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1.
Chest ; 162(4):A625, 2022.
Article in English | EMBASE | ID: covidwho-2060650

ABSTRACT

SESSION TITLE: Unusual Pneumonias SESSION TYPE: Rapid Fire Case Reports PRESENTED ON: 10/19/2022 12:45 pm - 1:45 pm INTRODUCTION: Cytomegalovirus (CMV) is an important infectious organism in the morbidity and mortality of immunocompromised patients. CMV is a known cause of pneumoina in transplant patients, such as lung transplant recepients. Pneumocytis Jiroveci Pneumonia (PCP) is also a known risk factor for potentially life-threatening infections in immunocompromised patients. In this , we are presenting a rare case of an immunocompromised patient who had penumonia caused by a concurrent CMV and PCP infections. CASE PRESENTATION: A 53 year-old female patient with history of Rheumatoid Arthritis treated with immunomodulating medications admitted for Shortness of breath, fatigue and tiredness but no fever. COVID-19 and influenza infections (PCR) tests were both negative. At presentation, her WBC was 9900. CT with contrast of her chest showed no embolism, but multi-focal widespread groundglass opacities. Blood culture was negative, MRSA screen was negativetoo, but Fungitell test was positive (with a value of more than 500) and serum LDH test was elevated to 382. CMV quantitaive PCR was elevated to 10,000 copies. A bronchoscopy was done and CMV PCR Bronchoalveolar lavage (BAL) is detected at 650 copies/ ml. A BAL EBV PCR tests was negative. Pneumocystis Jiroveci pneumonia was detected on BAL Direct fluorescent antibody test (DFA). CMV retinitis has been ruled out by an ophthalmology exam. Patient was diagnosed with concurrent CMV and PCP pneumonia infection and her respiratory status worsened mandating a brief ICU stay. Treatment was started with Bactrim, Valganciclovir and Ganciclovir with progressive improvement. In a follow up appointment at the infectious diease clinic two months later, the patient condition improved but was still in need for supplemental oxygen through nasal canula. DISCUSSION: A concurrent CMV and PCP microorganisms lung infection is rare, but patient with underlying immunocompromise constitue a major risk factor for that. CONCLUSIONS: Patients with underlying immuncompromise conditions are at risk of many infections with grave morbidity and mortality risks. Though it is a rare to have a concurrent CMV and PCP lung infection, a patient treated with immunomodulating medications including methotrexate, prednisone and rituximab was a culprit for severe infection. Reference #1: Peghin, M., Hirsch, H. H., Len, Ó., Codina, G., Berastegui, C., Sáez, B., Solé, J., Cabral, E., Solé, A., Zurbano, F., López-Medrano, F., Román, A., & Gavaldá, J. (2016). Epidemiology and immediate indirect effects of respiratory viruses in lung transplant recipients: A 5-year prospective study. American Journal of Transplantation, 17(5), 1304–1312. https://doi.org/10.1111/ajt.14042 DISCLOSURES: No relevant relationships by MohD Ibrahim

2.
Journal of South China Agricultural University ; 41(5):27-35, 2020.
Article in Chinese | CAB Abstracts | ID: covidwho-2040361

ABSTRACT

Objective: To prepare monoclonal antibodies against porcine epidemic diarrhea virus (PEDV) N protein, and develop an indirect immuno-fluorescence assay method used for detecting PEDV. Method: The expressed recombinantly PEDV N protein was used as an immunogen and 8-week-old female BALB/c mice were immunized. Then their spleen cells with high antibody titer were isolated and fused with SP2/0 cells. The hybridoma cell lines secreting monoclonal antibodies against PEDV N protein were screened. In Vero cells infected with PEDV, monoclonal antibody of anti-PEDV N protein was used as the primary antibody and FITC-goat-anti-mouse IgG was used as the secondary antibody to develop indirect immuno-fluorescence assay method used for detecting PEDV. Result: The prepared hybridoma cell lines could stably secrete anti-PEDV N protein antibodies, ELISA antibody titer in cell supernatant was above 1:3 200, and in mouse ascites above 1:1 000 000. While monoclonal antibodies were applied in established indirect immuno-fluorescence assay, the optimal conditions were that cells were fixed with 80% () acetone at -20 degrees C for 30 min;The primary antibody was diluted 1 000 times by PBS buffer solution and incubated at 4 degrees C overnight;The secondary antibody was diluted 100 times by PBS buffer solution and incubated at 37 degrees C for 1 h. Transmissible gastroenteritis virus (TGEV), classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine reproductive virus (PRV), porcine enteric a corone virus (PEAV), porcine rotavirus (PoRV) and PEDV were detected by established indirect immuno-fluorescence assay method, only PEDV showed positive, all the else viruses showed negative.

