Worldwide prevalence of microbial agents’ coinfection among COVID‐19 patients: A comprehensive updated systematic review and meta‐analysis

BackgroundTo provide information about pathogens’ coinfection prevalence with SARS‐CoV‐2 could be a real help to save patients’ lives. This study aims to evaluate the pathogens’ coinfection prevalence among COVID‐19 patients.MethodIn order to find all of the relevant articles, we used systematic search approach. Research‐based databases including PubMed, Web of Science, Embase, and Scopus, without language restrictions, were searched to identify the relevant bacterial, fungal, and viral coinfections among COVID‐19 cases from December 1, 2019, to August 23, 2021. In order to dig deeper, other scientific repositories such as Medrxiv were probed.ResultsA total of 13,023 studies were found through systematic search. After thorough analysis, only 64 studies with 61,547 patients were included in the study. The most common causative agents of coinfection among COVID‐19 patients were bacteria (pooled prevalence: 20.97%; 95% CI: 15.95–26.46; I ²: 99.9%) and less frequent were virus coinfections (pooled prevalence: 12.58%; 95% CI: 7.31–18.96; I ²: 98.7%). The pooled prevalence of fungal coinfections was also 12.60% (95% CI: 7.84–17.36; I ²: 98.3%). Meta‐regression analysis showed that the age sample size and WHO geographic region did not influenced heterogeneity.ConclusionWe identified a high prevalence of pathogenic microorganism coinfection among COVID‐19 patients. Because of this rate of coinfection empirical use of antibacterial, antifungal, and antiviral treatment are advisable specifically at the early stage of COVID‐19 infection. We also suggest running simultaneously diagnostic tests to identify other microbiological agents’ coinfection with SARS‐CoV‐2.     

Prolonged SARS‐Cov‐2 shedding with rapid IgG antibody decay in a COVID‐19 patient: A case report

BackgroundThe coronavirus disease 2019 (COVID‐19) epidemic is still spreading rapidly around the world. Recent cases with prolonged severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) RNA detection have been successively reported, and the phenomenon of false‐negative real‐time polymerase chain reaction (RT‐PCR) results of SARS‐CoV‐2 RNA or “repositive” was also described in COVID‐19 patients.MethodsWe report a 69‐year‐old female patient with hypertension, suspected lung tumor, and previous history of total hysterectomy for hysteromyoma who presented with moderate COVID‐19 symptoms and was positive for SARS‐CoV‐2 RNA by RT‐PCR when she traveled from the USA to China.ResultsThe patient required second and third re‐hospitalizations due to “repositive” SARS‐CoV‐2 throat swab test results during post‐charge solitary isolation and observation, and serum SARS‐CoV‐2‐IgG decayed rapidly before disappearing on illness Day 139 when the throat swab was still positive. The virus shedding lasted for at least 146 days (the last positive throat swab test result was on illness Day 146, and the first true‐negative test result was on illness Day 151) since her initial positive test.ConclusionProlonged SARS‐CoV‐2 RNA viral shedding is prone to occur in an immunocompromised host, wherein changes in the host immune status can lead to repeated positive SARS‐CoV‐2 detection. Moreover, the SARS‐CoV‐2‐IgG may decrease rapidly and disappear before virus removal, indicating there may be certain limitations on the protective effect of the SARS‐CoV‐2 antibody, which deserves clinical attention.     

The utility of SARS‐CoV‐2 nucleocapsid protein in laboratory diagnosis

BackgroundThe Coronavirus Disease 2019 (COVID‐19) is caused by the severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), which has now become a global pandemic owing to its high transmissibility. The SARS‐CoV‐2 nucleocapsid protein tests are playing an important role in screening and diagnosing patients with COVID‐19, and studies about the utility of SARS‐CoV‐2 nucleocapsid protein tests are increasing now.MethodsIn this review, all the relevant original studies were assessed by searching in electronic databases including Scopus, Pubmed, Embase, and Web of Science. “SARS‐CoV‐2”, “COVID‐19”, “nucleocapsid protein”, and “antigen detection” were used as keywords.ResultsIn this review, we summarized the utility of SARS‐CoV‐2 nucleocapsid protein in laboratory diagnosis. Among the representative researches, this review analyzed, the sensitivity of SARS‐CoV‐2 nucleocapsid protein detection varies from 13% to 87.9%, while the specificity could almost reach 100% in most studies. As a matter of fact, the sensitivity is around 50% and could be higher or lower due to the influential factors.ConclusionIt is well suggested that SARS‐CoV‐2 nucleocapsid protein is a convenient method with a short turnaround time of about half an hour, and the presence of N antigen is positively related to viral transmissibility, indicating that SARS‐CoV‐2 N protein immunoassays contribute to finding out those infected people rapidly and segregating them from the uninfected people.     

