Spectrum of Bacterial Pathogens in Critical COVID-19 Patients Admitted in Intensive Care Units of a Tertiary Care Hospital During the First and Second Wave of the Pandemic

Objective: This study intends to compare the clinical characteristics and the prevalence and spectrum of bacterial pathogens in COVID-19 patients admitted to ICU during the first and second waves at a tertiary care, teaching and referral hospital of eastern India. Method: This is a hospital-based retrospective study which analysed demographic details, clinical profile and bacterial culture results of severe and critically ill COVID-19 patients admitted in intensive care units (ICU) during April -Oct 2020 (1 st wave) and April –July 2021 (2 nd wave). Result: The patients admitted during the 2 nd wave were comparatively older and had multiple comorbidities compared to the 1 st wave. (23.8%) (45/189) and 50% (173/346) of the COVID-19 patients admitted to ICU developed bacterial infection during the 1 st and 2 nd wave respectively. Overall, there was predominance of multidrug resistant Gram negative bacilli in both the waves. There was increased isolation of intrinsic colistin resistant microorganisms. Conclusion: Multidrug resistant Gram negative bacterial infections, remain a dreaded complication in severe and critically ill hospitalised COVID-19 patients requiring ICU care and high usage of colistin spirals the emergence and spread of pathogens intrinsically resistant to colistin.

Role of reverse transcriptase polymerase chain reaction for the diagnosis of human rabies.

Background & objectives: Traditionally, rabies diagnosis is made by demonstration of rabies viral antigen by direct immunofluorescence (DIF) and mouse inoculation test (MIT). The present study was carried out to evaluate the role of reverse transcriptase-polymerase chain reaction (RT-PCR) in comparison with these conventional techniques for the diagnosis of rabies. Methods: Skin biopsies, corneal impression smears and saliva sample were collected ante-mortem and brain tissue and CSF were collected post-mortem from ten clinically suspected rabies patients. DIF, Seller staining, MIT and RT-PCR were performed on the patients’ samples for the diagnosis of rabies. The ability of RT-PCR to detect rabies virus earlier as compared to other assays was tested both for reference virus as well as clinical isolates. Results: All samples taken ante-mortem were negative for DIF test. Six of 10 post-mortem brain tissues of the clinically suspected patients were positive both by RT-PCR and MIT, of these six, five were positive by DIF test and four were positive by Seller stain. RT-PCR could detect the rabies virus earlier as compared to DIF, both from clinical isolates and fixed rabies virus. Interpretation & conclusions: The present results showed 100 per cent sensitivity and specificity of RT-PCR as compared to 83.3 per cent of DIF and 66.7 per cent of Sellers stain for diagnosis of rabies. RT-PCR also detected rabies viral infection earlier as compared to conventional tests and can also be used on ante-mortem samples. Thus, the present study shows the usefulness of RT-PCR as an alternative to MIT for the confirmation of rabies diagnosis.

Viral markers in patients with hemophagocytosis: A prospective study in a tertiary care hospital.

Background : Hemophagocytic syndrome (HPS) is a rare clinicopathological condition characterized by the activation of macrophages with prominent hemophagocytosis in bone marrow and other reticulo-endothelial systems. HPS can be familial or secondary to infections including viruses. Aim : To study the viral markers in patients with HPS. Materials and Methods : Serum samples of patients with HPS and control group were screened for anti EBV VCA IgM, and IgG, anti-Parvo B19 IgM, and anti-CMV IgM antibodies using commercially available ELISA kits and CMV and ParvoB19 DNA by polymerase chain reaction (PCR). Results and Discussion : The present prospective study reports the profile of viral markers in HPS cases from north India. Among the 14 HPS cases 43% (6/14) were positive for at least one viral marker tested, of which EBV was found to be the most prevalent (3/6: 50%) followed by parvovirus B19(2/6: 33%) and cytomegalovirus (1/6: 17%). Mortality was noted in 33% of virus associated HPS patients. Our study highlights the higher association of Epstein-Barr virus (EBV) with HPS as compared to other viruses along with higher rate of mortality in both parvovirus B 19 and EBV associated HPS.

Preservation of continuous cell lines at -85°C: A low-cost alternative for resource limited countries.

Background: Cell culture is the most popular method of virus propagation because of its high sensitivity. However, the need of high cost liquid nitrogen for storage of cell lines is one of the main limiting factor for its widespread use in developing countries. Objective: The present study was therefore carried out to standardize the preservation of continuous cell lines at deep freezer (-85ºC) for 6 months. Methods: Fixed number of Vero and Hep2 cells were preserved at -85ºC deep freezer in separate vials and were revived at one month interval to check the growth pattern. Results: Both the cell lines could be revived with healthy cells and monolayer was formed within 7-10 days, after storage at -85ºC for 4 months. Conclusion: The present study highlights the utility of -85ºC deep freezer as an alternative to liquid N 2 for preservation of these cell lines at least up to four months.

Congenital rubella and cytomegalovirus infections in and around Chandigarh.

AIMS: This study has analyzed the role of rubella and cytomegalovirus (CMV) in infections of children and pregnant women. SETTINGS AND DESIGN: The study was carried out in a tertiary care hospital. Data from blood samples from pregnant women (asymptomatic and also women with obstetric problems) and children (suspected of intrauterine infections) that were received in the laboratory over a period of 8 years were analysed. MATERIALS AND METHODS: The samples were tested for rubella- and CMV-specific IgM antibodies by capture enzyme linked immunosorbent assay. RESULTS: In children, the overall positivity for rubella- and CMV-specific IgM antibodies was 2.8% and 12.5%, respectively. In asymptomatic pregnant females, rubella positivity was 0.7% while in women with obstetric complications it was 3.4%. IgM antibody positivity in cases of CMV was 7.8% in both asymptomatic pregnant women and also in women with obstetric complications. CONCLUSIONS: The study indicated that infection with CMV is more common than the rubella virus. The incidence of rubella has reduced over the past few years. Hence, screening for rubella infection may be reserved for women with obstetric complications only. The routine screening for CMV among all antenatal cases is a debatable issue.

