ABSTRACT
BackgroundThe current global pandemic of Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2 led to the investigation with clinical, biochemical, immunological and genomic characterization from the patients to understand the pathophysiology of viral infection. MethodsSamples were collected from six asymptomatic and six symptomatic SARS-CoV-2 confirmed hospitalized patients in Bhubaneswar, Odisha, India. Clinical details, biochemical parameters, treatment regime were collected from hospital, viral load was determined by RT-PCR, levels of cytokines and circulating antibodies in plasma were assessed by Bioplex and isotyping respectively. In addition, the whole genome sequencing of viral strains and mutational analysis were carried out. FindingsAnalysis of the biochemical parameters highlighted the increased levels of C-Reactive protein (CRP), lactate dehydrogenase (LDH), serum SGPT, serum SGOT and ferritin in symptomatic patients indicating that patients with higher levels of few biochemical parameters might experience severe pathophysiological complications after SARS-CoV-2 infection. This was also observed that symptomatic patients were mostly with one or more comorbidities, especially diabetes (66.6%). Surprisingly the virological estimation revealed that there was no significant difference in viral load of oropharyngeal (OP) samples between the two groups. This suggests that the viral load in OP sample does not correlate with the disease severity and both asymptomatic and symptomatic patients are equally capable of transmitting the virus. Whereas, viral load was higher in plasma and serum samples of symptomatic patients suggesting that the development of clinical complications is mostly associated to high viral load in plasma and serum. This also demonstrated that the patients with high viral load in plasma and serum samples were found to develop sufficient amounts of antibodies (IgG, IgM and IgA). Interestingly, the levels of 7 cytokines (IL-6, IL-.1, IP-10, IL-8, IL-10, IFN-2, IL-15) were found to be highly elevated in symptomatic patients, while three cytokines (soluble CD40L, GRO and MDC) were remarkably higher in asymptomatic patients. Therefore, this data suggest that cytokines and chemokines may serve as "predictive indicator" of SARS-CoV-2 infection and contribute to understand the pathogenesis of COVID-19. The whole genome sequence analysis revealed that the current isolates were clustered with 19B, 20A and 20B clades, however acquired 11 additional changes in Orf1ab, spike, Orf3a, Orf8 and nucleocapsid proteins. The data also confirmed that the D614G mutation in spike protein is mostly linked with higher virus replication efficiency and severe SARS-CoV-2 infection as three patients had higher viral load and among them two patients with this mutation passed away. InterpretationThis is the first comprehensive study of SARS CoV-2 patients from India. This will contribute to a better understanding of the pathophysiology of SARS-CoV-2 infection and advance in the implementation of effective disease control strategies. FundingThis study was supported by the core funding of Institute of Life Sciences, Bhubaneswar, Dept of Biotechnology, India. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSAsymptomatic patients are a source of concern as measures taken to control the spread of the virus are severely impacted by their undetectability. Presently, there is an inadequate information about the characteristics of the asymptomatic and symptomatic patients. The association between SARS-CoV-2 viral load, cytokines and risk of disease progression remains unclear in COVID-19 in Indian scenario. PubMed was searched for articles published up to May, 2021, using the keywords "SARS CoV-2 patients in India", or "2019 novel coronavirus patients in India". No published work about the patients data on SARS CoV-2 in Indian scenario could be identified. Added value of this studyThis investigation highlights the ability of both asymptomatic and symptomatic patients to transmit the virus equally. This study also demonstrates that the D614G mutation in the spike protein is associated with severe SARS-CoV-2 infection and enhance levels of inflammatory markers such as CRP and ferritin which can be predictive biomarkers for critical condition of patients. This is the first comprehensive study of SARS CoV-2 patients from India and will contribute to a better understanding of the pathophysiology of SARS-CoV-2 infection by advancing the implementation of effective disease control strategies. Implications of all the available evidenceThe current global pandemic of Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2 led to the investigation with clinical, biochemical, immunological and viral genome sequencing to understand the pathophysiology of this virus infection. Samples were collected from six asymptomatic and six symptomatic SARS-CoV-2 confirmed hospitalized patients in Bhubaneswar, Odisha, India. This investigation highlights the ability of both asymptomatic and symptomatic patients to transmit the virus equally. This also demonstrated that the D614G mutation is mostly associated with higher virus replication capacity and severe SARS-CoV-2 infection and enhanced levels of inflammatory markers such as CRP and ferritin which are associated with critical conditions of patients. This is the first comprehensive study of SARS CoV-2 patients from India and will contribute to a better understanding of the pathophysiology of SARS-CoV-2 infection by advancing the implementation of competent disease control strategies.
ABSTRACT
We present a microfluidic chip for immobilizing Drosophila melanogaster larvae for high resolution in vivo imaging. The chip creates a low-temperature micro-environment that anaesthetizes and immobilizes the larva in under 3 minutes. We characterized the temperature distribution within the chip and analyzed the resulting larval body movement using high resolution fluorescence imaging. Our results indicate that the proposed method minimizes submicron movements of internal organs and tissue without affecting the larva physiology. It can be used to continuously immobilize larvae for short periods of time (minutes) or for longer periods (several hours) if used intermittently. The same chip can be used to accommodate and immobilize arvae across all developmental stages (1st instar to late 3rd instar), and loading larvae onto the chip does not require any specialized skills. To demonstrate the usability of the chip, we observed mitochondrial trafficking in neurons from the cell bodies to the axon terminals along with mitochondrial fusion and neuro-synaptic growth through time in intact larvae. Besides studying sub-cellular processes and cellular development, we envision the use of on chip cryo-anesthesia in a wide variety of biological in vivo imaging applications, including observing organ development of the salivary glands, fat bodies and body-wall muscles.