Hypersensitivity in the lungs is responsible for acute respiratory failure in COVID‐19 patients: Case series of patients who received high‐dose/short‐term methylprednisolone

BackgroundCOVID‐19 is a highly contagious respiratory disease caused by the SARS‐CoV‐2 virus. Patients with severe disease have a high fatality rate and face a huge medical burden due to the need for invasive mechanical ventilation. Hypoxic respiratory failure is the major cause of death in these patients. There are currently no specific anti‐SARS‐CoV‐2 drugs, and the effect of corticosteroids is still controversial.MethodsThe clinical data of 102 COVID‐19 patients, including 27 patients with severe disease, were analyzed. The serum levels of total IgE and anti‐SARS‐CoV‐2 specific IgE were compared in healthy controls and COVID‐19 patients, changes in the level of anti‐SARS‐CoV‐2 specific IgE and clinical response to methylprednisolone (MP) treatment were analyzed, and the effect of high‐dose/short‐term MP therapy for patients with critical illness and respiratory failure was determined.ResultsCOVID‐19 patients had elevated serum levels of anti‐SARS‐CoV‐2 specific IgE, and patients with severe disease, especially critical illness, had even higher levels. Application of short‐term/high‐dose MP significantly reduced the level of these IgE antibodies and also blocked the progression of hypoxic respiratory failure. Hypoxic respiratory failure in patients with COVID‐19 is related to pulmonary hypersensitivity.ConclusionsHypersensitivity in the lungs is responsible for acute respiratory failure in COVID‐19 patients. Application of high‐dose/short‐term MP appears to be an effective life‐saving method for COVID‐19 patients who have hypoxic respiratory failure.     

Performance evaluation of four rapid antibody tests for the detection of severe acute respiratory syndrome coronavirus 2

BackgroundThe prompt detection of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) is important in the therapeutic management of infected patients. Rapid diagnostic tests are widely used for this purpose. This study aimed to evaluate the clinical performance of four SARS‐CoV‐2 immunoglobulin IgG/IgM rapid diagnostic tests in the detection of SARS‐CoV‐2.MethodsNasopharyngeal and oropharyngeal swabs and/or sputum were collected from 30 patients infected with SARS‐CoV‐2 and 30 healthy volunteers. All specimens were tested using four SARS‐CoV‐2 IgG/IgM rapid diagnostic tests and real‐time polymerase chain reaction. We assessed the clinical sensitivity and specificity of the tests.ResultsThe clinical sensitivity of FREND™, SsmarTest™, BIOCREDIT™, and IVDLAB™ was 96.67%, 100.00%, 100.00%, and 96.67%, respectively, compared to real‐time polymerase chain reaction. The clinical specificity was 96.67%, 100.00%, 86.67%, and 96.67%, respectively.ConclusionThese findings could expedite the detection of SARS‐CoV‐2 and thus reduce the risk of further transmission of the virus.     

Large scale production and characterization of SARS‐CoV‐2 whole antigen for serological test development

BackgroundThe rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) has generated a pandemic with alarming rates of fatality worldwide. This situation has had a major impact on clinical laboratories that have attempted to answer the urgent need for diagnostic tools, since the identification of coronavirus disease 2019 (COVID‐19). Development of a reliable serological diagnostic immunoassay, with high levels of sensitivity and specificity to detect SARS‐CoV‐2 antibodies with improved differential diagnosis from other circulating viruses, is mandatory.MethodsAn enzyme‐linked immunosorbent assay (ELISA) using whole inactivated virus cultured in vitro, was developed to detect viral antigens. WB and ELISA investigations were carried out with sera of convalescent patients and negative sera samples. Both analyses were concurrently performed with recombinant MABs to verify the findings.ResultsPreliminary data from 10 sera (5 patients with COVID‐19, and 5 healthy controls) using this immunoassay are very promising, successfully identifying all of the confirmed SARS‐CoV‐2‐positive individuals.ConclusionThis ELISA appears to be a specific and reliable method for detecting COVID‐19 antibodies (IgG, IgM, and IgA), and a useful tool for identifying individuals which have developed immunity to the virus.     

