 started in December 2021 and peaked in mid-January 2022. Afterward, peaks were seen in the number of COVID-19 cases because of Omicron BA.2 and BA.5 variants. Our study aimed to observe the prevalence and viral load-related transmissibility rates of the Omicron BA.2 and BA.5 variant infections in our region between January 21 and July 01, 2022, using an easy and cost-effective PCR screening method. Methods: The frequency of BA.2 and BA.5 were determined by the two-stage allele-specific and drop-out RT-PCR method targeting NSP6 105-107del, spike 69-70del, and spike L452R mutation-specific primers. Transmissibility of the Omicron variants was assessed using cycle threshold (Ct) values (a proxy for SARS-CoV-2 viral load and infectivity). Also, using the next generation sequencing (NGS) method, existing mutations were analyzed by generating full-length sequences of the representative, randomly selected samples from the Omicron variants determined by PCR screening test. Results: We defined the first case of BA.2 on January 19, 2022, in Istanbul University-Cerrahpasa School of Medicine COVID-19 Molecular Diagnosis Laboratory. Following this, it was observed that BA.1 lost its dominance due to the increased transmissibility of BA.2. On May 5, we defined the first case of BA.5, and as of July this Omicron variant rapidly became preponderant, with a frequency of more than 85%. Compared with BA.1, BA.2 and BA.5 were associated with 2.82 (95% CI: 2.33-4.12) and 2.49 (95% CI: 2.16-3.55) fewer cycles, respectively, meaning higher transmissibility. As confirmed by the NGS results, it was concluded that screening with NSP6 105-107del, spike 69-70del and spike L452R mutation targeted PCR method, which is used uniquely in our hospital in Turkey, can be an easy and cost-effective method in the follow-up of Omicron variants. Conclusion: The higher viral load detection in infections with BA.2 and BA.5 reflects a prolonged disease period, and increased transmissibility, so rapid expansion of these Omicron variants in Turkey is inevitable. Even though the prevalence of the Omicron variants in the population can be monitored in near real-time by the PCR screening method, more sequencing studies are needed for the early identification of new mutations that will emerge.

Antibody response to inactivated COVID-19 vaccine (CoronaVac) in immune-mediated diseases: a controlled study among hospital workers and elderly.

OBJECTIVE: To assess antibody response to inactivated COVID-19 vaccine in patients with immune-mediated diseases (IMD) among hospital workers and people aged 65 and older. METHODS: In this cross-sectional study, we studied 82 hospital workers with IMD (mean age: 42.2 ± 10.0 years) and 300 (mean age: 41.7 ± 9.9 years) controls. Among + 65 aged population, we studied 22 (mean age: 71.4 ± 4.5 years) patients and 47 controls (mean age: 70.9 ± 4.8 years). All study subjects had a negative history for COVID-19. Sera were obtained after at least 21 days following the second vaccination. Anti-spike IgG antibody titers were measured quantitatively using a commercially available immunoassay method. RESULTS: Patients with IMD were significantly less likely to have detectable antibodies than healthy controls both among the hospital workers (92.7% vs 99.7%, p < 0.001) and elderly population (77.3% vs 97.9%, p = 0.011). Among patients with IMD, those using immunosuppressive or immune-modulating drugs (64/75, 85.3%) were significantly less likely to have detectable antibodies compared to those off treatment (29/29, 100%) (p = 0.029). Additionally, a negative association between age and the antibody titer categories among patients (r = - 0.352; p < 0.001) and controls (r = - 0.258; p < 0.001) were demonstrated. CONCLUSIONS: Among hospital workers, the vast majority of patients with IMD and immunocompetent controls developed a significant humoral response following the administration of the second dose of inactivated COVID-19 vaccine. This was also true for the elderly population, albeit with lower antibody titers. Immunosuppressive use, particularly rituximab significantly reduced antibody titers. Antibody titers were significantly lower among those aged ≥ 60 years both in patient and control populations. Whether these individuals should get a booster dose warrants further studies.

[Evaluation of the Diagnostic Performance of Different Principles of SARS-CoV-2 Commercial Antibody Tests in COVID-19 Patients]. / SARS-CoV-2 ile Ilgili Farkli Prensipli Ticari Antikor Testlerinin COVID-19 Hastalarindaki Tanisal Performanslarinin Degerlendirilmesi.

