SARS-CoV-2 is not detectable in cervicovaginal secretions from women with active COVID-19 infection- a pilot study.

OBJECTIVE: To detect the shedding of SARS-CoV-2 in cervicovaginal secretions of women with active COVID 19 infection. METHODS: A cross-sectional study from a COVID facility including women aged 20-45 years with active COVID-19 infection, cervicovaginal secretions were collected from cervix and posterior fornix using dacron swab within 7 days of symptom onset or 5 days of nasopharyngeal rRT-PCR test positivity in asymptomatic women. Cervicovaginal samples of women with mild symptoms were tested using rRT-PCR for SARS-CoV-2. RESULTS: SARS-CoV-2 was not detected in cervicovaginal secretions of any of the 11 women included in the study. CONCLUSIONS: SARS-CoV-2 does not shed in the cervicovaginal secretions of women with mild COVID 19 infection, ruling out sexual and vertical transmission of virus in mild and asymptomatic disease.

Gargle lavage & saliva: Feasible & cheaper alternatives to nasal & throat swabs for diagnosis of COVID-19.

Background & objectives: In the present scenario, the most common sample for diagnosis of COVID-19 by reverse transcription polymerase chain reaction (RT-PCR) is nasal and throat swab (NTS). Other sampling options such as gargle lavage have found limited application in clinical use mostly because of unavailability of an appropriate gargling liquid. This study was conducted to assess the stability of SARS-CoV-2 RNA in normal saline at 4°C that can serve as a gargling liquid as well as a transport medium. The study also looked at the agreement between NTS and gargle lavage/saliva for the detection of SARS-CoV-2. Methods: In 29 consecutive real-time RT-PCR (rRT-PCR) positive COVID-19 patients, paired NTS, gargle and saliva samples were taken. Samples were processed by rRT-PCR for the detection of SARS-CoV-2 RNA. To assess the SARS-CoV-2 RNA stability in normal saline, gargle lavage specimens were divided into two aliquots; one subset of the specimen was run within 4-6 h along with the routine samples (NTS and saliva) and the other subset was stored at 4°C and processed after 24-30 h. Agreement between cycle threshold (Ct) values from both the runs was compared using Bland-Altman (BA) analysis. Results: The positivity rates of rRT-PCR in NTS, saliva and gargle lavage samples were 82.7 (24/29), 79.3 (23/29) and 86.2 per cent (25/29), respectively. BA plot showed a good agreement between the Ct values of fresh and stored gargle samples, stipulating that there were no significant differences in the approximate viral load levels between the fresh and stored gargle lavage samples (bias: E gene -0.64, N gene -0.51, ORF gene -0.19). Interpretation & conclusions: Our study results show stability of SARS-CoV-2 RNA in the gargle samples collected using normal saline up to 24-30 h. Gargle lavage and saliva specimen collection are cost-effective and acceptable methods of sampling for the detection of SARS-CoV-2 RNA by rRT-PCR. These simplified, inexpensive and acceptable methods of specimen collection would reduce the cost and workload on healthcare workers for sample collection.

Rapid chromatographic immunoassay-based evaluation of COVID-19: A cross-sectional, diagnostic test accuracy study & its implications for COVID-19 management in India.

Background & objectives: Coronavirus disease 2019 (COVID-19) has so far affected over 41 million people globally. The limited supply of real-time reverse transcription-polymerase chain reaction (rRT-PCR) kits and reagents has made meeting the rising demand for increased testing incompetent, worldwide. A highly sensitive and specific antigen-based rapid diagnostic test (RDT) is the need of the hour. The objective of this study was to evaluate the performance of a rapid chromatographic immunoassay-based test (index test) compared with a clinical reference standard (rRT-PCR). Methods: A cross-sectional, single-blinded study was conducted at a tertiary care teaching hospital in north India. Paired samples were taken for RDT and rRT-PCR (reference standard) from consecutive participants screened for COVID-19 to calculate the sensitivity and specificity of the RDT. Further subgroup analysis was done based on the duration of illness and cycle threshold values. Cohen's kappa coefficient was used to measure the level of agreement between the two tests. Results: Of the 330 participants, 77 were rRT-PCR positive for SARS-CoV-2. Sixty four of these patients also tested positive for SARS-CoV-2 by RDT. The overall sensitivity and specificity were 81.8 and 99.6 per cent, respectively. The sensitivity of RDT was higher (85.9%) in participants with a duration of illness ≤5 days. Interpretation & conclusions: With an excellent specificity and moderate sensitivity, this RDT may be used to rule in COVID-19 in patients with a duration of illness ≤5 days. Large-scale testing based on this RDT across the country would result in quick detection, isolation and treatment of COVID-19 patients.

