Liquid biopsy testing in metastatic or advanced breast cancer patients during the COVID-19 pandemic

Objective: During the COVID-19 pandemic, cancer patients had restricted access to standard of care tissue biopsy. Liquid biopsy assays using next generation sequencing technology provides a less invasive method for determining circulating tumour mutations (ctDNA) associated with targeted treatments or prognosis. As part of deploying technology to help cancer patients obtain molecular testing, a clinical program was initiated to offer liquid biopsy testing for Canadian patients with advanced or metastatic breast cancer. Method(s): Blood was drawn in two 10 mL StreckTM DNA BCTs and sent to the CAP/CLIA/DAP accredited Imagia Canexia Health laboratory for testing using the clinically validated Follow ItTM liquid biopsy assay. Plasma was isolated using a double spin protocol and plasma cell-free DNA (cfDNA) extracted using an optimized Promega Maxwell RSC method. Extracted cfDNA was amplified using the multiplex amplicon-based hotspot 30 or 38 gene panel and sequenced. An inhouse developed bioinformatics pipeline and reporting platform were used to identify pathogenic single nucleotide variants (SNVs), indels (insertions and deletions), and gene amplification. Included in the panel are genes associated with metastatic breast cancer: AKT1, BRAF, ERBB2, ESR1, KRAS, PIK3CA, TP53. Result(s): To identify biomarkers, 1214 metastatic or advanced breast cancer patient cfDNA samples were tested. There were 15 cases sent for repeat testing. We reported 48% of samples harboring pathogenic ctDNA mutations in TP53 (22%), PIK3CA (19%), ESR1 (18%), AKT1 (2%), ERBB2 (1.5%). Co-occurring variants were identified in samples with ESR1/PIK3CA as well as TP53/PIK3CA (both p-values <0.001). Interestingly, 29% of samples with mutated ESR1 harbored >= 2 ESR1 ctDNA mutations. In 56% of cases, previous molecular testing indicated the cancer subtype as hormone receptor (ER, PR) positive with/without HER2 negative status. In this specific subgroup, 49% harbored ctDNA mutations with 63% of those being PIK3CA and/or ESR1 mutations. Conclusion(s): A population of Canadian women with metastatic breast cancer were tested using a liquid biopsy gene panel during the COVID-19 pandemic for identification of biomarkers for targeted therapeutic options. Over 50% of the samples were identified as hormone positive, with greater than 60% harboring PIK3CA and ESR1 ctDNA mutations. Studies have shown that metastatic PIK3CA mutated ER-positive/HER2-negative tumors are predictive to respond to alpelisib therapy and have FDA and Health Canada approval. Additionally, ESR1 mutations are associated with acquired resistance to antiestrogen therapies, and interestingly we identified 29% of ESR1 mutated samples with multiple mutations possibly indicating resistance subclones. In future studies, longitudinal monitoring for presence of multiple targetable and resistance mutations could be utilized to predict or improve clinical management.

The pathogenesis of COVID-19: Hypercoagulation and D-dimer in thrombotic complications