Nucleocapsid protein of SARS‐CoV‐2 is a potential target for developing new generation of vaccine

BackgroundSARS‐CoV‐2 has spread worldwide causing more than 400 million people with virus infections since early 2020. Currently, the existing vaccines targeting the spike glycoprotein (S protein) of SARS‐CoV‐2 are facing great challenge from the infection of SARS‐CoV‐2 virus and its multiple S protein variants. Thus, we need to develop a new generation of vaccines to prevent infection of the SARS‐CoV‐2 variants. Compared with the S protein, the nucleocapsid protein (N protein) of SARS‐CoV‐2 is more conservative and less mutations, which also plays a vital role in viral infection. Therefore, the N protein may have the great potential for developing new vaccines.MethodsThe N protein of SARS‐CoV‐2 was recombinantly expressed and purified in Escherichia coli. Western Blot and ELISA assays were used to demonstrate the immunoreactivity of the recombinant N protein with the serum of 22 COVID‐19 patients. We investigated further the response of the specific serum antibodies and cytokine production in BALB/c mice immunized with recombinant N protein by Western Blot and ELISA.ResultsThe N protein had good immunoreactivity and the production of IgG antibody against N protein in COVID‐19 patients was tightly correlated with disease severity. Furthermore, the N protein was used to immunize BALB/c mice to have elicited strong immune responses. Not only high levels of IgG antibody, but also cytokine‐IFN‐γ were produced in the N protein‐immunized mice. Importantly, the N protein immunization induced a high level of IgM antibody produced in the mice.ConclusionSARS‐CoV‐2 N protein shows a great big bundle of potentiality for developing a new generation of vaccines in fighting infection of SARS‐CoV‐2 and its variants.     

Establishment of an in‐house real‐time RT‐PCR assay for the detection of severe acute respiratory syndrome coronavirus 2 using the first World Health Organization international standard in a resource‐limited country

BackgroundThe COVID‐19 pandemic caused by SARS‐CoV‐2 remains public health burdens and many unresolved issues worldwide. Molecular assays based on real‐time RT‐PCR are critical for the detection of SARS‐CoV‐2 in clinical specimens from patients suspected of COVID‐19.ObjectiveWe aimed to establish and validate an in‐house real‐time RT‐PCR for the detection of SARS‐CoV‐2.MethodologyPrimers and probes sets in our in‐house real‐time RT‐PCR assay were designed in conserved regions of the N and E target genes. Optimized multiplex real‐time RT‐PCR assay was validated using the first WHO International Standard (NIBSC code: 20/146) and evaluated clinical performance.ResultsThe limit of detection validated using the first WHO International Standard was 159 IU/ml for both E and N target genes. The evaluation of clinical performance on 170 clinical samples showed a positive percent agreement of 100% and the negative percent agreement of 99.08% for both target genes. The Kappa value of 0.99 was an excellent agreement, the strong correlation of C _t values observed between two tests with r ² = 0.84 for the E gene and 0.87 for the N gene. Notably, we assessed on 60 paired saliva and nasopharyngeal samples. The overall agreement was 91.66%, and Kappa value of 0.74 showed a high agreement between two types of samples. When using nasopharyngeal swabs as the reference standard, positive percent agreement, and negative percent agreement were 91.83% and 90.90%, respectively.ConclusionIn the present study, we established and validated an in‐house real‐time RT‐PCR for molecular detection of SARS‐CoV‐2 in a resource‐limited country.     

Establishing an internal quality control method for the stable extraction of nucleic acids of severe acute respiratory syndrome coronavirus 2 and RT‐PCR‐based detection

BackgroundSevere acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), the causative agent of the coronavirus disease 2019 (COVID‐19), is detected using real‐time RT‐PCR. However, there are limitations pertaining to quality control, particularly with respect to establishing quality control measures for extraction of viral nucleic acids. Here, we investigated the quality control measures for the various processes using an extrinsic quality control substance and quality control charts.MethodsAn extrinsic quality control substance was added to the sample, and then, real‐time RT‐PCR was performed. Samples with negative test results and the corresponding data were analyzed; a quality control chart was created and examined.ResultsData analysis and the quality control charts indicated that SARS‐CoV‐2 could be reliably detected using real‐time RT‐PCR, even when different nucleic acid extraction methods were used or when different technicians were employed.ConclusionWith the use of quality control substances, it is possible to achieve quality control throughout the process—from nucleic acid extraction to nucleic acid detection—even upon using varying extraction methods. Further, generating quality control charts would guarantee the stable detection of SARS‐CoV‐2.     

