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Objective: The aim of the study was to compare the efficacy of Clevira tablets in Human adult patients, with Influenza A&B and Vital flu. Methods: This study was an open label, balanced, randomized, multi-dose, two-treatment, parallel, and comparative Phase III clinical trial to determine the safety and efficacy of Clevira Tablets. Twenty patients were enrolled and received Clevira Tablet along with Standard Treatment for Influenza A&B and other respiratory viral infections. Enrollment was based on the diagnosis of hematology, biochemistry, serology, RT-PCR, and chest X-ray and inclusion, and none of the exclusion criteria and included in the study. Results: All the patients demonstrated safety measures with respect to blood pressure and pulse rate. Furthermore, statistically significant (P < 0.0001) improvement showed in temperature from baseline (102.03 ± 0.64) and at the end of the study period (98.14 ± 0.70). Conclusion: The study demonstrated an expedited clinical cure with normal vital signs and hematological results which validated that Clevira is safe and efficacious in patients with Influenza A&B and Viral flu. The data further entrusted that Clevira can be used in infected patients with Influeza A&B and Viral Flu, and relieve the signs and symptoms, with a rapid recovery, without any adverse side effects.
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Background: It is well-recognized that uncontrolled glycemia reflects the severity and mortality rates of respiratory virus epidemics. The aim of the study was to report the features and course of therapy of diabetic individuals admitted to a tertiary care facility with COVID-19 infection. Additionally, we tried to assess how hyperglycemia affected the clinical results. Methods: This study used observational methods to conduct a retrospective chart review of 125 cases from October 2021 to March 2022 at a single center. Results: Males made up 94.6% of the 125 examined cases. Within the age range of 21 to 78 years, the study group's average age was 49.6�.4 years. Of the patients, 66.4% had prior knowledge of diabetes. When compared to pre-existing diabetes individuals with the newly diagnosed diabetes individuals, the latter had a higher death rate (p=0.03) and needed mechanical breathing (p=0.02). Conclusions: Hyperglycemia that is uncontrolled harms COVID-19-infected patients. There is an increased risk of hyperglycemia problems that have been discovered or unknown.to screen for cases of undiscovered diabetes in hospital patients, which may be especially relevant during the COVID-19 pandemic, optimal glycemia optimization is essential.
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Background@#The Panbio™ COVID-19 Ag Rapid Test is a Food and Drug Administration (FDA)-approved point-of-care test (POCT) used for SARS-CoV-2 detection which has met minimum sensitivity and specificity requirements by the World Health Organization (WHO).@*Objective@#The study aimed to compare the clinical performance of a commercial lateral flow assay (LFA) to reverse transcriptase polymerase reaction (RT-PCR) in SARS-CoV-2 infection diagnosis@*Methodology@#Clinical data and simultaneous LFA and RT-PCR samples collected from June 2021 to June 2022 were obtained to analyze the diagnostic accuracy of LFA compared to RT-PCR.@*Results@#A total of 265 samples was obtained. 34.45% of RT-PCR positive samples were reliably detected by LFA. COVID-19 was reliably ruled out by LFA in 99.32% RT-PCR negative samples. LFA sensitivity among symptomatic patients with ≤7 days of illness was 51.61%, slightly higher than those with >7 days of illness (18.92%), and significantly higher than asymptomatic patients (16.67%). Asymptomatic subjects have a varied range of Ct-values, indicating different stages of infection or viral loads. Individuals with symptoms for more than 7 days have higher Ct-values, suggesting they are in later stages of infection or have lower viral loads. The probability of a positive LFA result decreases significantly when the Ct-value is beyond 28-30.@*Conclusion@#The LFA evaluated in this study did not show significant sensitivity and specificity during the early disease course wherein viral loads are suggestively high. However, its utility to accurately rule out COVID-19 is quite reliable in subjects with symptoms that are >7 days since Ct-values are suggestively beyond 28-30 which implies a significantly decreased probability of a positive LFA result.