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) caused a new coronavirus disease (COVID-19), which is highly contagious and its pathogenesis has not been fully elucidated. In COVID-19, the inflammation and blood coagulation systems are excessively activated. SARS-CoV-2 damages endothelial cells and pneumocytes, which leads to disruption of hemostasis in SARS. Thromboembolism is the main cause of mortality in patients with COVID-19. Clots, including pulmonary embolism (PE) and deep vein thrombosis (DVT), ranging from minor to fatal complications of the SARS-CoV-2 infection are known. Individuals with pre-existing diseases are more susceptible to the development of blood clots and poor outcomes. High levels of circulating cytokines and D-dimer (DD) are influential biomarkers of poor outcomes in COVID-19. The latter occurs as a result of hyperfibrinolysis and hypercoagulation. Plasmin is a key player in fibrinolysis and is involved in the cleavage of many viral envelope proteins, including SARS-CoV. Due to this function penetration of viruses into the host cell occurs. In addition, plasmin is involved in the pathophysiology of acute respiratory distress syndrome (ARDS) in SARS and promotes the secretion of cytokines, such as IL-6 and TNF, from activated macrophages. The focus of existing treatment to alleviate fibrinolysis in patients with COVID-19 is the use of systemic fibrinolytic therapy given thrombotic pathology in severe forms of COVID-19 which may lead to death. However, fibrinolytic therapy may be harmful in the advanced stages of COVID-19, when the status of disseminated intravascular coagulation (DIC) changes from suppressed fibrinolysis to its enhancement during the progression of the disease. This narrative review aims to elucidate the pathogenesis of COVID-19, which will further help in precise diagnosis and treatment. Take-home message: The COVID-19 virus disrupts haemostasis and thromboembolism by over activating the inflammation and blood coagulation systems. High levels of cytokines and D-dimer are indicators of pre-existing diseases and blood clots. Systemic fibrinolytic treatment can reduce severe fibrinolysis in COVID-19, which is caused by plasmin. In the late stages of DIC, when fibrinolysis increases, it may be dangerous. To improve therapy and results, understanding COVID-19 pathogenicity is critical. © 2023 by the authors.

Circulating Small RNA Profiling of Patients with Alveolar and Cystic Echinococcosis.

Alveolar (AE) and cystic (CE) echinococcosis are two parasitic diseases caused by the tapeworms Echinococcus multilocularis and E. granulosus sensu lato (s. l.), respectively. Currently, AE and CE are mainly diagnosed by means of imaging techniques, serology, and clinical and epidemiological data. However, no viability markers that indicate parasite state during infection are available. Extracellular small RNAs (sRNAs) are short non-coding RNAs that can be secreted by cells through association with extracellular vesicles, proteins, or lipoproteins. Circulating sRNAs can show altered expression in pathological states; hence, they are intensively studied as biomarkers for several diseases. Here, we profiled the sRNA transcriptomes of AE and CE patients to identify novel biomarkers to aid in medical decisions when current diagnostic procedures are inconclusive. For this, endogenous and parasitic sRNAs were analyzed by sRNA sequencing in serum from disease negative, positive, and treated patients and patients harboring a non-parasitic lesion. Consequently, 20 differentially expressed sRNAs associated with AE, CE, and/or non-parasitic lesion were identified. Our results represent an in-depth characterization of the effect E. multilocularis and E. granulosus s. l. exert on the extracellular sRNA landscape in human infections and provide a set of novel candidate biomarkers for both AE and CE detection.

INTERIM RESULTS OF THE PML-16, PML-19 PROTOCOLS FOR PRIMARY MEDIASTINAL LARGE B-CELL LYMPHOMA THERAPY

Introduction. Primary mediastinal lymphoma (PML) is an aggressive lymphoid tumor treatment success of which is determined by induction therapy. To date, none of the standard chemotherapy regimens (CT) have demonstrated an advantage in efficacy. Intensive therapy programs are associated with high toxicity. Aim - to evaluate the efficacy and toxicity of two pilot prospective treatment protocols PML-16 and PML-19 as well as the possibility of using the analysis of freely circulating tumor DNA (ctDNA) to assess MRD in patients with PML. Materials and methods. From January 2016 to January 2022, 34 previously untreated PML patients were included in the study;average age - 32;stage > I - in 60 %;extramediastinal lesions - in 14.7 %;bulky disease - in 73.5 % of patients. Positron emission tomography combined with computed tomography (PET-CT) was performed;ctDNA was determined to assess the completeness of remission. Results. Eighteen patients received treatment according to the PML-16 protocol (6 courses of chemotherapy;2 blocks of RmNHL-BFM-90 + 4 courses of R-EPOCH). After the end of therapy, all 18 patients achieved PET-negative remission. The next 16 patients received treatment according to the PML-19 protocol (4 courses of chemotherapy;2 blocks of R-mNHL-BFM-90 + 2 courses of R-EPOCH) in combination with lenalidomide. After the end of therapy, 9 (56 %) patients achieved PET-negative remission;7 (44 %) retained pathological activity (D4-5 points). After 3 and 6 months 15 (94 %) patients achieved normalization of metabolic activity. Considering the high frequency of false-positive results in patients with PML, a ctDNA study was performed to determine the depth of remission in 15 patients. After the end of therapy, all 15 patients had complete elimination of ctDNA. Of these, 5 (33 %) remained PET-positive at the end of treatment. During further observation, after 3-6 months, in 4 patients the level of metabolic activity decreased to physiological without the use of consolidating therapy. After the end of therapy, one patient suffered the new coronavirus infection, COVID-19. A month later, residual formation of SUVmax 14.2 remained in the mediastinum. The patient is currently under observation. With a median follow-up of 36 months (9 to 76 months) all 34 patients are in remission. Conclusion. The effectiveness of PML-16 made it possible to abandon the consolidation therapy and refuted the idea of the need for 6 courses of CT. The combination of programs based on the application of the principle of high-dose short-pulse induction of remission (R-mNHL-BFM-90) in combination with the prolonged administration of medium doses (R-EPOCH) was crucial in achieving a successful result. The inclusion of lenalidomide in the "PML-19" program made it possible to achieve complete remission in 100 % of cases after 4 courses. The possibility of using DNA analysis to assess MRD in patients with PML was shown.Copyright © 2022 Izdatel'stvo Meditsina. All rights reserved.

