INTRODUCTION:

In China's Wuhan city, the corona virus outbreak was first reported in December 2019. The World Health Organisation (WHO) declared the corona virus pandemic (occurring over a wide geographic area and affecting an exceptionally high proportion of the population) on March 11, 2020 when cases began to appear all across the world (Priyanka et al., 2021)¹. The last pandemic documented in the world was the H1N1 flu in 2009 Kishor R, Ramhari B, 2020².

As of December 31st, 2023, the WHO stated that there were 773,819,856 confirmed cases of COVID-19 worldwide, resulting in 7,010,568 fatalities. As of November 26, 2023 13.59 billion vaccine doses had been administered since its introduction on 22 July 2020 with 67% and 32% of global population being administered primary and booster dose, respectively (WHO covid-19 dashboard, 2022)³.

From 3 January 2020 to 31 December 2023, there were 45,013,172 confirmed cases of COVID-19 in India, with 533,361 fatalities recorded to the World Health Organisation. As of November 26, 2023 2.21 Billion doses of vaccines have been given since its introduction in India on 16 January 2021 with 69% and 17% of the population being administered primary and booster dose, respectively. (WHO covid-19 dashboard, 2022)³.

On November 28, 2022, the Jammu and Kashmir section alone reported 479363 verified COVID-19 cases to the J&K Govt (NHM J&K) with 4785(1.00%) deaths, 474552(99.00%) recovered cases, and 26(0.01%) active cases⁴. A total of 26859586 lab tests for COVID-19 had been completed by November 28th, 2022 ⁴, with 479363 positive cases identified and 26380223 negative tests reported. As a result, the positivity and negativity rates were, respectively, 1.78% and 98.22%. As of February 13, 2022, District Doda in a particular area of J & K reported 11,074 confirmed instances of COVID-19, including 144(1.3%) fatal cases, 10,362(93.6%) recovered cases, and 568(5.1%) active cases. Similar statistics may be seen for District Kishtwar, which reported 5828 confirmed cases with 45 fatalities, 5742 recovered cases, and 41 active cases (NHM, 2022)⁴.

Reverse transcription-polymerase chain reaction (RT-PCR)-based viral detection cases have been documented in some investigations at various stages of the disease course (Zou L et al., 2020; Pan Y et al., 2020; Wang W et al., 2020)^5,6,7. But these publications kept an eye on SARS-CoV-2 during the early stages of infection. In the Chenab valley of Jammu and Kashmir, no study has yet described the viral dynamics of SARS-CoV-2 infection during a lengthy observation period. Understanding the viral profile in a patient's respiratory specimens can help with diagnosis and provide insight into the course of the illness. As a result, we obtained clinical samples from 60 patients, identified COVID-19 as positive on RTPCR, and published findings of SARS-CoV-2 detection over the course of the illness.

Transmission:

Covid-19 spreads when virus particles from infected patients' respiratory droplets are coughed or sneezed out and come into contact with a healthy person's eye, nose, or mouth mucosa. Sarkar et al., 2021⁸. The usage of infected bed linens, blankets, kitchen utensils, thermometers and stethoscopes has also been linked to the transmission of viruses (Parasher A. 2021)⁹.

The incubation phase, also known as the presymptomatic period or prepatent period, lasts for 5.1 days from the moment of virus exposure to the beginning of symptoms, with 97.5% of the population experiencing symptoms by 11.5 days (Lauer SA et al., 2019)¹⁰.

Patho physiology:

Although little is known about the progression of a disease, COVID-19 has been classified into three stages (Parasher A.2021)⁹ based on the distinct yet overlapping symptoms of dyspnea, fever, pain, weakness, tightness in the chest, loss of taste and smell, and dyscrasia, which are all accompanied by acute respiratory distress syndrome (ARDS) (Huang C et al., 2020; Vaira LA et al.^11,12.

SARS-CoV-2 attaches to upper respiratory tract nasal epithelial cells during the asymptomatic stage. As the primary receptor for viral entrance into host cells, ACE-2 (Angiotensin Converting Enzyme) is expressed in adult respiratory tract (nasal epithelium) (Wan Y et al., 2020)¹³ (Hoffmann M et al., 2020)¹⁴. As the virus infects the respiratory tract's cilia, it multiplies quickly (Sims AC et al., 2005)¹⁵. This phase lasts for the first few days since the patient has a low viral load and only experiences a minor immunological reaction. Patient is very contagious, and a nasal swab test can find the virus (Parasher A.2021)⁹.

Upper respiratory tract infection: A virus enters the upper respiratory system through the nasal epithelial cells, causing fever, a dry cough, and other flu-like symptoms. The majority of patients do not move on to the next stage because a stronger immune response is enough to stop the infection. The virus-infected cells secrete INF-beta, INF-lembda, and CXCL-10 (C-X-C pattern chemokine ligand 10) (Tang NL et al., 2005)¹⁶.