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COVID-19 , SARS-CoV-2ABSTRACT
Objectives@#In the Philippines, patients on chronic hemodialysis with COVID-19 remain admitted in hospitals despite clinical recovery because most free-standing dialysis units require proof of negative conversion via Reverse Transcriptase – Polymerase Chain Reaction (RT-PCR). This study aims to determine the time to negative conversion of COVID-19 RT-PCR testing among adult patients on chronic hemodialysis with COVID-19 admitted at the Philippine General Hospital (PGH) and bring insight in using the symptom or time-based procedure as recommended by local guideline, and ultimately, to ensure delivery of adequate hemodialysis despite being infected with COVID-19, shorten isolation period, and conserve resources especially in resource-limited settings.@*Methods@#This is a retrospective cohort study on all adult patients on chronic hemodialysis who were admitted in PGH after the diagnosis of COVID-19 by RT-PCR between March 2020 and February 2021. Descriptive statistics was used in summarizing the data.@*Results@#A total of 90 patients on chronic hemodialysis who tested positive for COVID-19 via RT-PCR admitted at PGH were included in the study. Most of these patients had moderate COVID-19 at 53.3%. The median number of days from onset of symptoms to clinical recovery was 14.5 days. The median time to first negative conversion was 18 days. Most of these patients had negative conversion at the second week. The correlation coefficient between time to clinical recovery and negative conversion was 0.214. @*Conclusion@#Among adult patients on chronic hemodialysis who were admitted in PGH after the diagnosis of COVID-19, the time to negative conversion was longer compared to the time to clinical recovery with a very weak correlation between the two.
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COVID-19 , Renal DialysisABSTRACT
@#COVID-19, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), remains a global health threat. Timely identification of infected cases is important for appropriate patient management and the control of viral spread. Simple and cost-effective tests are required to increase access to testing and early case detection. Here, we describe a colorimetric reverse transcription-loop-mediated isothermal amplification (RT-LAMP) method to detect SARS-CoV-2. The RT-LAMP could amplify the orf1ab sequence detectable by visual color change within 45 min at 63 °C. The limit of detection (LoD) for SARS-CoV-2 RNA was less than 100 copies (13.36) per reaction with no cross-amplification with other related viruses. Clinical evaluation using leftover RNA samples extracted from 163 nasopharyngeal swab specimens showed perfect agreement in negative (n = 124) and positive samples with cycle thresholds (Ct) < 34 cycles (n = 33) detected by real-time reverse transcription-polymerase chain reaction (RT-PCR), targeting RdRp and N genes as a reference. Overall, the diagnostic accuracy, sensitivity, specificity, positive and negative predictive values of RT-LAMP in testing were 96.32% (95% CI: 92.16-98.64%), 84.62% (95% CI: 68.47-94.14%), 100% (95% CI: 97.07-100.0%), 100% (95% CI: 89.42-100.0%), and 95.38% (95% CI: 90.22-98.29), respectively. This RT-LAMP assay is simple and reliable, with the potential to be an alternative for the rapid detection of SAR-CoV-2 with minimal time and fewer resources compared to real-time RT-PCR.
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Objectives:As a part of an ongoing research programme on vector-borne viral diseases especially Dengue, a retrospective analytical study on the occurrence and distribution of Dengue virus serotype(s) in the state of Manipur – a small state situated in the northeastern region of the Indian subcontinent was carried out at Viral Research & Diagnostic Laboratory (VRDL), Department of Microbiology, Regional Institute of Medical Sciences (RIMS), Imphal which is a tertiary care hospital.Materials & Methods:A total of 914 blood samples from clinically dengue-suspected patients were screened for the presence of Dengue infection by adopting the ELISA (IgM) technique during the period from 01/06/2022 to 02/12/2022. Further, anti-Dengue IgM antibody-positive samples having high Optical Density (OD) value(s) were selected and subjected to RT PCR to determine the serotype(s) of the Dengue virus.Results:Of the 914 blood samples examined for the presence of Dengue infection, 111 (12.14%) were found positive for anti- Dengue IgM antibody indicating acute infection of Dengue virus.Of the positive patients, there were 56 (50.45%) males and 55 (49.54%) females.Predominant clinical features observed among the Dengue-confirmed patients included – fever (74%), headache (19%), arthralgia / joint pain (9%), myalgia (6%) and vomiting (6%) respectively.The study revealed that while the circulation of three (03) Dengue virus serotypes, namely DENV – 1, DENV – 2 & DENV – 3 were observed in the Tengnoupal district,the circulation of two Dengue virus serotypes i.e., DENV – 1 & DENV – 2 was evident in Imphal West & Bishnupur districts respectively.The present study also reveals the occurrence of Dengue virus serotype – 1 (DENV - 1) in Churachandpur district. Conclusion:The present study reveals the circulation/distribution of three Dengue virus serotypes namely, DENV – 1, DENV – 2 and DENV – 3 among the studied samples.