The shed P2X7 receptor is an index of adverse clinical outcome in COVID-19 patients.

Introduction: The pathophysiology of the Corona Virus Disease 2019 (COVID-19) is incompletely known. A robust inflammatory response caused by viral replication is a main cause of the acute lung and multiorgan injury observed in critical patients. Inflammasomes are likely players in COVID-19 pathogenesis. The P2X7 receptor (P2X7R), a plasma membrane ATP-gated ion channel, is a main activator of the NLRP3 inflammasome, of the ensuing release of inflammatory cytokines and of cell death by pyroptosis. The P2X7R has been implicated in COVID-19-dependent hyperinflammation and in the associated multiorgan damage. Shed P2X7R (sP2X7R) and shed NLRP3 (sNLRP3) have been detected in plasma and other body fluids, especially during infection and inflammation. Methods: Blood samples from 96 patients with confirmed SARS-CoV-2 infection with various degrees of disease severity were tested at the time of diagnosis at hospital admission. Standard haematological parameters and IL-6, IL-10, IL-1ß, sP2X7R and sNLRP3 levels were measured, compared to reference values, statistically validated, and correlated to clinical outcome. Results: Most COVID-19 patients included in this study had lymphopenia, eosinopenia, neutrophilia, increased inflammatory and coagulation indexes, and augmented sNLRP3, IL-6 and IL-10 levels. Blood concentration of sP2X7R was also increased, and significantly positively correlated with lymphopenia, procalcitonin (PCT), IL-10, and alanine transaminase (ALT). Patients with increased sP2X7R levels at diagnosis also showed fever and respiratory symptoms, were more often transferred to Pneumology division, required mechanical ventilation, and had a higher likelihood to die during hospitalization. Conclusion: Blood sP2X7R was elevated in the early phases of COVID-19 and predicted an adverse clinical outcome. It is suggested that sP2X7R might be a useful marker of disease progression.

INTERIM RESULTS OF THE PML-16, PML-19 PROTOCOLS FOR PRIMARY MEDIASTINAL LARGE B-CELL LYMPHOMA THERAPY.