Lower respiratory tract infection/acute respiratory distress syndrome: Via ACE-2 receptors, the virus enters the host type-II alveolar cells and quickly replicates, producing additional nucleocapsids. According to Tang NL et al. (2005)¹⁶, the virus can be found in sputum and nasal swabs, and the disease is said to be clinically evident. The virus infected cells release various cytokines and inflammatory markers like interleukins (IL-1, IL-6, IL-8), TNF-alpha, INF-beta, INF-lembda, monocyte chemoattractant proteins (MCP-1), macrophages, CXCL-10 bringing 'cytokine storm' which further attracts neutrophils, CD-4 helper T-cells and CD-8 cytotoxic T-cells leading to sequestration of lungs. After replication, the virus is liberated by host cell death and continues to infect neighbouring type-II pneumocytes. Acute respiratory distress syndrome (ARDS) is caused by the loss of both type-I and type-II pneumocytes as a result of the prolonged dissemination of viral particles and the ensuing inflammatory response (Cascella M et al., 2020; Xu Z et al., 2020)^17,18. The fibrin-rich hyaline membrane with multinucleated gaint cells is the pathological outcome of alveolar involvement (Xu Z et al., 2020: Gu J, Korteweg C 2007) ^18,19. Vomiting, discomfort in the abdomen, and loose stools are also common (Parasher A 2021)⁹.

METHOD AND MATERIAL:

Location:

This study comprised 100 hospitalised patients with proven SARS-CoV-2 infection in 3 districts of Chenab valley (Doda, Baderwah, and Kishtwar) between January 21, 2021, and February 12, 2021.

According to the interim guidance from the Centres for Disease Control and Prevention (CDC, 2020: CNHC, 2020) and the diagnosis and treatment guidelines for SARS-CoV-2 from the Chinese National Health Committee (version 5) [20,21], all enrolled patients had a confirmed diagnosis of COVID-19.

Sampling:

Patients were sampled on several days following the beginning of symptoms using a nasal or throat swab.

The COVID-19 nucleic acid detection kit (Meril COVID-19 One step RT-PCR Kit) was used in an RT-PCR experiment to detect SARS-CoV-2 according to the manufacturer's instructions. The real-time RT-PCR technique specifically examined 2 target genes, namely nucleo-capsid protein (N) and open reading frame 1ab (ORF1ab). The time from the onset of symptoms and the date of the first negative RT-PCR test result was considered the viral nucleic acid conversion time if two consecutive negative findings were obtained. All information (test dates and RT-PCR assay results) was gathered up until the last follow-up date (3rd March 2021).

Ethics clearance:

According to GMCDoda/IEC/2022/34, the Institutional Ethical Committee of GMC Doda has ethically authorised the current study.

RESULTS:

This study includes 100 patients with COVID-19 diagnoses in total. All of the included patients had mild to moderate illness, according the criteria (CNHC, 2020) ²¹. The intensive care unit (ICU) did not receive any new patients. With 61 (61.00%) men and 39 (39.00%) women, the median age was 55 years (interquartile range (IQR) 42-68 years; range 25-83 years). All patients made a full recovery and were let go from the hospital at the conclusion of a follow-up.

500 SARS-CoV-2 RT-PCR assays in all, with 5 tests per patient, were performed on 100 patients. The longest time between two SARS-CoV-2 RT-PCR testing was 42 days following the onset of symptoms. 24 days (IQR, 18–31 days) was the median amount of time between the beginning of symptoms and nucleic acid conversion. Table 1 and Figure 1 display the percentages of RT-PCR assays that returned positive and negative findings. The majority of the SARS-CoV-2 RT-PCR results were positive in the first three weeks following the onset of symptoms. The number of negative RT-PCR results rose starting in week 3 after the onset of symptoms. At week 6 following the onset, all RT-PCR test outcomes were unfavourable (Table 1). The percentage of RT-PCR test results that were positive was 100% in week 1, 89.3% in weeks 2, 3, 4, 5, and 6, and 0% in week 6 (Figure 2).

Table 1: Results of RT-PCR

	Week 1	Week 2	Week 3	Week 4	Week 5	Week 6	Total
Positive	34	50	50	34	17	0	185
Negative	0	17	66	116	83	33	315
Total	34	67	116	150	100	33	500

Fig 1: Results of RT-PCR

Fig 2: Positivity rate

DISCUSSION:

A number of instances of pneumonia caused by the -corona virus were reported in Wuhan, China, in December 2019. Chinese researchers isolated a SARS-CoV-2 from a patient on January 7th, 2020, and sequenced the virus' genome. The 2019-novel corona virus (2019-nCoV) was identified by the World Health Organisation (WHO) on January 12th, 2020²². Coronavirus is a single-stranded, positive sense RNA-containing, 120–160nm-sized enclosed particle. There are six different coronavirus types that have been identified as infecting people. COVID-19 is the most recent coronavirus with the one and seven digits. Coronavirus-related illnesses, which can range from the common cold to the severe acute respiratory syndrome (SARS-CoV) Balai MK, 2021²³, are zoonotic diseases that are spread from animals to people. Transmembrane protease serine 2 (TMPRSS2), an endothelial cell surface enzyme, has the ability to stimulate the viral melding into respiratory epithelium through binding to ACE2. The viral primary spike (S) protein of the SARS-CoV-2 ties to the angiotensin-converting enzyme 2 (ACE2) receptor in an alveoli cell. Alaraj M. 2022²⁴.