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Background: Genital herpes is caused predominantly by Herpes simplex virus type 2 (HSV-2) and less commonly by HSV-1. Genital HSV-1 infection results from oral sex, with fewer recurrences, mild symptoms, less asymptomatic shedding and poor genital transmission. The present study was undertaken to identify frequency of occurrence of HSV-1 and HSV-2 in genital herpes by microscopy, serology and Real Time Polymerase Chain Reaction (RT-PCR). Methods: Genital ulcer swabs and serum were collected at regional STI centre, Govt Medical College and Hospital, Nagpur. A total of 53 patients of Genital Ulcer Disease from December 2020 -22 were examined for etiology by microscopy, serum IgM and IgG against HSV-1 and 2 by ELISA and HSV-1 and 2 DNA by RT- PCR. Results: Out of 53 genital swabs processed, 6 (11.3%) and 28 (52.8%) samples were positive for HSV-1 and 2 DNA respectively. Of the 6 HSV-1 DNA positive samples, seropositivity for HSV-1 IgM was in 2 (33.3%) samples and for HSV-1 IgG in 4 (66.7%) samples. Of the 28 HSV-2 DNA samples, HSV-2 IgM was positive in 4 (14.3%) samples and HSV-2 IgG was positive in 7 (25%) samples, multi nucleated giant cells were seen in 2 (7.14%) samples. The remaining 15 (53.6%) HSV-2 DNA positive samples were seronegative. Conclusions: HSV-1 was detected in 6 (11.3%) samples. Though these genital ulcers may be mild, it is important to counsel the patients for abstinence or safe sex practices to prevent their partners from acquiring painful non-genital ulcers due to HSV-1.
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Background: Initial wave of COVID-19 created a massive health crisis everywhere including India due to a limited understanding of the disease pathology. Most physicians used sepsis as a prototype to manage COVID-19, as there are similarities. Heat-killed Mycobacterium w (Mw) (inj. sepsivac®) is a known immunomodulator approved for the treatment of gram-negative sepsis. Our purpose of this observation is to evaluate the safety and efficacy of Inj sepsivac in COVID-19 patients along with the standard of care. Methods: Total 49 patients data with reverse transcriptase-polymerase chain reaction (RT-PCR) confirmed critically ill COVID-19 patients who were admitted at Velocity Hospital, Surat between May 4, 2021, and May 18, 2021 were evaluated. They were evaluated for vital parameters like pulse, blood pressure, respiratory rate and temperature as well as laboratory parameters like ALT, S IL-6, serum creatinine and CRP during three follow-up visits after the administration of Inj sepsivac. Further follow-up was done until the discharge/death of the patient. Results: There was a statistically significant reduction of mean CRP observed compared to the baseline value during all follow-up visits. The rest of the laboratory parameters as well as clinical assessment did not show any significant change as compared to baseline. Out of 49, two patients died (mortality rate; 4%). Inj. sepsivac was found to be well tolerated without any systemic side effects. Conclusions: The addition of Mw to the standard of care can improve laboratory parameters like CRP, without any safety concerns. These results should be further substantiated by larger randomised clinical trials.
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Abstract The rocketing number of COVID-19 cases highlighted the critical role that diagnostic tests play in medical and public health decision-making to contain and mitigate the SARS-CoV-2 pandemic. This study reports the evaluation and implementation of different tests for the molecular detection of SARS-CoV-2 in the central region of Argentina. We evaluated 3 real time RT-PCR kits (GeneFinder COVID-19 Plus RealAmp Kit, DisCoVery SARS-CoV-2 RT-PCR Detection Kit and WGene SARS-CoV-2 RT Detection), 2 nucleic acid extraction methods [MagaBio plus Virus DNA/RNA Purification Kit II (BioFlux), 35-min vs. 9-min, a pre-analytical reagent (FlashPrep®) and 2 isothermal amplification tests (Neokit Plus and ELA CHEMSTRIP®). The order according to the best performance of the 3 real-time RT-PCR kits evaluated was: DisCoVery > GeneFinderTM> WGene. The 2 RNA extraction methods showed similar good results: MagaBio plus Virus RNA Purification Kit II (BioFlux) 9-min was selected due to its faster performance. FlashPrep® reagent showed excellent results to perform direct RNA detection. Isothermal amplification assays showed acceptable sensitivity and specificity values (>80%), except in samples with Ct> 30. Our data show optimal real time RT-PCR kits and alternative molecular methods for SARS-CoV-2 diagnostic. These alternative assays proved to be aceptable.