Introduction. Primary mediastinal lymphoma (PML) is an aggressive lymphoid tumor treatment success of which is determined by induction therapy. To date, none of the standard chemotherapy regimens (CT) have demonstrated an advantage in efficacy. Intensive therapy programs are associated with high toxicity. Aim - to evaluate the efficacy and toxicity of two pilot prospective treatment protocols PML-16 and PML-19 as well as the possibility of using the analysis of freely circulating tumor DNA (ctDNA) to assess MRD in patients with PML. Materials and methods. From January 2016 to January 2022, 34 previously untreated PML patients were included in the study;average age - 32;stage > I - in 60 %;extramediastinal lesions - in 14.7 %;bulky disease - in 73.5 % of patients. Positron emission tomography combined with computed tomography (PET-CT) was performed;ctDNA was determined to assess the completeness of remission. Results. Eighteen patients received treatment according to the PML-16 protocol (6 courses of chemotherapy;2 blocks of RmNHL-BFM-90 + 4 courses of R-EPOCH). After the end of therapy, all 18 patients achieved PET-negative remission. The next 16 patients received treatment according to the PML-19 protocol (4 courses of chemotherapy;2 blocks of R-mNHL-BFM-90 + 2 courses of R-EPOCH) in combination with lenalidomide. After the end of therapy, 9 (56 %) patients achieved PET-negative remission;7 (44 %) retained pathological activity (D4-5 points). After 3 and 6 months 15 (94 %) patients achieved normalization of metabolic activity. Considering the high frequency of false-positive results in patients with PML, a ctDNA study was performed to determine the depth of remission in 15 patients. After the end of therapy, all 15 patients had complete elimination of ctDNA. Of these, 5 (33 %) remained PET-positive at the end of treatment. During further observation, after 3-6 months, in 4 patients the level of metabolic activity decreased to physiological without the use of consolidating therapy. After the end of therapy, one patient suffered the new coronavirus infection, COVID-19. A month later, residual formation of SUVmax 14.2 remained in the mediastinum. The patient is currently under observation. With a median follow-up of 36 months (9 to 76 months) all 34 patients are in remission. Conclusion. The effectiveness of PML-16 made it possible to abandon the consolidation therapy and refuted the idea of the need for 6 courses of CT. The combination of programs based on the application of the principle of high-dose short-pulse induction of remission (R-mNHL-BFM-90) in combination with the prolonged administration of medium doses (R-EPOCH) was crucial in achieving a successful result. The inclusion of lenalidomide in the "PML-19" program made it possible to achieve complete remission in 100 % of cases after 4 courses. The possibility of using DNA analysis to assess MRD in patients with PML was shown.Copyright © 2022 Izdatel'stvo Meditsina. All rights reserved.

CRISPR-Cas assisted diagnostics: A broad application biosensing approach

In addition to its remarkable genome editing capability, the CRISPR-Cas system has proven to be very effective in many fields of application, including the biosensing of pathogenic infections, mutagenic defects, or early cancer diagnosis. Thanks to their many advantages in terms of simplicity, efficiency, and reduced time, several CRISPR-Cas systems have been described for the design of sensitive and selective analytical tools, paving the way for the development and further commercialization of next-generation diagnostics. However, CRISPR-Cas-based biosensors still need further research efforts to improve some drawbacks, such as the need for target amplification, low reproducibility, and lack of knowledge of exploited element robustness. This review aims to describe the latest trends in the design of CRISPR-Cas biosensing technologies to better highlight the insights of their advantages and to point out the limitations that still need to be overcome for their future market entry as medical diagnostics.Copyright © 2023 Elsevier B.V.

Early detection of recurrent HPV+ hypopharyngeal squamous cell carcinoma by circulating tumor DNA

Circulating tumor DNA is a liquid biomarker that offers a highly specific method to assess HPV-associated tumor burden via a blood draw. It has the potential for many clinical applications in cancer care, including prognostication, monitoring treatment response, and surveillance for disease recurrence. In this case report, we present a case of recurrent HPV-associated hypopharyngeal squamous cell carcinoma first detected by circulating tumor HPV DNA that demonstrates the role of circulating tumor HPV DNA tests in posttreatment surveillance and the utility of HPV testing in all HPV-mediated tumors, regardless of subsite.Copyright © 2023 Elsevier Inc.