3.
Acta Veterinaria et Zootechnica Sinica ; 53(6):2024-2028, 2022.
Article in Chinese | CAB Abstracts | ID: covidwho-2025545

ABSTRACT

This study aimed to analyze the proliferation characteristics of porcine deltacoronavirus (PDCoV) in suspension cultured porcine kidney cells LLC-PK1, so as to provide Candidate cell for large-scale production of PDCoV inactivated vaccine. LLC-PK1 cells were suspended by gradually decreasing serum method. PDCoV adaptive monoclonal cell lines were screened by limited dilution method. Indirect immunofluorescence method was used to identify the infectivity of PDCoV. The initial cell density, MOI, time of receiving virus collection and TPCK pancreatin concentration were screened to determine the best suspension culture conditions. The suspension cell strain LLC-PK1Sa which can proliferate PDCoV efficiently was screened out;PDCoV can specifically infect LLC-PK1 cells;PDCoV inoculated LLC-PK1Sa cells with a density of 2 x 106 cells.mL-1 according to the MOI of 10-3, When the final concentration of TPCK pancreatin reached 7.5 g.mL-1, the titer of virus solution harvested 48 h after inoculation was the highest. In this study, the efficient proliferation of PDCoV in LLC-PK1Sa suspension cells was realized for the first time, and the suspension culture conditions were preliminarily optimized, which could provide theoretical reference for large-scale production of PDCoV inactivated vaccine.

4.
Wiener Tierarztliche Monatsschrift ; 109(Artikel 11), 2022.
Article in English | CAB Abstracts | ID: covidwho-2025202

ABSTRACT

We have evaluated the diagnostic performance of immunochromatographic point-of-care tests (POCT) for the detection of rotavirus, coronavirus, Escherichia (E.) coli F5, Cryptosporidium (C.) parvum, Clostridium (Cl.) perfringens and Giardia (G.) intestinalis in fresh and thawed faecal samples from calves aged up to six months with diarrhoea. We performed POCTs to detect rotavirus, coronavirus, E. coli F5, C. parvum, Cl. perfringens and G. intestinalis on fresh samples in a field study and re-evaluated the performance for C. parvum, Cl. perfringens and G. intestinalis using thawed samples. We calculated the performance based on the results of the reference methods, which were RT-qPCR for the detection of rota- and coronavirus and bacteriological culturing and PCR to detect E. coli F5 and Cl. perfringens a and ss2 toxins. C. parvum was detected by phase-contrast microscopy and G. intestinalis by immunofluorescence microscopy. We collected 177 faecal samples from diarrhoeic calves. We found good performance for the POCT targeting rotavirus (sensitivity (SE)=92.9%;specificity (SP)=95.6%) and C. parvum (SE=63.3%;SP=96.2%). For E. coli F5, the number of true positive samples (n=1) was too low to evaluate the performance. The POCT to detect coronavirus gave a poor performance (SE=3.3%;SP=96.6%) and the POCT to detect Cl. perfringens a moderate performance (SE=52.8%;SP=78.2%). G. intestinalis POCT showed a higher sensitivity to immunofluorescence microscopy in thawed than in fresh faecal samples (SE=43.9% versus SE=29.2%). There are substantial differences in diagnostic performance between the commercially available immunochromatographic POCTs. Still, POCT can make a valuable contribution to the diagnosis and prevention of calf diarrhoea.

5.
Acta Microbiologica Sinica ; 2:672-685, 2022.
Article in Chinese | CAB Abstracts | ID: covidwho-1841702

ABSTRACT

[Objective] To explore whether porcine deltacoronavirus (PDCoV) can infect and proliferate in different animal species-derived cell lines. [Methods] The Sichuan isolate CHN-SC2015of PDCoV was inoculated in twelve cell lines derived from hamster,poultry,monkey, human and swine. After at least five blindly passages in each cell line, the virus was identified by RT-PCR,RT-q PCR, indirect immunofluorescence assay (IFA), and sequencing. [Results] PDCoV caused distinct cytopathic effect (CPE) in Vero,PAM,PK15,ST, and LLC-PK1 cells at the 1st passage (P1) and proliferated to various degrees in PAM,PK15,ST, and LLC-PK1 cells, while the CPE gradually disappeared during subsequent passages in Vero and PAM cells. Except that in the three susceptible cell lines (PK15,LLC-PK1, and ST), the viral copies of the infected cell lines gradually decreased with the increase in passages, and PDCoV could not be detected at P4 or P5 of DEF,Marc-145,HEK-293,ZYM-SIEC02, and PAM cells. PCR results showed that PDCoV could be detected only in CEF and Vero cells at P5. The IFA results showed that PDCoV could infect other cell lines except BHK-21 and ZYM-SIEC02, and specific immunofluorescence was observed in PK15,LLC-PK1, and ST cells at P1,P3, and P9. Therefore, only three cell lines (PK15,LLC-PK1, and ST) were suitable for serial passage, with the virus titers up to 107.11,107.00, and 107.37 TCID50/mL at P9,respectively. After passage in different cell lines,CHN-SC2015 accumulated 14 nucleotide mutations corresponding to 12 amino acid mutations. [Conclusion] This study indicates that PDCoV can infect a variety of cells in vitro, suggesting that it may have the potential of cross-species transmission.