Resumen La explosión de casos de COVID-19 resaltó el papel fundamental que desempeñan las pruebas de diagnóstico en la toma de decisiones médicas y de salud pública para contener y mitigar la pandemia de SARS-CoV-2. Este estudio reporta la evaluación y la implementación de diferentes test para la detección molecular de SARS-CoV-2 en la región central de Argentina. Evaluamos tres kits de RT-PCR en tiempo real (GeneFinder COVID-19 Plus RealAmp Kit, DisCoVery SARS-CoV-2 RT-PCR Detection Kit y WGene SARS-CoV-2 RT Detection), dos métodos de extracción de ácidos nucleicos (MagaBio plus Virus DNA/RNA Purification Kit II [BioFlux, 35-min vs. 9-min), un reactivo pre-analítico (FlashPrep®) y dos test de amplificación isotérmica (Neokit Plus and ELA CHEMSTRIP®). El orden de rendimiento de los tres kits de RT-PCR en tiempo real evaluados fue el siguiente: DisCoVery GeneFinder™ WGene. Los dos métodos de extracción de RNA mostraron buenos y similares resultados; se seleccionó MagaBio plus Virus RNA Purification Kit II (BioFlux) 9-min debido a su rápido tiempo de procesamiento. El reactivo FlashPrep® mostró excelentes resultados para realizar detección directa de RNA. Los ensayos de amplificación isotérmica mostraron valores de sensibilidad y de especificidad aceptables (80%), excepto en muestras con Ct 30. Nuestros resultados muestran kits de RT-PCR en tiempo real óptimos, como así también métodos moleculares alternativos para el diagnóstico de SARS-CoV-2 que resultan aceptables para su uso en contextos adversos, de descentralización y en diferentes escenarios epidemiológicos, para la detección rápida y precisa del SARS-CoV-2.
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Introducción. La pandemia por COVID-19 ha puesto de manifiesto la necesidad de pruebas diagnósticas rápidas. La prueba de referencia es la reacción en cadena de la polimerasa en tiempo real (RT-PCR). Requiere un equipo y personal capacitado, y su resultado puede llevar un tiempo de espera prolongado. El sistema BD Veritor® es el método rápido cromatográfico utilizado para la detección del antígeno del coronavirus de tipo 2 del síndrome respiratorio agudo grave, en individuos sintomáticos. El objetivo primario del siguiente trabajo es evaluar sensibilidad y especificidad del test de antígeno (TA) comparadas con la RT-PCR en población pediátrica. Población y métodos. Estudio prospectivo, de prueba diagnóstica. Se incluyó a todo menor de 17 años en los primeros 5 días de inicio de síntomas, que consultó desde julio de 2021 hasta febrero de 2022. Se calculó un mínimo de 300 muestras para lograr una precisión de ± 8,76 % y de ± 3,68 % para sensibilidad y especificidad respectivamente. Se analizaron en paralelo las muestras por ambas metodologías. Resultados. De 316 muestras pareadas, 33 fueron positivas por ambos métodos; 6 fueron positivas solo por RT-PCR. La especificidad del TA fue del 100 %; la sensibilidad, del 84,6 %, con un valor predictivo positivo y negativo del 100 % y del 98 % respectivamente. Conclusiones. El TA demostró ser útil en el diagnóstico de pacientes pediátricos con COVID-19 en los primeros 5 días de inicio de síntomas, aunque aquellos con TA negativo y alta sospecha clínica deberían confirmar su resultado con la RT-PCR.
Introduction. The COVID-19 pandemic has brought to light the need for rapid diagnostic tests. The gold standard test is reverse transcription-polymerase chain reaction (RT-PCR). RT-PCR requires equipment and trained personnel, and results may take a long waiting time. The BD Veritor® System is a rapid chromatographic method used for the detection of severe acute respiratory syndrome coronavirus 2 antigen in symptomatic individuals. The primary objective of this study is to assess the sensitivity and specificity of the antigen test (AT) compared to the RT-PCR in the pediatric population. Population and methods. Prospective study with a diagnostic test. All children younger than 17 years in the first 5 days of symptom onset, who consulted between July 2021 and February 2022, were included. A minimum of 300 specimens was estimated to achieve an accuracy of ±8.76% and ±3.68% for sensitivity and specificity, respectively. Specimens were analyzed in parallel using both methodologies. Results. Of 316 paired samples, 33 were positive by both methods; 6 were positive only by RT-PCR. The specificity of the AT was 100%; sensitivity was 84.6%, with a positive and negative predictive value of 100% and 98%, respectively. Conclusions. The AT proved to be useful in the diagnosis of pediatric patients with COVID-19 in the first 5 days of symptom onset, although those with a negative AT and high clinical suspicion should confirm their result with a RT-PCR.