Cytomegalovirus pneumonia in adult acute lymphoblastic leukemia.

Cytomegalovirus (CMV) is an important cause of morbidity and mortality in immunosuppressed patients. Though acute lymphoblastic leukemia (ALL) is an immunosuppressed state, CMV disease has been reported infrequently. We present a patient of adult B lineage ALL who was on maintenance chemotherapy and developed CMV pneumonia. Patient was managed with intravenous ganciclovir and had successful outcome. However, three weeks later patient had a relapse of ALL and died shortly after high dose chemotherapy.

Respiratory syncytial virus is not an important community acquired pathogen in adult hematological malignancy patients.

Respiratory syncytial virus (RSV) has been reported as a leading cause of upper and lower viral respiratory tract infection in high-risk adult populations. We prospectively studied 48 patients with hematological malignancies over a period of 2 years. Throat and nasal washings were subjected to immunofluorescence and cell culture for virus isolation. Of these 48 patients, 31 had acute leukemia, 6 had chronic leukemia, 10 had lymphoma and one had multiple myeloma. The median age of the patients was 20 years with a male to female ratio of 4:1. No RSV was detected in any of the samples. RSV was not found as a major cause of community acquired upper respiratory tract infections in adults with hematological malignancies in India.

Otosclerosis--do we have a viral aetiology?

The etiology of otosclerosis remains an enigma though there are evidences suggesting a viral involvement. This study aimed to find out the relationship between viral infections and otosclerosis. Twenty two patients with otosclerosis and 10 healthy controls were included in the study. IgM antibodies to varicella zoster virus (VZV), measles, rubella, human cytomegalovirus (HCMV) and herpes simplex virus (HSV) were detected using micro ELISA. Paul Bunnel Davidsohn test was performed to rule out Ebstein Barr virus (EBV) infection. Overall, 5(22.7%) patients showed antibodies to one or more viruses. IgM antibodies against measles and VZV could be demonstrated in 4(18.1%) and 1(4.5%) patients respectively. None of the samples were found to be positive for HSV, HCMV, rubella and EBV antibodies. Controls were negative for all the viruses tested. The difference in seropositivity between the patient and control group was not statistically significant (p>0.05). Thus, this study suggests that otosclerosis is not commonly associated with a systemic viral infection.

Heterophile antibody positive infectious mononucleosis.

OBJECTIVE: The present study has been carried out to analyse the trend of heterophile antibody positive infectious mononucleosis cases. METHODS: A total of 1741 cases of clinically suspected infectious mononucleosis from various age groups were investigated during the period January, 1986 to December, 2000 and were analysed for infectious mononucleosis (IM) specific heterophile antibody by Paul-Bunnel-Davidsohn (PBD) test. Forty seven heterophile antibody negative samples were also tested simultaneously for the presence of the IgG antibody to viral capsid antigen (VCA) and Epstein Barr nuclear antigen (EBNA) to detect the exposure to Epstein Barr Virus (EBV) infection. RESULTS: The overall percentage of EBV specific heterophile (Paul-Bunnel) antibody positivity was found to be 11.1% (194/1741). The average Paul-Bunnel antibody positivity between 1986 to 1990 was 20.5% which declined drastically to 5.7% during 1991-2000. Males comprised of 55.2% of the serologically proven IM cases. Of the 47 heterophile antibody negative cases, 38 (80.9%) and 33 (70.2%) were found to be positive for anti-VCA IgG and anti-EBNA IgG antibodies respectively. Paul Bunnel antibody positivity was found to be higher in >14 year age group patients than those below 14 years. CONCLUSION: These findings suggest that the EBV infection still continues to be endemic in this part of the country, however, a declining trend in IM cases was observed during the last decade.

Rapid detection of mutations in rpoB gene of rifampicin resistant Mycobacterium tuberculosis strains by line probe assay.

BACKGROUND & OBJECTIVES: Multidrug resistant (MDR) tuberculosis (TB) is a problem of increasing importance in the world due to limited treatment options. Resistance to rifampicin results from nucleotide changes in the gene encoding the beta subunit of the RNA polymerase (rpoB) of Mycobacterium tuberculosis. Rifampicin resistance is considered as a marker for MDR TB. The nature and frequency of mutations in the rpoB gene of rifampicin resistant clinical isolates vary considerably according to the geographical location and very little information is available on specific mutational patterns in India. This study was undertaken to detect and characterize the rpoB gene mutation associated with rifampicin resistance in M. tuberculosis by line probe assay. METHODS: A total of 36 strains of M. tuberculosis were analysed by INNO-LiPA Rif TB and compared with the results of conventional susceptibility testing method. After PCR amplification of the region of RNA polymerase involved in rifampicin resistance, the amplified product was hybridized with a set of 10 oligonucleotides immobilized onto a membrane strip. From the pattern obtained, the presence or absence of rifampicin resistance in the M. tuberculosis strains was assessed. RESULTS: It was found that the M. tuberculosis probe was 100 per cent specific; the most frequently observed mutation was His-526-Tyr in the rpoB gene; and correlation between the results of the LiPA and those obtained by the classical susceptibility testing was excellent (100%). INTERPRETATION & CONCLUSION: INNO LiPA was found to be a reliable, simple, rapid and informative tool for the early detection and characterization of rpoB mutation associated with rifampicin resistance in M. tuberculosis in the clinical laboratory setting and may constitute an important molecular method for the control of tuberculosis.