6.
J Int Med Res ; 50(1): 3000605211069279, 2022 Jan.
Article in English | MEDLINE | ID: covidwho-1598885

ABSTRACT

BACKGROUND: During the coronavirus disease 2019 (COVID-19) pandemic, antibody screening is a critical tool to assess anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immunity. We examined variation in antibody titers associated with age and sex among patients with confirmed COVID-19. METHODS: Blood IgG levels were tested in 1081 patients with positive SARS-CoV-2 quantitative reverse transcription polymerase chain reaction (RT-qPCR) tests between 1 September and 31 December 2020. Patients who did not experience reinfection were identified. Serum IgG levels were measured by immunofluorescence assay. Antibody positivity and antibody titers were analyzed according to time since infection, sex, and age. RESULTS: The mean (standard deviation) age was 41.2 (14.2) years and 41.2% of patients were women. The lowest antibody positivity rate between the first and ninth month post-infection was detected in the sixth month. The lowest antibody titers among patients aged 20 to 80 years occurred in those aged 30 to 39 years. The IgG titer was positively correlated with age in years (r = 0.125) and decades (r = 0.126). CONCLUSIONS: Six months after infection, anti-SARS-CoV-2 antibody titers increased. Anti-SARS-CoV-2 antibody titers also increased with age. Immunity and pathogenicity should be investigated in addition to antibody positivity rates and antibody titers.


Subject(s)
COVID-19 , Adult , Antibodies, Viral , Female , Fluorescent Antibody Technique , Humans , Immunoglobulin G , Pandemics , SARS-CoV-2
7.
Oman Med J ; 36(5): e307, 2021 Sep.
Article in English | MEDLINE | ID: covidwho-1502661

ABSTRACT

OBJECTIVES: We sought to determine whether SARS-CoV-2 infections are associated with anosmia and if this virus infects other neuronal cells. We utilized male and female olfactory neuronal cell lines and other olfactory cell lines to determine the viral targets. METHODS: We used four undifferentiated and two partially differentiated human developing neuronal cell lines. Infectivity was confirmed by reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), immunofluorescence assay (IFA) probing with anti-SARS-CoV-2 antibody, evaluation of cytopathic effects, and neurite formation. We induced partial differentiation of all cell lines (since both olfactory cell lines were terminally differentiated) with retinoic acid (RA) to determine whether differentiation was a factor in viral permissiveness. The expression of serine protease, transmembrane serine protease 2 (TMPRSS2), and angiotensin-converting enzyme II (ACE2) receptors were examined by RT-qPCR and IFA to determine the mechanism of viral entry. RESULTS: Four to five days after exposure, both olfactory cell lines exhibited morphological evidence of infection; IFA analyses indicated that ~30% of the neurons were SARS-CoV-2 positive. At two weeks, 70-80% were positive for SARS-CoV-2 antigens. The partially differentiated (CRL-2266 and CRL-2267) and undifferentiated cell lines (CRL-2142, CRL-2149, CRL-127, and CDL-2271) were essentially non-permissive. After RA treatment, only CRL-127 exhibited slight permissiveness (RT-qPCR). The TMPRSS2 receptor showed high expression in olfactory neurons, but low expression in RA treated CRL-127. ACE2 exhibited high expression in olfactory neurons, whereas other cell lines showed low expression, including RA-treated cell lines. ACE2 expression slightly increased in CRL-127 post RA-treatment. CONCLUSIONS: Our studies confirm neurotropism of SARS-CoV-2 to olfactory neurons with viral entry likely mediated by TMPRSS2/ACE2. Other neuronal cell lines were non-permissive. Our results established that the nerve cells were infected regardless of male or female origin and strengthened the reported association of COVID-19 with loss of smell in infected individuals.