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Humans , Infant, Newborn , Infant , Child, Preschool , Child , Adolescent , COVID-19/diagnosis , Prospective Studies , Sensitivity and Specificity , Reverse Transcriptase Polymerase Chain Reaction , Reverse Transcription , Pandemics , COVID-19 Testing , SARS-CoV-2ABSTRACT
A humanidade foi impactada por uma Pandemia que expôs a população ao contato com um vírus de elevado contágio e com o índice de letalidade alarmante. Este estudo objetivou avaliar a possibilidade da persistência de material genético do SARS-CoV-2 na superfície dos equipamentos de estabelecimentos de prática de atividades físicas indoor e outdoor. Foram coleta das amostras de equipamentos utilizados para a prática de exercícios físicos em cinco academias, cinco praças e entre os frequentadores desses ambientes. Aplicou-se a técnica RT-PCR para a detecção doRNA do SARS-CoV-2 e posterior análise dos resultadose foi detectada a existência de partículas de RNA viral do SARS-CoV-2 em 48,57% das amostras coletadas dos equipamentos das academias e 12,85% das amostras coletadas nas praças, evidenciando uma incidência menor em equipamentos utilizados em locais abertos em todas as áreas comparadas.Além disso, constatou-se que 35,7% dos participantes do estudo testaram positivo para COVID-19.Os casos positivos para COVID-19 detectados apresentaram sintomas classificados como levesa moderados e uma recuperação rápida.A presença de material genético nos equipamentos,por sua vez, leva-nos a perceber a importância da higienização adequada das superfícies, como forma de prevenção.
Humanity was impacted by a Pandemic that exposed the population to contact with a highly contagious virus with an alarming lethality rate. The present study aimed to evaluate the possibility of the persistence of genetic material from SARS-CoV-2 on the surface of equipment used to practice indoor and outdoor physical activities. A sample of equipment used for the practice of physical activity was collected in five gyms and five squares and among the regulars of these environments. The RT-PCR technique was applied to detect the RNA of SARS-CoV-2 and subsequent analysis of the results. The existence of SARS-CoV-2 viral RNA particles was detected in 48.57% of the samples collected from gym equipment and 12.85% of the samples collected in squares, evidencing a lower incidence in equipment used in open spaces in all areas compared and it was found that 35.7% of the study participants tested positive for COVID-19. The positive cases for COVID-19 detected, had symptoms classified as mild to moderate and a quick recovery. The presence of genetic material in the equipment, in turn, leads us to realize the importance of proper cleaning of surfaces, as a form of prevention.
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Background: In children and adolescents, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is mostly responsible for mild respiratory symptoms, in contrast with severe forms reported in adults. An association between the disease caused by SARS-CoV-2, coronavirus disease 2019 (COVID-19), and late manifestations of vasculitis has been increasingly suspected.Method: All children and adolescents (aged ?18 years) who were diagnosed with Kawasaki disease (KD) and KD shock syndrome (KDSS) as per AHA and Kanegaye et al criteria. From each patient we obtained at least two nasopharyngeal swabs to test for SARS-CoV-2 using reverse transcription- polymerase chain reaction (RT-PCR). We also took blood samples to test for IgG antibodies against SARS-CoV-2. Results: Leucocytosis was found in majority with median leucocyte count of 10100 predominant neutrophila. Inflammatory markers D dimer, serum ferritin and fibrinogen level were raised. The 43% patients had coronary artery aneurysm. The 75% patient had some form of Left ventricular systolic dysfunction. All patients received IVIG while 13 patients received both IVIG and methyl predinisolone. Ionotropes were used in 68%. Two patients received tocluzumab.Conclusions: In this study we found increased incidence of Kawasaki like illness temporally associated with COVID-19. Older age of presentation with more atypical presentation.