A Case Report Of The Effects Of Chemotherapy On Post-heart-transplant Gene Expression Profiling

Introduction: Malignancies are a major complication of heart transplant (HT). Noninvasive surveillance after HT using gene expression (GEP) profiling and donor derived cell free DNA (dd-cfDNA) are noninferior to biopsy and are widely utilized. The interpretation of % dd-cfDNA, is not well understood in malignancies with a conceptual increase in the recipient fraction. The effect of chemotherapy on GEP in the setting of post-HT surveillance has not been described to the best of our knowledge. Hypothesis: Induction of chemotherapy will cause global transcriptional reduction in GEP. Method(s): GEP was performed with AlloMap (AM, CareDx), which evaluates expression levels of 11 mononuclear cell genes, involved in lymphocyte activation, T-cell priming, cell migration, hematopoietic proliferation, steroid sensitivity, and platelet activation. Scores range from 0-40, higher scores have a stronger correlation with rejection. At our center a total of 995 draws were analyzed from 2019-2022. In parallel dd-cfDNA, which informs about graft injury was analyzed using AlloSure (AS, CareDx). Case Events: A 71-year-old male HT recipient for nonischemic cardiomyopathy and no rejection history was diagnosed with metastatic gastric adenocarcinoma at 16 months post-HT. Following diagnosis, mycophenolic acid was stopped, prednisone 5 mg was started, and tacrolimus trough goal was gradually lowered to 4-6 given infectious complications. Palliative chemotherapy with folinic acid, fluorouracil (5-FU), oxaliplatin (FOLFOX) was initiated at 18 months post-HT with planned dose reduction of oxaliplatin and holding of 5-FU bolus to reduce risk of myelosuppression given comorbidities. Oxaliplatin was stopped at 18 months post HT. Due to COVID he last received 5-FU at 33 months post-HT. Graft function remained stable and DSA negative. At 36 months post-HT, he developed a bowel obstruction without surgical options for interventions and expired shortly thereafter. Result(s): With initiation of prednisone and following chemotherapy there was a drastic decrease in AM scores (Fig. A). Steroid therapy led to an 18% decline in AM scores, the greatest decrease occurred with chemotherapy, with 67% decline from the mean when compared to all center patients (Fig B). Dd-cfDNA levels remained stable during the course aside from one early elevation. Conclusion(s): To the best of our knowledge this is the first published case on the effect of chemotherapy on GEP profiling in the setting of post-HT surveillance. This case advises caution when interpreting GEP in the setting of chemotherapy showing great reduction in GEP scores. While dd-cfDNA levels remained relatively stable after malignancy diagnosis and treatment initiation further studies will need to inform on the use of both GEP and dd-cfDNA in these patients.Copyright © 2022

Biosensor Design for the Detection of Circulating Tumor Cells Using the Quartz Crystal Resonator Technique.

A new mass-sensitive biosensing approach for detecting circulating tumor cells (CTCs) using a quartz crystal resonator (QCR) has been developed. A mathematical model was used to design a ring electrode-based QCR to eliminate the Gaussian spatial distribution of frequency response in the first harmonic mode, a characteristic of QCRs, without compromising the sensitivity of frequency response. An ink-dot method was used to validate the ring electrode fabricated based on our model. Furthermore, the ring electrode QCR was experimentally tested for its ability to capture circulating tumor cells, and the results were compared with a commercially available QCR with a keyhole electrode. An indirect method of surface immobilization technique was employed via modification of the SiO2 surface of the ring electrode using a silane, protein, and anti-EpCAM. The ring electrode successfully demonstrated eliminating the spatial nonuniformity of frequency response for three cancer cell lines, i.e., MCF-7, PANC-1, and PC-3, compared with the keyhole QCR, which showed nonuniform spatial response for the same cancer cell lines. These results are promising for developing QCR-based biosensors for the early detection of cancer cells, with the potential for point-of-care diagnosis for cancer screening.