8.
Biomedica ; 40(Supl. 2): 148-158, 2020 10 30.
Article in English, Spanish | MEDLINE | ID: covidwho-916545

ABSTRACT

Introduction: SARS-CoV-2 has been identified as the new coronavirus causing an outbreak of acute respiratory disease in China in December, 2019. This disease, currently named COVID-19, has been declared as a pandemic by the World Health Organization (WHO). The first case of COVID-19 in Colombia was reported on March 6, 2020. Here we characterize an early SARS-CoV-2 isolate from the pandemic recovered in April, 2020. Objective: To describe the isolation and characterization of an early SARS-CoV-2 isolate from the epidemic in Colombia. Materials and methods: A nasopharyngeal specimen from a COVID-19 positive patient was inoculated on different cell lines. To confirm the presence of SARS-CoV-2 on cultures we used qRT-PCR, indirect immunofluorescence assay, transmission and scanning electron microscopy, and next-generation sequencing. Results: We determined the isolation of SARS-CoV-2 in Vero-E6 cells by the appearance of the cytopathic effect three days post-infection and confirmed it by the positive results in the qRT-PCR and the immunofluorescence with convalescent serum. Transmission and scanning electron microscopy images obtained from infected cells showed the presence of structures compatible with SARS-CoV-2. Finally, a complete genome sequence obtained by next-generation sequencing allowed classifying the isolate as B.1.5 lineage. Conclusion: The evidence presented in this article confirms the first isolation of SARSCoV-2 in Colombia. In addition, it shows that this strain behaves in cell culture in a similar way to that reported in the literature for other isolates and that its genetic composition is consistent with the predominant variant in the world. Finally, points out the importance of viral isolation for the detection of neutralizing antibodies, for the genotypic and phenotypic characterization of the strain and for testing compounds with antiviral potential.


Introducción. El nuevo coronavirus causante de un brote de enfermedad respiratoria aguda en China en diciembre de 2019 se identificó como SARS-CoV-2. La enfermedad, denominada COVID-19, fue declarada pandemia por la Organización Mundial de la Salud (OMS). El primer caso de COVID-19 en Colombia se reportó el 6 de marzo de 2020; en este estudio se caracterizó un aislamiento temprano del virus SARS-CoV-2 de una muestra ecolectada en abril de 2020. Objetivos. Describir y caracterizar una cepa temprana a partir de un aislamiento de SARSCoV-2 durante la pandemia en Colombia. Materiales y métodos. Se obtuvo una muestra de un paciente con COVID-19 confirmada por qRT-PCR; la muestra fue inoculada en diferentes líneas celulares hasta la aparición del efecto citopático. Para confirmar la presencia de SARS-CoV-2 en el cultivo, se utilizó la qRT-PCR a partir de los sobrenadantes, la inmunofluorescencia indirecta (IFI) en células Vero-E6, así como microscopía electrónica y secuenciación de nueva generación (nextgeneration sequencing). Resultados. Se confirmó el aislamiento de SARS-CoV-2 en células Vero-E6 por la aparición del efecto citopático tres días después de la infección, así como mediante la qRT-PCR y la IFI positiva con suero de paciente convaleciente positivo para SARS-CoV-2. Además, en las imágenes de microscopía electrónica de trasmisión y de barrido de células infectadas se observaron estructuras compatibles con viriones de SARS-CoV-2. Por último, se obtuvo la secuencia completa del genoma, lo que permitió clasificar el aislamiento como linaje B.1.5. Conclusiones. La evidencia presentada en este artículo permite confirmar el primer aislamiento de SARS-CoV-2 en Colombia. Además, muestra que esta cepa se comporta en cultivo celular de manera similar a lo reportado en la literatura para otros aislamientos y que su composición genética está acorde con la variante predominante en el mundo. Finalmente, se resalta la importancia que tiene el aislamiento viral para la detección de anticuerpos, para la caracterización genotípica y fenotípica de la cepa y para probar compuestos con potencial antiviral.


Subject(s)
Betacoronavirus/isolation & purification , Coronavirus Infections/virology , Pandemics , Pneumonia, Viral/virology , RNA, Viral/genetics , Animals , Betacoronavirus/genetics , Betacoronavirus/physiology , Betacoronavirus/ultrastructure , COVID-19 , Chlorocebus aethiops , Colombia/epidemiology , Convalescence , Coronavirus Infections/epidemiology , Cytopathogenic Effect, Viral , Fluorescent Antibody Technique, Indirect , Genome, Viral , Humans , Microscopy, Electron , Molecular Typing , Nasopharynx/virology , Pneumonia, Viral/epidemiology , Reverse Transcriptase Polymerase Chain Reaction , SARS-CoV-2 , Sequence Analysis, RNA , Species Specificity , Vero Cells , Virion/ultrastructure , Virus Cultivation
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