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COVID-19 patients commonly present with lower respiratory symptoms with other systemic involvement. Haematological manifestation such as low haemoglobin, thrombocytopenia, lymphocytopenia also common in COVID19 patients. In this study, we investigated prevalence, association with serum ferritin in post COVID-19 anaemic patients, after human umbilical cord blood transfusion in relation to control group. Among 155 COVID-19 RT-PCR positive patients 36 (23%) was anaemic. In our study 18 patients was transfused human umbilical cord blood, 12 patients were treated with haematinics and 6 patients denied taking any of the above. In most cases anaemia was moderate to severe that may be due to inflammation or due to pre-existing iron deficiency.Umbilical cord blood transfusion to post COVID -19 patients for the treatment of anaemia because of the unique composition of UCB. Haematological analysis and serum ferritin estimation reflecting the treatment out come in post COVID-19 anaemic patients. There was a difference between the dependent variable's serum ferritin (p <.001) in anaemic COVID-19 patients. In conclusion, our result highlight serum ferritin is widely used in diagnosis and monitoring of COVID-19 disease.
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Background & objectives: High transmissibility of the SARS-CoV-2 has significant implications on healthcare workers’ safety, preservation, handling, transportation and disposal of the deceased bodies. The objective of this study was to detect SARS-CoV-2 antigen in nasopharyngeal samples and its implications in handling and care of COVID-19 deceased bodies. Methods: A study was conducted at a dedicated COVID-19 centre on deceased individuals from April to December 2020. Rapid antigen test (RAT) and reverse transcription (RT)-PCR was compared on all the SARS-CoV-2 positive cadavers recruited in the study. Results: A total of 115 deceased individuals were included in the study. Of these, 79 (68.7%) were male and 36 (31.3%) were female and majority were in the age group of 51-60 yr [31 (27%)]. SARS-CoV-2 antigen test was positive in 32 (27.8%) and negative in 83 (72.1%) individuals. The mean time interval between deaths to the sample collection was 13.2 h with interquartile range of eight to 20 h. Reverse transcription (RT)-PCR was used as the reference test and 24 (20.9%) cases were true positive; 93.6 per cent [95% confidence interval (CI) 88.8-98.4%] sensitivity, 45.2 per cent (95% CI 35.5-55%) specificity, 60.2 per cent (95% CI 50.6-69.8%) positive predictive value and 88.8 per cent (95% CI 82.7-95%) negative predictive value of antigen test was computed. Interpretation & conclusions: SARS-CoV-2 antigen test was positive beyond 19 h in COVID-19 deceased individuals. Antigen test was found to be highly sensitive in the deceased. Patients, suspected of having died due to COVID-19, can be screened by this method. As infectiousness of the virus in the deceased bodies cannot be directly concluded from either the antigen or RT-PCR test, yet possible transmission cannot be completely ruled out. Strict infection control measures need to be followed during the handling and clearance of COVID-19 cadavers.
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Objetivo: Optimizar el control interno de calidad de RT-PCR en tiempo real para detección cualitativa de SARS-CoV-2, utilizando los valores Cq de controles negativos y positivos. Material y método: Estudio prospectivo-longitudinal. La muestra estuvo constituida por 143 valores Cq para los controles negativos de alicuotado y extracción, así como para el control positivo. Se analizó la distribución normal de los valores Cq mediante la prueba de Anderson-Darling (AD) y se aplicaron pruebas de aleatoriedad. Se calculó límites de control a partir de 51 valores Cq, para luego, mediante gráficas de control, monitorizar 92 valores Cq obtenidos desde noviembre del 2020 hasta marzo del 2021. Se evaluó aceptación de lote e índices Cpk como indicadores de optimización. Los cálculos se hicieron con el programa Minitab. Resultados: Se aceptaron los lotes de valores Cq y se obtuvieron índices Cpk superiores a 1.33 para los tres tipos de control. Discusión: No existen estudios publicados que apliquen control estadístico de calidad a la detección cualitativa de SARS-CoV-2. Conclusiones: Es posible utilizar los valores Cq de los controles para optimizar el control interno de calidad de RT-PCR en tiempo real para detección cualitativa de SARS-CoV-2, como si se tratara de una técnica de tipo cuantitativo.
Objective: To optimize the internal quality control of real-time RT-PCR for the qualitative detection of SARS-CoV-2, using the Cq values of negative and positive controls. Material and method : Prospective-longitudinal study. The sample consisted of 143 Cq values for the negative aliquot and extraction controls, as well as for the positive control. The normal distribution of Cq values was analyzed using the Anderson-Darling (AD) test and randomness tests were applied. Control limits were calculated from 51 Cq values, and then, using control charts, to monitor 92 Cq values obtained from November 2020 to March 2021. Lot acceptance and Cpk indices were evaluated as optimization indicators. The calculations were made with the Minitab program. Results: The batches of Cq values were accepted and Cpk indices higher than 1.33 were obtained for the three types of control. Discussion : There are no published studies that apply statistical quality control to the qualitative detection of SARS-CoV-2. Conclusions : It is possible to use the Cq values of the controls to optimize the internal quality control of real-time RT-PCR for qualitative detection of SARS-CoV-2, as if it were a quantitative technique.