This study is the first case series from 100 COVID-19 patients with 500 RT-PCR test samples for the detection of SARS-CoV-2. Our first findings are noteworthy for showing that patients with SARS-CoV-2 infection have a dynamic character. For the detection of SARS-CoV-2 Vaishnavi, K, 202025 advocate real-time reverse transcription polymerase chain reaction (RT-PCR) is the most widely used assay. According to genomic research, SARS-CoV-2 and SARS-CoV, which generated an epidemic with 8096 confirmed cases worldwide in 2002–2003 (WHO, 2004), have around 80% of the same genetic makeup²⁶. Based on epidemiological and clinical data, it was hypothesised that SARS-CoV-2 was spread from person to person (Ping Y et al., 2020; Chan JF et al., 2020)^27,28. The viral nucleic acid shedding pattern of patients with SARS-CoV-2 infection is different from that of SARS-CoV, which had modest viral loads in the early stage and peaked approximately 10 days after symptom onset (Peiris JS et al., 2003)²⁹. SARS-CoV-2 shares similar sequencing characteristics with SARS-CoV and Middle East respiratory syndrome corona virus (MERS-CoV)³⁰. Our study gathered the serial RT-PCR test results from 100 COVID-19-recovered patients and looked at the dynamic profile over the course of the illness. We demonstrated that after 3 weeks of the onset of symptoms, the majority of patients tested positive for SARS-CoV-2 on RT-PCR assays. From week 4 after the onset of symptoms, the negative findings on RT-PCR testing for SARS-CoV-2 started to predominate, and by the end of the follow-up (6 weeks), all RT-PCR test results were negative. Throughout the observation period of six weeks, the positive RT-PCR test result rate continued to decrease (Figures 1 and 2). According to the aforementioned research, SARS-CoV-2 viral replication lasts for a sizable amount of time in infected patients.

In this investigation, we sought to determine whether COVID-19 patients' clinical traits and viral shedding were related. According to our research, patients with delayed viral nucleic acid conversion are typically older and have more comorbid conditions. Older people may be more susceptible to coronavirus infection, which may be caused by immunological pathogenesis and the creation of a proinflammatory cytokine storm (Guan WJ et al., 2020)³⁰. Older people with compromised immune systems may experience a protracted virus clearance time.

False-negative results on RT-PCR for SARS-CoV-2 are quite frequent in clinical settings as a result of mistakes in sample and testing. The Chinese National Health Committee's (CNHC) current diagnosis and treatment recommendations for SARS-CoV-2 recommend that the criteria for releasing a patient include improvement in radiography, relief of symptoms, and two consecutive negative RT-PCR results for SARS-CoV-2 (CNHC, 2020)²¹. In a recent study by [Lan L et al., 2020]³¹, it was discovered that patients who had recovered from COVID-19 had positive RT-PCR test findings. These afflicted patients might be the ones that spread the infection. The aforementioned findings cast doubt on the existing discharge standards. Evidence revealed that the rapid person-to-person transmission ability of COVID-19 may be associated to its outbreaks (Zunyou W and Jennifer MG, 2020)³². Traditional public health measures including isolation, quarantine, and community containment are essential to restrict the spread of COVID-19 because particular therapy has not been established (Li Q et al., 2020:McCloskey B and Heymann DL, 2020)^33,34. In non-ICU patients who had COVID-19 during the course of the illness, this preliminary analysis discovered evidence of the dynamic profile of SARS-CoV-2. We recommend extended surveillance and repeat confirmation of RT-PCR testing from respiratory specimens for safe discharge and the end of quarantine based on the findings of our study.

CONCLUSION:

According to this study, patients who have prolonged viral nucleic acid conversion are typically older and have more co-morbid conditions like diabetes and hypertension.

CONFLICT OF INTEREST:

None of the authors have any conflicts of interest.

AUTHORS CONTRIBUTION:

Sameera Akhtar collected the samples, processed them, and compiled the results along with writing of manuscript. Shoket Ali did formal analysis and methodology, Imran Bashir edited the manuscript, and Sufhia Akhter conceptualization of the work and data curation.

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Received on 24.11.2022 Modified on 15.05.2023

Research J. Pharm. and Tech 2024; 17(3):1277-1281.

DOI: 10.52711/0974-360X.2024.00200