Neurological manifestations in patients with COVID‐19: A systematic review and meta‐analysis

IntroductionThe intensification of coronavirus disease 2019 (COVID‐19) complications, severe symptoms, and high mortality rate has led researchers to focus on this significant issue. While respiratory and cardiac complications have been described as high‐risk manifestations in patients with COVID‐19, neurological complications can also enhance mortality. This study aimed to evaluate the prevalence of neurological complications arises from SARS‐CoV‐2 and assess the mortality rate from neurological complications.Material and MethodsLiterature review was conducted by searching in PubMed/Medline, Web of Sciences, and Embase. After performing search strategies with relevant terms, a number of articles were excluded, including review articles, systematic review or meta‐analysis, duplicate publication of same researchers, congress abstracts, animal studies, case reports, case series, and articles reporting a history of neurological features prior to COVID‐19 infection. After retrieving the data, statistical analysis was performed using the STATA Version 14 software.ResultsFrom 4455 retrieved publications, 20 articles were selected for further analysis. Among 18,258 included patients, 2791 showed neurological symptoms, which were classified into different groups. Headache, confusion, and fatigue were reported as the most non‐specific neurological features in confirmed COVID‐19 patients. Psychiatric symptoms, CNS disorders, cerebrovascular disorders, CNS inflammatory disorders, PNS disorders, neuromuscular disorders, etc., were defined as specific neurological manifestations. The pooled prevalence of neurological manifestations and mortality rate of COVID‐19 patients with neurological features were estimated to be 23.0% (95% CI: 17.8–29.2) and 29.1% (95% CI: 20.3–39.8), respectively.ConclusionNeurological manifestations may commonly happen in patients with COVID‐19. This study reported a high prevalence of neurological complications and mortality rates in COVID‐19 patients. Therefore, patients with COVID‐19 who indicated neurological symptoms should be taken seriously and should receive early treatment to prevent undesirable events.     

Clinical and biochemical indexes of 11 COVID‐19 patients and the genome sequence analysis of the tested SARS‐CoV‐2

BackgroundAt present, SARS‐CoV‐2 epidemic in the world rapidly spread. It is a serious global public health emergency.MethodsIn this study, we described the clinical characteristics of 11 COVID‐19 patients hospitalized in the Meizhou People''s Hospital, and viral genome sequences of SARS‐CoV‐2 from these patients were analyzed.ResultsOf the 11 patients, six cases developed fever, 9 cases developed a cough, and two cases developed headache and chills. Four patients (36.4%) had underlying diseases. Pneumonia is the most common complication. The laboratory test results showed that there were no adult patients with increased lymphocyte/lymphocyte percentage (LYM/LYM%). Most patients had normal total protein (TP) and albumin (ALB), but only two patients had decreased. Most patients had increased or normal levels of erythrocyte sedimentation rate (ESR), C reactive protein (CRP), activated partial thromboplastin time (APTT), fibrinogen (FIB), creatine kinase isoenzymes (CK‐MB), and lactate dehydrogenase (LDH). Neutrophil (NEU) (r = 0.664, p = 0.026), CK‐MB (r = 0.655, p = 0.029) and blood urea nitrogen (BUN) (r = 0.682, p = 0.021) were significantly associated with SARS‐CoV‐2 virus cycle threshold (Ct) value. Multiple sequence alignment (MSA) shows that two different SNPs were identified at positions 8781 and 28144, and have a complete linkage genetic form of 8781C‐28144T and 8781T‐28144C.ConclusionsThe reports of the 11 COVID‐19 patients in our hospital will provide useful information for the diagnosis, treatment, and drug development of SARS‐CoV‐2.     

Performance comparison of two nucleic acid amplification systems for SARS‐CoV‐2 detection: A multi‐center study

BackgroundMany rapid nucleic acid testing systems have emerged to halt the development and spread of COVID‐19. However, so far relatively few studies have compared the diagnostic performance between these testing systems and conventional detection systems. Here, we performed a retrospective analysis to evaluate the clinical detection performance between SARS‐CoV‐2 rapid and conventional nucleic acid detection system.MethodsClinical detection results of 63,352 oropharyngeal swabs by both systems were finally enrolled in this analysis. Sensitivity (SE), specificity (SP), and positive and negative predictive value (PPV, NPV) of both systems were calculated to evaluate their diagnostic accuracy. Concordance between these two systems were assessed by overall, positive, negative percent agreement (OPA, PPA, NPA) and κ value. Sensitivity of SARS‐CoV‐2 rapid nucleic acid detection system (Daan Gene) was further analyzed with respect to the viral load of clinical specimens.ResultsSensitivity of Daan Gene was slightly lower than that of conventional detection system (0.86 vs. 0.979), but their specificity was equivalent. Daan Gene had ≥98.0% PPV and NPV for SARS‐CoV‐2. Moreover, Daan Gene demonstrated an excellent test agreement with conventional detection system (κ = 0.893, p = 0.000). Daan Gene was 99.31% sensitivity for specimens with high viral load (C _t < 35) and 50% for low viral load (C _t ≥ 35).ConclusionsWhile showing an analytical sensitivity slightly below than that of conventional detection system, rapid nucleic acid detection system may be a diagnostic alternative to rapidly identify SARS‐CoV‐2‐infected individuals with high viral loads and a powerful complement to current detection methods.     