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Objective To investigate the gene expression of sigma factors in vivo, and to explore the sigma factors that may be closely related to the virulence of pathogenic Mycobacterium tuberculosis Methods Tuberculosis (TB) patients diagnosed in the outpatient department of Tianjin Tuberculosis Control Center from January to December 2018 were selected, and 20 sputum-positive specimens were randomly selected from TB patients confirmed with Xpert-positive for the present study. Two immediate sputum specimens were collected from each case of pulmonary tuberculosis before treatment, one for RNA extraction and one for in vitro culture. In vitro cultured strains in the logarithmic phase of growth were harvested for RNA extraction. The specific primers for 13 sigma factors were designed. The differential expression of the 13 sigma factors between sputum isolates and in vitro cultured strains was analyzed by fluorescence quantitative PCR. Taking ribosomal 16s as the reference gene, the transcription level of sigma factors was analyzed by 2ΔCt. Using the stably expressed sigA as the control reference, the expression differences of other sigma factors were analyzed by one-way ANOVA. Results Within 0 days, stress-associated sigma factors have a different expression profile in clinical isolate strains vs H37Rv or in vitro. All the sigma factors induced up regulation in sputum ,while no difference transcription between clinical isolate strains vs H37Rv（P＞0.05）. When compared to in vitro culture ,only sigM transcript highest in sputum(P＜0.05). Conclusion SigM plays an important role in the initial stages of bacterial infection, but its exact role is unclear.We assumed it could have a role in the interplay between the host immune defenses and the bacterial escape mechanisms.
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Since the outbreak of the COVID-19 global pandemic in 2019, monitoring COVID-19 infection status and trend through wastewater, known as wastewater-based epidemiology (WBE), has been widely used in many countries and regions. WBE consists of five steps:wastewater sample collection, viral concentration, viral nucleic acid extraction, quantification of virus using quantitative RT-PCR, and dissemination of the wastewater surveillance results. This method could be used for early warning of COVID-19 outbreak in a population, monitoring COVID-19 distributions and epidemic trend, prediction of COVID-19 prevalence rate, understanding of temporal trend of SARS-COV-2 variants, and simultaneous surveillance of multiple pathogens. WBE and clinical surveillance can be used concurrently and the former is a good complement to the latter.
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@#Abstract: Objective To develop a real-time fluorescent quantitative RT-PCR (qRT-PCR) method for qualitative and quantitative Chikungunya virus (CHIKV) analysis. Methods Based on the systematic analysis of the genomic sequences of Chikungunya and its related arboviruses, the specific nucleic acid sequences for Chikungunya virus were screened and identified, and then the primers and TaqMan probe were designed. Meanwhile, the human GAPDH gene was used as an internal reference. The reaction system for qRT-PCR was systematically optimized by L9(34) orthogonal design, and a rapid detection method for Chikungunya by qRT-PCR based on TaqMan probe methods was established. The sensitivity, specificity, reproducibility, and coverage of the established method were analyzed in detail. The standard curve was made, and the absolute quantitative method was established using the cloned nucleic acid fragments as positive samples. Results A real-time fluorescent quantitative RT-PCR assay was developed for the qualitative and quantitative analysis of Chikungunya virus. The reaction system included Chikungunya virus and reference internal gene specific primers and probe, RT/Taq enzyme mixture, reaction buffer, and negative and positive reference. The established method obtained positive results with the ROSS strain of ECSA subtype, LR2006 strain of IOL branch, 181/25 strain of Asian type and Dongguan 2010 epidemic strains of Chikungunya virus, but there was no cross-reaction with other 18 arboviruses belonging to Flaviviruses, Alphaviruses and Bunyavirus. The minimum detection limit of the established method was 5.80 copies/mL, and a linear relationship was observed between the amount of input plasmid DNA and fluorescence signal value over a range of 5.80×102 copies/mL to 5.80×1010 copies/mL, and the correlation coefficient was 0.999 5. The qRT-PCR amplification efficiency was 91%, and the intra-assay variations and inter-assay variations were 0.01-0.07 and 0.03-0.11, respectively. Conclusions The TaqMan qRT-PCR method developed in this study can qualitatively and quantitatively detect Chikungunya virus rapidly with specificity and sensitivity, providing a technical method for the prevention and control of this viral disease.