Circulating cfDNA is an excellent marker of severity in Covid-19

Introduction: LDH is released by cytokine-mediated tissue damage, reflecting injury and possible organ dysfunction. It is associated with severity and mortality in patients with Covid-19. Circulating free DNA (cfDNA) is a marker of tissue damage, released by the nucleus and mitochondria after cell destruction. Recent studies have showed its possible role as a marker of tissue damage in Covid-19. Method(s): Prospective, observational, longitudinal study of 91 hospitalized patients with different degrees of severity. The evolution distinguished viral, early inflammatory and late inflammatory phases. Clinical data, immune cells, proinflammatory cytokine levels(TNF-alpha,IL-1beta,IL-8,IL-6,INF-gamma,IL-17A andG-CSF),serum inflammatory markers (CRP,PCT,D-dimer,ferritin) and tissue damage markers (LDH and cfDNA) were included. Result(s): LDH levels increased in parallel with severity, but did not discriminate mortality in the first sample and only showed significantly higher levels in critically ill patients. The IL-6/LDH correlation was close and significant in all degrees of severity and at all stages of the disease. The cfDNA value was higher in more severe and patients who died, and was the only biomarker that remained higher in these patients during the 3 phases of the disease. The multivariate study showed that cfDNA, and not IL-6, was an independent risk factor for critical status and ICU admission. Conclusion(s): cfDNA is an excellent marker of tissue damage, severity and mortality in Covid-19 disease, better than LDH. The IL-6/LDH association seems to reflect better timely inflammation, and cfDNA probably reflects better the extent of tissue damage.

The immediate impact of the COVID-19 pandemic on polio immunization and surveillance activities.

In addition to affecting individual health the COVID-19 pandemic has disrupted efforts to deliver essential health services around the world. In this article we present an overview of the immediate programmatic and epidemiologic impact of the pandemic on polio eradication as well as the adaptive strategic and operational measures taken by the Global Polio Eradication Initiative (GPEI) from March through September 2020. Shortly after the World Health Organization (WHO) declared a global pandemic on 11 March 2020, the GPEI initially redirected the programme's assets to tackle COVID-19 and suspended house-to-house supplementary immunization activities (SIAs) while also striving to continue essential poliovirus surveillance functions. From March to May 2020, 28 countries suspended a total of 62 polio vaccine SIAs. In spite of efforts to continue poliovirus surveillance, global acute flaccid paralysis (AFP) cases reported from January-July 2020 declined by 34% compared with the same period in 2019 along with decreases in the mean number of environment samples collected per active site in the critical areas of the African and Eastern Mediterranean regions. The GPEI recommended countries should resume planning and implementation of SIAs starting in July 2020 and released guidelines to ensure these could be done safely for front line workers and communities. By the end of September 2020, a total of 14 countries had implemented circulating vaccine-derived poliovirus type 2 (cVDPV2) outbreak response vaccination campaigns and Afghanistan and Pakistan restarted SIAs to stop ongoing wild poliovirus type 1 (WPV1) transmission. The longer-term impacts of disruptions to eradication efforts remain to be determined, especially in terms of the effect on poliovirus epidemiology. Adapting to the pandemic situation has imposed new considerations on program implementation and demonstrated not only GPEI's contribution to global health security, but also identified potential opportunities for coordinated approaches across immunization and health services.

Covid-19 and NETs

Introduction: Neutrophil extracellular traps (NETs) have recently been linked to an important role in the pathogenesis of Covid-19. Method(s): Prospective observational study of 91 hospitalized patients. We studied longitudinally the viral phase, early inflammatory and late, and the 4 most specific components of NETs: cell free-DNA (cfDNA), MPO-DNA and NE-DNA complexes and citrullinated Histone 3 (citH3). Result(s): We observed elevated levels vs controls of MPO-DNA and NE-DNA complexes and cfDNA at admission and in the 3 phases of the disease. CitH3 was elevated from the early inflammatory phase onwards. There was a significant correlation in survivors (r=0.798) and in all severity degrees between MPO and NE and between cfDNA and H3 cit (r=0.3), but not in the rest of combinations among the 4, nor in dead patients. We did not observe any correlation in any group between MPO or NE with citH3. There was an increase of only cfDNA levels in more severe patients. The area under the ROC curve for critical severity and mortality was high for cfDNA (0.7327 and 0.7482) and much poorer for the other 3 NETs markers. Conclusion(s): -We found evidence of neutrophil activation of NETs components in Covid-19, during the 3 phases of the disease, but without a clear relationship with severity and mortality. -cfDNA was related to severity and mortality, and its sources appeared to be more related to tissue damage than to NETs -The best correlation between them was MPO-NE, and these more neutrophil-specific markers reflect probably better NET formation. NETs role has maybe been overestimated using other less specific markers.