Evaluation of automated molecular tests for the detection of SARS‐CoV‐2 in pooled nasopharyngeal and saliva specimens

BackgroundPooling of samples for SARS‐CoV‐2 testing in low‐prevalence settings has been used as an effective strategy to expand testing capacity and mitigate challenges with the shortage of supplies. We evaluated two automated molecular test systems for the detection of SARS‐CoV‐2 RNA in pooled specimens.MethodsPooled nasopharyngeal and saliva specimens were tested by Qiagen QIAstat‐Dx Respiratory SARS‐CoV‐2 Panel (QIAstat) or Cepheid Xpert Xpress SARS‐CoV‐2 (Xpert), and the results were compared to that of standard RT‐qPCR tests without pooling.ResultsIn nasopharyngeal specimens, the sensitivity/specificity of the pool testing approach, with 5 and 10 specimens per pool, were 77%/100% (n = 105) and 74.1%/100% (n = 260) by QIAstat, and 97.1%/100% (n = 250) and 100%/99.5% (n = 200) by Xpert, respectively. Pool testing of saliva (10 specimens per pool; n = 150) by Xpert resulted in 87.5% sensitivity and 99.3% specificity compared to individual tests. Pool size of 5 or 10 specimens did not significantly affect the difference of RT‐qPCR cycle threshold (C_T) from standard testing. RT‐qPCR C_T values obtained with pool testing by both QIAstat and Xpert were positively correlated with that of individual testing (Pearson''s correlation coefficient r = 0.85 to 0.99, p < 0.05). However, the C_T values from Xpert were significantly stronger (p < 0.01, paired t test) than that of QIAstat in a subset of SARS‐CoV‐2 positive specimens, with mean differences of −4.3 ± 2.43 and −4.6 ± 2 for individual and pooled tests, respectively.ConclusionOur results suggest that Xpert SARS‐CoV‐2 can be utilized for pooled sample testing for COVID‐19 screening in low‐prevalence settings providing significant cost savings and improving throughput without affecting test quality.     

Clinical manifestations,treatment options,and comorbidities in COVID‐19 relapse patients: A systematic review

IntroductionInterest revolving around coronavirus disease 2019 (COVID‐19) reinfection is escalating rapidly. By definition, reinfection denotes severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2), PCR redetection, and COVID‐19 recurrence within three months of the initial symptoms. The main aim of the current systematic review was to evaluate the features of COVID‐19 relapse patients.Materials and methodsFor this study, we used a string of terms developed by a skilled librarian and through a systematical search in PubMed, Web of Science, and Embase for eligible studies. Clinical surveys of any type were included from January 2019 to March 2021. Eligible studies consisted of two positive assessments separated by a negative result via RT‐PCR.ResultsFifty‐four studies included 207 cases of COVID‐19 reinfection. Children were less likely to have COVID‐19 relapse. However, the most patients were in the age group of 20–40 years. Asthenia (66.6%), headache (66.6%), and cough (54.7%) were prevalent symptoms in the first SARS‐CoV‐2 infection. Asthenia (62.9%), myalgia (62.9%), and headache (61.1%) were most frequent in the second one. The most common treatment options used in first COVID‐19 infection were lopinavir/ritonavir (80%), oxygen support (69.2%), and oseltamivir (66.6). However, for the treatment of second infection, mostly antibiotics (100%), dexamethasone (100%), and remdesivir (80%) were used. In addition, obesity (32.5%), kidney failure (30.7%), and hypertension (30.1%) were the most common comorbidities. Unfortunately, approximately 4.5% of patients died.ConclusionWe found the potency of COVID‐19 recurrence as an outstanding issue. This feature should be regarded in the COVID‐19 management. Furthermore, the first and second COVID‐19 are similar in clinical features. For clinically practical comparison of the symptoms severity between two epochs of infection, uniform data of both are required. We suggest that future studies undertake a homogenous approach to establish the clinical patterns of the reinfection phenomena.