Rapid detection of SARS-CoV-2 infection by multicapillary column coupled ion mobility spectrometry (MCC-IMS) of breath. A proof of concept study

AbstractThere is an urgent need for screening patients of having a communicable viral disease to cut infection chains.We could recently demonstrate that MCC-IMS of breath is able to identify Influenza-A infected patients. With decreasing Influenza epidemic and upcoming SARS-CoV-2 infections we extended our study to the analysis of patients with suspected SARS-CoV-2 infections.51 patients, 23m, 28f, aged 64 ± 16 years, were included in this study.Besides RT-PCR analysis of nasopharyngeal swabs all patients underwent MCC-IMS analysis of breath. 16 patients, 7m, 9f, were positive for SARS-CoV-2 by RT-PCR. There was no difference in gender or age according to the groups.Stepwise canonical discriminant analysis could correctly classify the infected and non-infected subjects in 98% by cross-validation. Afterwards we combined the Influenza-A sub study and the SARS-CoV-2-sub study for a total of 75 patients, 34m, 41f, aged 64.8 ± 1.8 years, 14 positive for Influenza-A, 16 positive for SARS-CoV-2, the remaining 44 patients were used as controls. In one patient RT-PCR was highly suspicious of SARS-CoV-2 but inconclusive.There was no imbalance between the groups for age or gender.97.3% of the patients could be correctly classified to the respective group by discriminant analysis. Even the inconclusive patient could be mapped to the SARS-CoV-2 group applying the discrimination function.ConclusionMCC-IMS is able to detect SARS-CoV-2 infection and Influenza-A infection in breath. As this method provides exact, fast non-invasive diagnosis it should be further developed for screening of communicable viral diseases.Study registration: NCT04282135

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Rapid detection of SARS-CoV-2 infection by multicapillary column coupled ion mobility spectrometry (MCC-IMS) of breath. A proof of concept study

10.21203/rs.3.rs-70139/v1 ◽

2020 ◽

Author(s):

Claus Steppert ◽

Isabel Steppert ◽

William Sterlacci ◽

Thomas Bollinger

Keyword(s):

Ion Mobility ◽

Influenza A ◽

Viral Disease ◽

Pcr Analysis ◽

Proof Of Concept ◽

Influenza Epidemic ◽

Rt Pcr ◽

Respective Group ◽

Non Invasive ◽

Multicapillary Column

Abstract There is an urgent need for screening of patients having a communicable viral disease to cut infection chains. We could recently demonstrate that MCC-IMS of breath is able to identify Influenza-A infected patients. With decreasing Influenza epidemic and upcoming SARS-CoV-2 infections we went on and also analysed patients with suspected SARS-CoV-2 infections.75 patients, 34m, 41f, aged 64.4 ± 15.4 years, 14 positive for Influenza-A, 16 positive for SARS-CoV-2, the remaining 44 patients were used as controls. In one patient RT-PCR was highly suspicious of SARS-CoV-2 but initially inconclusiveBesides RT-PCR analysis of nasopharyngeal swabs all patients underwent MCC-IMS analysis of breath. There was no difference in gender or age according to the groups.97.3% of the patients could be correctly classified to the respective group by discriminant analysis. Even the inconclusive patient could be mapped to the SARS-CoV-2 group applying the discrimination function.ConclusionMCC-IMS is able to detect SARS-CoV-2 infection and Influenza-A infection in breath. As this method provides exact, fast non-invasive diagnosis it should be further developed for screening of communicable viral diseases.Trial registrationClinicalTrial.gov, NCT04282135 Registered 20 February 2020 - Retrospectively registered, https://clinicaltrials.gov/ct2/show/NCT04282135?term=IMS&draw=2&rank=1

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Artemia sp. as a DNA vaccine vector for common carp Cyprinus carpio larvae

Jurnal Akuakultur Indonesia ◽

10.19027/jai.12.54-61 ◽

2014 ◽

Vol 12 (1) ◽

pp. 54

Author(s):