miRNAs: The Road from Bench to Bedside.

miRNAs are small noncoding RNAs that control gene expression at the posttranscriptional level. It has been recognised that miRNA dysregulation reflects the state and function of cells and tissues, contributing to their dysfunction. The identification of hundreds of extracellular miRNAs in biological fluids has underscored their potential in the field of biomarker research. In addition, the therapeutic potential of miRNAs is receiving increasing attention in numerous conditions. On the other hand, many operative problems including stability, delivery systems, and bioavailability, still need to be solved. In this dynamic field, biopharmaceutical companies are increasingly engaged, and ongoing clinical trials point to anti-miR and miR-mimic molecules as an innovative class of molecules for upcoming therapeutic applications. This article aims to provide a comprehensive overview of current knowledge on several pending issues and new opportunities offered by miRNAs in the treatment of diseases and as early diagnostic tools in next-generation medicine.

Comparison of circulating bacterial profiles between mild and severe COVID-19 patients.

Recent reports revealed that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected patients can develop bacteremia; however, the circulating bacterial profile is not well studied. Therefore, this study has aimed to investigate circulating bacterial profile in mild (n = 15) and severe (n = 13) SARS-CoV-2-infected patients as well as healthy controls (n = 10), using 16S rDNA (V4) sequencing approach. The alpha diversity indexes and Bray-Curtis dissimilarity matrix revealed that the bacterial profiles between the two conditions are significantly different. Correspondingly, the relative abundance indicates that the predominant bacterial phylum in both conditions was Proteobacteria. At genus level, the dominant bacterial genera in the mild patients belonged to Sphingomonas, Stenotrophomonas, and Achromobacter, while bacterial genera belonging to Enhydrobacter, Comamonas, and Acinetobacter were dominant in the severe patients. Furthermore, Linear discriminant analysis (LDA) Effect Size (LEfSe). revealed that Stenotrophomonas, Delftia, Achromobacter, and Neisseria were enriched in the mild condition, while Agrobacterium, Comamonas, Pseudomonas, Corynebacterium, Alkaliphilus, and Kocuria were enriched in the severe patients. These results revealed a distinct circulating bacterial profile in the mild and severe SARS-CoV-2-infected patients, which may provide an insight for further therapeutic strategy.

T follicular helper cell responses to SARS-CoV-2 vaccination among healthy and immunocompromised adults.

The worldwide rollout of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccinations in the last 2 years has produced a multitude of studies investigating T-cell responses in the peripheral blood and a limited number in secondary lymphoid tissues. As a key component to an effective immune response, vaccine-specific T follicular helper (Tfh) cells are localized in the draining lymph node (LN) and assist in the selection of highly specific B-cell clones for the production of neutralizing antibodies. While these cells have been noted in the blood as circulating Tfh (cTfh) cells, they are not often taken into consideration when examining effective CD4+ T-cell responses, particularly in immunocompromised groups. Furthermore, site-specific analyses in locations such as the LN have recently become an attractive area of investigation. This is mainly a result of improved sampling methods via ultrasound-guided fine-needle biopsy (FNB)/fine-needle aspiration (FNA), which are less invasive than LN excision and able to be performed longitudinally. While these studies have been undertaken in healthy individuals, data from immunocompromised groups are lacking. This review will focus on both Tfh and cTfh responses after SARS-CoV-2 vaccination in healthy and immunocompromised individuals. This area of investigation could identify key characteristics of a successful LN response required for the prevention of infection and viral clearance. This furthermore may highlight responses that could be fine-tuned to improve vaccine efficacy within immunocompromised groups that are at a risk of more severe disease.