Evaluation of Polymerase Chain Reaction over Routine Microbial Diagnosis for the Diagnosis of Fungal Keratitis.

SIGNIFICANCE: The significance of the study is that, although conventional culture remains the criterion standard for identifying the causative fungal pathogens, polymerase chain reaction (PCR) may serve as a powerful and high-throughput tool for the early and definitive diagnosis of high-risk patients with mycotic keratitis owing to high sensitivity and specificity. PURPOSE: This study was focused on comparing the results of PCR with traditional microbial studies for the detection and identification of fungal pathogens in patients with clinically suspected fungal keratitis. METHODS: Corneal scrapings were collected from 59 patients with clinically suspected fungal keratitis for routine culture, staining, and seminested PCR assay for fungal pathogen identification. The results of PCR were compared with a conventional microbial workup (smear and culture). The samples that were unidentified by culture but were amplified by PCR were further identified by nucleotide sequencing. RESULTS: Of the 59 patients with suspected fungal keratitis, 38 (64.40%) were found to be positive by PCR assay, 24 (40.67%) by culture, 18 (20.3%) by potassium hydroxide wet mount, and 8 (13.5%) by Gram stains for fungal pathogens. All the 24 isolates found positive with culture were also positive with PCR, so they had not been sequenced for molecular identification. The remaining 14 isolates that were positive with PCR but negative with culture were further identified as Cladosporium cladosporioides, Simplicillium species, Fusarium solani, Alternaria tenuissima, Chaetomium globosum, Penicillium citrinum, and Rhizopus microsporus by sequencing up to the species level. CONCLUSIONS: The PCR was able to detect the presence of fungal pathogens in a high proportion of culture-negative cases. This study suggests that PCR may serve as a rapid, important complement to traditional culture with high-throughput means of fungal pathogen identification in patients with clinically suspected fungal keratitis.

Topical lignocaine anaesthesia for oropharyngeal sampling for COVID-19.

OBJECTIVES: To ascertain if topical lignocaine application in oropharynx prior to swab sampling to test for COVID-19 improves a patient's comfort and to assess its effect on the swab sample taken to conduct the RT-PCR. METHODS: Adult patients testing positive on the RT-PCR COVID-19 test were sampled again within 48 h after administering topical oropharyngeal anaesthesia. Patients were asked to rate their discomfort on a visual analog scale (VAS) for both sample A and B. A qualitative real-time RT-PCR for detection of SARS-CoV-2 RNA, was performed, and the cycle threshold value (Ct), used as a surrogate marker for the viral load, was measured for the sample taken without lignocaine (sample A) and the sample taken post-lignocaine application (sample B). The difference in Ct values of both the groups was checked for any statistical significance using paired t-test. Wilcoxon signed rank test was used on VAS scores to determine any significant decrease in discomfort. RESULTS: Forty patients were included in the study. Twenty-nine patients (72.5%) reported the procedure to be more comfortable post-lignocaine application. Median (IQR) discomfort on VAS decreased from 7 (1) to 5 (2) after lignocaine use, which was statistically significant (p < 0.05). Mean Ct value for sample A was 17.21 ± 5.25 and for sample B was 18.44 ± 4.8 (p > 0.05), indicating a non-significant effect of lignocaine on SARS-CoV-2 concentration in the sample. CONCLUSION: Topical lignocaine, while improving the comfort of the procedure of oropharyngeal sampling for patient did not alter the SARS-CoV-2 viral load that was detected in nasal and oropharyngeal samples taken together.

Chronic osteomyelitis by Aeromonas hydrophila: A silent cause of concern.

Aeromonas is a Gram-negative bacillus, widely found in aquatic environment. Osteoarticular pathology caused by Aeromonas hydrophila is rarely encountered. To the best of our knowledge, this is the first case of chronic osteomyelitis by A. hydrophila reported from India. We report a case of chronic osteomyelitis of the lower limb due to A. hydrophila, which occurred as a delayed complication following open reduction and internal fixation. Prompt medical and surgical intervention supplemented by a comprehensive microbiological workup aided in pathogen identification and specific antimicrobial administration resulting in the successful outcome of our patient. This case illustrates the utility of multidisciplinary management approach involving microbiologists and orthopedicians in investigating and appropriately managing such cases.