Sri Nuryati ◽

Sekar Sulistyaning Hadiwibowo ◽

. Alimuddin

Keyword(s):

Dna Vaccine ◽

Common Carp ◽

Cyprinus Carpio ◽

Herpes Virus ◽

Oral Vaccine ◽

Viral Disease ◽

Pcr Analysis ◽

Rt Pcr ◽

Glycoprotein Gene ◽

Herpès Virus

ABSTRACT Koi herpes virus (KHV) is one of the most common impetuses for disease on common carp Cyprinus carpio. Generally, viral disease is difficult to cure because virus is intra-cellular parasite, that virus survives, multiplies, and lives only if it on the host cell. Oral vaccine delivery through Artemia sp. is of one alternative way to overcome this problem. This experiment was carried out by analysis DNA vaccine expression encoding of glycoprotein gene (GP-11) on C. carpio. Bacteria containing plasmid Krt-GP-11 as vaccine is served through Artemia sp. as a vector. Artemia sp. was given for one and two times a week to three weeks old common carp. Organs of fish fed by Artemia sp. were analyzed every three days after vaccination. The expression of GP-11 in kidney in each treatment is also observed by the use of RT-PCR method, within ten days after vaccination. The experiment showed that dose of DNA vaccine in whole bacteria could be expressed is 106 cfu/mL in a once or twice provisions a week. DNA vaccine could be detected in three organs. RT-PCR analysis also showed that the expression of GP-11 can be detected in all tested organs. In conclusion, Artemia sp. can be used as a vector to carry plasmid GP-11 vaccine for common carp Cyprinus carpio larvae. Keywords: DNA vaccine, KHV, Artemia sp., common carp ABSTRAK Salah satu penyakit pada ikan mas (Cyprinus carpio) yang disebabkan oleh virus adalah koi herpes virus (KHV). Penyakit yang disebabkan oleh virus umumnya sulit untuk disembuhkan karena virus merupakan parasit intraseluler, yaitu virus hanya dapat hidup, bertahan hidup, dan memperbanyak diri di dalam sel inang. Metode pemberian vaksin DNA secara oral melalui Artemia sp. merupakan salah satu alternatif pengobatan yang diharapkan dapat menangani permasalahan penyakit pada ikan yang disebabkan oleh virus. Pada penelitian ini dilakukan uji ekspresi vaksin DNA yang menyandikan glikoprotein 11 (GP-11) pada ikan mas. Bakteri yang mengandung plasmid Krt-GP-11 sebagai vaksin diberikan melalui Artemia sp. sebagai pembawa vaksin. Pemberian Artemia sp. dilakukan satu dan dua kali seminggu pada ikan mas umur tiga minggu. Keberadaan DNA vaksin di usus, ginjal, dan insang dianalisis menggunakan metode PCR. Organ diambil setiap tiga hari setelah pemberian vaksin. Ekspresi gen GP-11 juga diamati pada organ ginjal di setiap perlakuan dengan menggunakan metode RT-PCR, pada sepuluh hari setelah pemberian vaksin. Hasil penelitian menunjukkan bahwa DNA vaksin yang diberikan dengan dosis 106 cfu/mL pada perlakuan satu dan dua kali seminggu dapat terdeteksi pada ketiga organ. Hasil RT-PCR menunjukkan bahwa ekspresi GP-11 dapat terdeteksi pada semua organ uji di setiap perlakuan. Dengan demikian Artemia sp. dapat digunakan sebagai vektor pembawa vaksin plasmid GP-11 dengan frekuensi pemberian vaksin untuk larva ikan mas. Kata kunci: vaksin DNA, KHV, Artemia sp., ikan mas

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Rapid non-invasive detection of Influenza-A-infection by multicapillary column coupled ion mobility spectrometry

Journal of Breath Research ◽

10.1088/1752-7163/abb762 ◽

2020 ◽

Vol 15 (1) ◽

pp. 011001

Author(s):