Concurrent outbreaks of circulating vaccine-derived poliovirus types 1 and 2 affecting the Republic of the Philippines and Malaysia, 2019-2021.

Concurrent outbreaks of circulating vaccine-derived poliovirus serotypes 1 and 2 (cVDPV1, cVDPV2) were confirmed in the Republic of the Philippines in September 2019 and were subsequently confirmed in Malaysia by early 2020. There is continuous population subgroup movement in specific geographies between the two countries. Outbreak response efforts focused on sequential supplemental immunization activities with monovalent Sabin strain oral poliovirus vaccine type 2 (mOPV2) and bivalent oral poliovirus vaccines (bOPV, containing Sabin strain types 1 and 3) as well as activities to enhance poliovirus surveillance sensitivity to detect virus circulation. A total of six cVDPV1 cases, 13 cVDPV2 cases, and one immunodeficiency-associated vaccine-derived poliovirus type 2 case were detected, and there were 35 cVDPV1 and 31 cVDPV2 isolates from environmental surveillance sewage collection sites. No further cVDPV1 or cVDPV2 have been detected in either country since March 2020. Response efforts in both countries encountered challenges, particularly those caused by the global COVID-19 pandemic. Important lessons were identified and could be useful for other countries that experience outbreaks of concurrent cVDPV serotypes.

Review of use of inactivated poliovirus vaccine in campaigns to control type 2 circulating vaccine derived poliovirus (cVDPV) outbreaks.

Delivering inactivated poliovirus vaccine (IPV) with oral poliovirus vaccine (OPV) in campaigns has been explored to accelerate the control of type 2 circulating vaccine-derived poliovirus (cVDPV) outbreaks. A review of scientific literature suggests that among populations with high prevalence of OPV failure, a booster with IPV after at least two doses of OPV may close remaining humoral and mucosal immunity gaps more effectively than an additional dose of trivalent OPV. However, IPV alone demonstrates minimal advantage on humoral immunity compared with monovalent and bivalent OPV, and cannot provide the intestinal immunity that prevents infection and spread to those individuals not previously exposed to live poliovirus of the same serotype (i.e. type 2 for children born after the switch from trivalent to bivalent OPV in April 2016). A review of operational data from polio campaigns shows that addition of IPV increases the cost and logistic complexity of campaigns. As a result, campaigns in response to an outbreak often target small areas. Large campaigns require a delay to ensure logistics are in place for IPV delivery, and may need implementation in phases that last several weeks. Challenges to delivery of injectable vaccines through house-to-house visits also increases the risk of missing the children who are more likely to benefit from IPV: those with difficult access to routine immunization and other health services. Based upon this information, the Strategic Advisory Group of Experts in immunization (SAGE) recommended in October 2020 the following strategies: provision of a second dose of IPV in routine immunization to reduce the risk and number of paralytic cases in countries at risk of importation or new emergences; and use of type 2 OPV in high-quality campaigns to interrupt transmission and avoid seeding new type 2 cVDPV outbreaks.

Nanotechnology for ultrafast nucleic acid amplification

Nucleic acid detection has been one of the most valued tools in point-of-care diagnostics from life science, agriculture, food safety and environmental surveillance, because of its high sensitivity, great specificity and simple operation. Since polymerase chain reactions (PCR) were discovered, more and more researchers attach importance to exploring ultrafast nucleic acid amplification methods for further expediting the process of detection and curbing infectious diseases' high spread rate, especially after the coronavirus disease 2019 (COVID-19) worldwide pandemic event. Nowadays, nanotechnology as one of the most cut-ting-edge technologies has aroused growing attention. In this review, we describe new advances in na-notechnology research for ultrafast nucleic acid amplification. We have introduced commonly used nanotechnologies, namely nanofluidics, nanoporous materials, nanoparticles and so on. Recent advances in these nanotechnologies for ultrafast sample pretreatments, accelerated enzymatic amplification and rapid heating/cooling processes was summarized. Finally, challenges and perspectives for the future applications of ultrafast nucleic acid amplification are presented.(c) 2022 Elsevier Ltd. All rights reserved.