Following the emergence of severe acute respiratory syndrome coronavirus-2 (SARS CoV-2) and using only PCR for diagnosis, antibody tests have been rapidly developed by various commercial companies. There are differences between the sensitivity and specificity of these tests due to the usage of different viral target proteins and antibody subclasses. In order to evaluate the diagnostic use of these tests, we aimed to examine the diagnostic performance, especially sensitivity and specificity, of SARS-CoV-2 IgM, IgA and IgG tests of various companies (Abbott, Roche, Euroimmun, Dia.Pro, Anshlabs, Vircell, UnScience and RedCell), which have different principles (ECLIA/CLIA, EIA, LFA). Current (n= 180) and past (n= 180) COVID-19 patients with clinical and molecular diagnosis of COVID-19 admitted to Istanbul University-Cerrahpasa, Cerrahpasa Faculty of Medicine Hospital, Pandemic Polyclinic with suspected COVID-19 infection, were included in our study. The patients admitted within the first 3 weeks after the onset of symptoms were included in the current patient group, and those admitted at the third and after the third week were included in the past patient group. Serum samples (n= 180) obtained from Istanbul Sisli Hamidiye Etfal Training and Research Hospital, Blood Center between April and June 2018 before the COVID-19 pandemic were included in the study as a control group. All the tests included in our study were studied with the recommendations of the manufacturer companies. Between the IgG detection tests with different principles in patients with past COVID-19, the sensitivity and specificity values of the most effective tests were; 86.7%/99.4% (Abbott), 86.1%/98.9% (Dia.Pro), 91.3%/95% (RedCell). Between the IgM detection tests with different principles in current COVID-19 patients, the sensitivity and specificity values were; 67.8%/99.4% (Abbott), 68.9%/98.6% (Vircell), 50%/97.5% (RedCell). Abbott IgM with a kappa coefficient of 0.67 and Vircell IgM + IgA test with a kappa coefficient of 0.65 showed the best fit in patients with current COVID-19 infection. In patients with past COVID-19, Abbott IgG with 0.86 kappa coefficient and Dia.Pro IgG test with 0.85 kappa coefficient showed the best match. Due to the low sensitivity of IgM detection antibody tests, they should not be preferred instead of real-time reverse transcriptase polymerase chain reaction in routine diagnosis. IgG detection tests may be preferred to detect the antibody response and the titers in people who have had COVID-19 for population seroprevalence and especially therapeutic immune plasma production. However, it is thought that the combined use of both ECLIA/CLIA-based and EIA/ELISA-based tests together may be more effective in routine use for SARS-CoV-2 IgG tests.

[Investigation of Regulatory T Cells and Secreted Immunomodulatory Cytokine IL-10 Levels in Patients with Hepatitis B]. / Hepatit B Hastalarinda Regülatör T Hücre ve Salinan Immünmodülatör Sitokin IL-10 Düzeylerinin Degerlendirilmesi.

Hepatitis B infection is still among the most important public health problems worldwide, even great improvements have been made in the treatment strategies. Hepatitis B virus (HBV) replicates itself by entering the liver cells and simultaneously with the antigen release, many antagonistic immune responses are induced by the regulatory cells including T cell (Treg), T helper 17 (Th17), T helper 1 (Th1) and T helper 2 (Th2) cells. The main function of Treg cells is to develop an appropriate immune response against infection and to suppress the immune response if it is not required. Tregs suppress the effector T cells via secreting immune system supressor cytokines such as Transforming Growth Factor-Beta and interleukin (IL)-10 or contact dependent way. Tregs protect cells from immunopathologic damage of HBV specific T cell immune response and also cause viral persistence, cirrhosis, hepatocellular carsinoma (HCC) and autoimmunity but the mechanisms are not clear, yet. In this study, we aimed to determine whether evaluation of Treg cells and cytokine IL-10 levels together in hepatitis B patients is useful that may indicate the disease survey and response to the treatment. The peripheral blood samples of ninety-one volunteers, including 61 HBV infected patients and 30 healthy controls selected from applicants of Infectious Diseases Outpatient/Clinic Service, were taken. Their CD4+CD25highFOXP3+CD152+CD127lowTreg cell distribution were measured by flow cytometry method, using the recently defined markers. The level of IL-10 cytokine released by immunomodulatory cells was determined by quantitative ELISA method. Treg cell percentages of the patients with acute hepatitis B were below the normal range (2-4%) (median= 1.50%, 0.6-3.5) and the difference was statistically significant (p= 0.005). Treg cell percentages of the patients with chronic hepatitis B were higher than the control group (p< 0.05), and it was found to be related to the parameters used in the diagnosis, staging and follow-up of the disease. IL-10 levels were significantly higher in all hepatitis B clinical stages compared to the healthy controls (median= 11.7, 17.3-44.9) (p< 0.05). Also, in parallel with Treg cells, IL-10 levels were correlated with HBV DNA load and HBsAg levels (r= 0.48, p< 0.02). Treg cells and the related cytokine IL-10 are thought to play an important role in the immunology of HBV infection and therefore, promising to follow up the disease and to develop new therapeutic strategies targeting the Treg cell.