Claus Steppert ◽

Isabel Steppert ◽

Thomas Bollinger ◽

William Sterlacci

Keyword(s):

Ion Mobility Spectrometry ◽

Ion Mobility ◽

Influenza A ◽

Non Invasive ◽

Multicapillary Column

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Molecular detection of Mycobacterium tuberculosis in pulmonary and extrapulmonary samples in a hospital-based study

African Health Sciences ◽

10.4314/ahs.v20i4.14 ◽

2020 ◽

Vol 20 (4) ◽

pp. 1617-23

Author(s):

Kalal Iravathy Goud ◽

Matam Kavitha ◽

Adi Mahalakshmi ◽

Ravi Vempati ◽

Abdulaziz A Alodhayani ◽

...

Keyword(s):

Mycobacterium Tuberculosis ◽

Pcr Analysis ◽

Sequencing Analysis ◽

Rt Pcr ◽

Tissue Samples ◽

Non Invasive ◽

Invasive Test ◽

Keywords Tuberculosis ◽

Bronchoalveolar Fluid

Objective: Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), remains a deadly infectious disease. India contributes to one-third of the global TB burden. However, no studies have been carried out in the Telangana (Hyderabad) population using real-time polymerase chain reaction (RT-PCR). Therefore, the current study evaluated the role of RT-PCR as a rapid and non-invasive test to diagnose TB by testing for pulmonary tuberculosis (PTB) and extrapulmonary tubercu- losis (EPTB). Materials and methods: This hospital-based study examined 1670 samples (900 EPTB; 770 PTB) comprising tissue (n = 537), peritoneal fluid (n = 420), sputum (n = 166), bronchial fluid (n = 126), cerebrospinal fluid (n = 145), ascetic fluid (n = 76), sputum pus (n = 78), urine (n = 79), and bronchoalveolar fluid (n = 43) samples. DNA from samples was separated using specific isolation kits and subjected to RT-PCR. Results: In this study, we enrolled 1670 subjects and categorized 54.4% as females and 45.6% as males. The collected sam- ples were categorized as 48.5% of fluid samples, followed by tissue (32.2%), sputum (9.9%), urine (4.7%), and pus-swab (4.6%). RT-PCR analysis revealed that 4.7% patients were positive for Mtb. Our results revealed that 61% of the affected patients were male and 39% were female. Among the specimen types, tissue samples gave the highest proportion of positive results (36.3%). Conclusion: The results showed that RT-PCR should be implemented and given top priority in TB diagnosis to save time and facilitate a definitive diagnosis. Tissue samples are highly recommended to screen the Mtb through the technique RT- PCR. Future studies should extend the technique to the global population and exome sequencing analysis should be per- formed to identify TB risk markers. Keywords: Tuberculosis (TB); EPTB; PTB; Mycobacterium tuberculosis (Mtb).

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Does sampling saliva increase detection of SARS-CoV-2 by RT-PCR? Comparing saliva with oro-nasopharyngeal swabs

10.1101/2020.07.26.20158618 ◽

2020 ◽

Cited By ~ 1

Author(s):

Ozlem Akgun Dogan ◽

Betsi Kose ◽

Nihat Bugra Agaoglu ◽

Jale Yildiz ◽

Gizem Alkurt ◽

...

Keyword(s):