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@#Abstract: Objective To establish the duplex TaqMan RT-PCR method for detection of Entamoeba histolytica and Giardia lamblia in fecal samples. Methods Primer pairs and probes for Entamoeba histolytica and Giardia lamblia were designed and duplex TaqMan RT-PCR amplification system was constructed. PCR products were inserted into the pUC57 plasmid, and the lower limit of detection of the method was determined. Clinical stool samples were tested in order to evaluated the efficacy of the method. Results The detection limits of duplex TaqMan RT-PCR were 31.6 copies/μL for Entamoeba histolytica and 32 copies/μL for Giardia lamblia, respectively. Of the total of 212 clinical stool samples tested, all 3 samples with E. histolytica-positive patients by microscopy were positive by PCR, while 1 from 209 samples with E. histolytica-negative patients by microscopy were positive by PCR, and the remaining samples were negative. For Giardia lamblia, all 8 samples positive by microscopy were positive by PCR, and 1 from 204 sample with a microscopy-negative patient was positive by PCR, and the remaining samples were negative. The amplification product sequencing and blast analysis were used to confirm that the amplified sequence in the specimen of a patient with negative microscopy but positive PCR belongs to the targeted pathogen, supported by clinical symptoms and laboratory test results. PCR results for other diarrhea-causing pathogens were negative, indicating no cross-reactivity. Conclusions The dual TaqMan RT-PCR method developed in this study can not only detect microscopy-positive samples of Entamoeba histolytica and Giardia lamblia but also can detect samples that were missed by microscopy, with higher sensitivity than the microscopy method. Further, this detection method does not cross-react with other diarrhea-causing pathogens, including cross-react with other diarrhea-causing pathogens including Iodamoeba butschlii, Blastocystis hominis, Plesiomonas, Aeromonas, Salmonella, Shigella, Sphaerozoum fuscum, and Entamoeba hartmani, thus has a good specificity.
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Aims@#Strawberry (Fragaria ananassa Duch.) is a member of the family Rosaceae and one of the most important vegetable crops in Egypt for local fresh consumption and exportation. Suspected symptoms of Strawberry mild yellow edge virus (SMYEV) on naturally infected strawberry plants were observed in Al Qaluobia Governorate. The main goal of this study was to develop a novel eco-friendly biogenic silver nanoparticle using clove (Syzygium aromaticum) (SzAgNPs) aqueous extract to control viral infection.@*Methodology and results@#Mechanically inoculated sixteen plant species representing seven families, Apocynaceae, Chenopodiaceae, Cucurbitaceae, Fabaceae, Moraceae, Rosaceae and Solanaceae, were used as hosts for SMYEV. Systemic mosaic, chlorotic local lesions, vein clearing, marginal chlorosis, marginal necrosis, yellowing and vein banding were observed 7 to 20 days post-inoculation. Transmission electron microscopy (TEM) of diseased tissues revealed severe degeneration of the chloroplasts and nucleus structures and the formation of cell wall protrusions. Sz-AgNPs were characterized using UV-Vis spectroscopy, TEM, dynamic light scattering (DLS) and Fourier transform infrared (FTIR). Foliar application of Sz-AgNPs (200 µg/mL) 24 h post and/or concurrently with SMYEV inoculation dropped the infection rate by 90% and 55%, respectively, whereas it was only reduced by 35% when applied 24 h prior to viral inoculation compared to control groups. DAS-ELISA and RT-PCR verified SMYEV inhibition (75-90%) in all plants treated with 200, 150 and 100 µg/mL Sz-AgNPs 24 h post-viral inoculation. Complete viral eradication was attained by applying Sz-AgNPs at a concentration of 200 μg/mL 24 h post-virus inoculation. Moreover, the total phenols, indoles, Vitamin C, total flavonoids and citric acid contents were not significantly affected by Sz-AgNPs treatment compared to healthy control groups.@*Conclusion, significance and impact of study@#In conclusion, Sz-AgNPs suppressed SMYEV by 75-90% when applied 24 h post-virus inoculation. Eco-friendly Sz-AgNPs could be used for controlling viral infections and avoiding the rejection of exportable strawberries due to the use of harmful pesticides.