Gold Standard ◽

Viral Rna ◽

Pcr Analysis ◽

Rt Pcr ◽

Standard Samples ◽

Rna Detection ◽

Detection Rates ◽

Gold Standard Method ◽

Non Invasive ◽

Isolation Period

The gold standard method in the diagnosis of SARS-CoV-2 infection is the detection of viral RNA in nasopharyngeal sample by RT-PCR. Recently, saliva samples has been suggested as an alternative due to being fast, reliable and non-invasive, rather than nasopharyngeal samples. We compared RT-PCR results in nasopharyngeal, oro-nasopharyngeal and saliva samples of COVID-19 patients. 98 of 200 patients were positive in RT-PCR analysis performed before the hospitalization. In day 0, at least one sample was positive in 67% of 98 patients. Positivity rate was 83% for both oro-nasopharyngeal and nasopharyngeal samples, while it was 63% for saliva samples (p<0.001). On day 5, RT-PCR was performed in 59 patients, 34% had at least one positive result. The positivity rate was 55% for saliva and nasopharyngeal samples, while it was 60% for oro-nasopharyngeal samples. Our study shows that the sampling saliva does not increase the sensitivity of RT-PCR tests at early stages of infection. However, on 5th day, viral RNA detection rates in saliva were similar to nasopharyngeal and oro-nasopharyngeal samples. In conclusion, we suggest that, in patients receiving treatment, virus presence in saliva, in addition to the standard samples, is important to determine the isolation period and to control the transmission.

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Discovery of molecular biomarker signatures for the detection of bladder cancer.

Journal of Clinical Oncology ◽

10.1200/jco.2013.31.6_suppl.302 ◽

2013 ◽

Vol 31 (6_suppl) ◽

pp. 302-302

Author(s):

Evan Gomes ◽

Virginia Urquidi ◽

Yunpeng Cai ◽

Yijun Sun ◽

Charles Joel Rosser ◽

...

Keyword(s):

Bladder Cancer ◽

Disease Status ◽

Healthcare Systems ◽

Urothelial Cell ◽

Pcr Analysis ◽

Monitoring And Control ◽

Rt Pcr ◽

Non Invasive ◽

Diagnostic Signature ◽

Independent Cohort

302 Background: Bladder cancer (BCa) is among the five most common malignancies world-wide, and due to high rates of recurrence, one of the most prevalent. Improvements in non-invasive urine-based assays to detect BCa would benefit both patients and healthcare systems. In this study, the goal was to identify urothelial cell transcriptomic signatures associated with BCa. Methods: Gene expression profiling (Affymetrix U133 Plus 2.0 arrays) was applied to exfoliated urothelia obtained from a cohort of 92 subjects with known bladder disease status. Computational analyses identified candidate biomarkers of BCa and an optimal predictive model was derived. Selected targets from the profiling analyses were monitored in an independent cohort of 81 subjects using quantitative real-time PCR (RT-PCR). Results: Data analysis identified 52 genes associated with BCa (p≤0.001), and gene models that optimally predicted class label were derived. RT-PCR analysis of 48 selected targets in an independent cohort identified a 14-gene diagnostic signature that predicted the presence of BCa with a specificity of 100% at 90% sensitivity. Conclusions: Exfoliated urothelia sampling provides a robust analyte for the evaluation of patients with suspected BCa. The refinement and validation of the multi-gene urothelial cell signatures identified in this preliminary study may lead to accurate, non-invasive assays for the detection of BCa. The development of an accurate, non-invasive BCa detection assay would benefit both the patient and healthcare systems through better detection, monitoring and control of disease.

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Real-Time PCR Analysis of Peripheral Blood: A Non-Invasive Technology that can be Used to Assess Tumor Burden in Breast Cancer Patients

10.21236/ada412843 ◽

2002 ◽

Author(s):

Michael Mitas

Keyword(s):

Breast Cancer ◽

Real Time ◽

Cancer Patients ◽

Real Time Pcr ◽

Peripheral Blood ◽

Tumor Burden ◽

Pcr Analysis ◽

Breast Cancer Patients ◽

Non Invasive

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Real-Time PCR Analysis of Peripheral Blood: A Non-Invasive Technology That Can Be Used to Assess Tumor Burden in Breast Cancer Patients

10.21236/ada424096 ◽

2003 ◽

Author(s):

Michael Mitas

Keyword(s):

Breast Cancer ◽

Real Time ◽

Cancer Patients ◽

Real Time Pcr ◽

Peripheral Blood ◽

Tumor Burden ◽

Pcr Analysis ◽

Breast Cancer Patients ◽

Non Invasive

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Phenotypical and Functional Characteristics of Human Regulatory T Cells during Ex Vivo Maturation from CD4+ T Lymphocytes

Applied Sciences ◽

10.3390/app11135776 ◽

2021 ◽

Vol 11 (13) ◽

pp. 5776

Author(s):

Varvara G. Blinova ◽

Natalia S. Novachly ◽

Sofya N. Gippius ◽

Abdullah Hilal ◽

Yulia A. Gladilina ◽

...

Keyword(s):

T Cells ◽

Regulatory T Cells ◽

Ex Vivo ◽

Pcr Analysis ◽

Mrna Levels ◽

Rt Pcr ◽

Inflammatory Reactions ◽

Effector Cells ◽

Cycle Regulation ◽

Further Development

Regulatory T cells (Tregs) participate in the negative regulation of inflammatory reactions by suppressing effector cells. In a number of autoimmune disorders, the suppressive function and/or the number of Tregs is compromised. The lack of active functioning Tregs can be restored with adoptive transfer of expanded ex vivo autologous Tregs. In our study, we traced the differentiation and maturation of Tregs CD4+CD25+FoxP3+CD127low over 7 days of cultivation from initial CD4+ T cells under ex vivo conditions. The resulting ex vivo expanded cell population (eTregs) demonstrated the immune profile of Tregs with an increased capacity to suppress the proliferation of target effector cells. The expression of the FoxP3 gene was upregulated within the time of expansion and was associated with gradual demethylation in the promotor region of the T cell-specific demethylation region. Real-time RT-PCR analysis revealed changes in the expression profile of genes involved in cell cycle regulation. In addition to FOXP3, the cells displayed elevated mRNA levels of Ikaros zinc finger transcription factors and the main telomerase catalytic subunit hTERT. Alternative splicing of FoxP3, hTERT and IKZF family members was demonstrated to be involved in eTreg maturation. Our data indicate that expanded ex vivo eTregs develop a Treg-specific phenotype and functional suppressive activity. We suggest that eTregs are not just expanded but transformed cells with enhanced capacities of immune suppression. Our findings may influence further development of cell immunosuppressive therapy based on regulatory T cells.

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Multiplexed detection of SARS-CoV-2 and other respiratory infections in high throughput by SARSeq

Nature Communications ◽

10.1038/s41467-021-22664-5 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Ramesh Yelagandula ◽

◽

Aleksandr Bykov ◽

Alexander Vogt ◽

Robert Heinen ◽

...

Keyword(s):

Influenza A ◽

Respiratory Infections ◽

High Sensitivity ◽

Cost Effective ◽

Massively Parallel ◽

Rt Pcr ◽

Viral Spread ◽

Saliva Analysis ◽

Double Blinded ◽

Generation Sequencing

AbstractThe COVID-19 pandemic has demonstrated the need for massively-parallel, cost-effective tests monitoring viral spread. Here we present SARSeq, saliva analysis by RNA sequencing, a method to detect SARS-CoV-2 and other respiratory viruses on tens of thousands of samples in parallel. SARSeq relies on next generation sequencing of multiple amplicons generated in a multiplexed RT-PCR reaction. Two-dimensional, unique dual indexing, using four indices per sample, enables unambiguous and scalable assignment of reads to individual samples. We calibrate SARSeq on SARS-CoV-2 synthetic RNA, virions, and hundreds of human samples of various types. Robustness and sensitivity were virtually identical to quantitative RT-PCR. Double-blinded benchmarking to gold standard quantitative-RT-PCR performed by human diagnostics laboratories confirms this high sensitivity. SARSeq can be used to detect Influenza A and B viruses and human rhinovirus in parallel, and can be expanded for detection of other pathogens. Thus, SARSeq is ideally suited for differential diagnostic of infections during a pandemic.

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