Molecular Crowding Modulates SARS-CoV-2 Aptamer Affinity

SARS-CoV-2 aptamer is a favorable candidate for the recognition and detection of SARS-CoV-2, owing to its small size and easy synthesis. However, the issue of compromised binding affinities in real samples and targeting mutant SARS-CoV-2 hinder wide applications of the aptamer. In this study, it is discovered that molecular crowding could increase binding affinity of CoV2-6C3 aptamer against RBD (Receptor Binding Domain) of SARS-CoV-2 via increasing the absolute value of the enthalpy change. The values of the equilibrium dissociation constant in molecular crowding decrease by 70% and 150%, respectively, against wild-type and mutant RBD compared with those in buffer without crowding. Moreover, the detection limit of SARS-CoV-2 pseudovirus is up to 5 times lower under molecular crowding compared to dilute conditions. The discovery deepens the understanding of aptamer-target interaction mechanisms in crowding conditions and provides an effective way to apply SARS-CoV-2 aptamer for virus recognition and detection.

Development of Duplex LAMP Technique for Detection of Porcine Epidemic Diarrhea Virus (PEDV) and Porcine Circovirus Type 2 (PCV 2).

Porcine epidemic diarrhea virus (PEDV) and porcine circovirus type 2 (PCV2) are both important global pathogenic viruses which have a significant impact on the swine industry. In this study, a duplex loop-mediated isothermal amplification (duplex LAMP) method was developed in combination with lateral flow dipstick (LFD) for simultaneous detection of PEDV and PCV2 using specific sets of primers and probes designed based on the conserved regions of a spike gene (KF272920) and an ORF gene (EF493839), respectively. The limit of detection (LOD) values of the duplex LAMP-LFD for the detection of PEDV and PCV2 were 0.1 ng/µL and 0.246 ng/µL, respectively. The LOD of duplex LAMP-LFD was 10-times more sensitive than conventional PCR and RT-PCR-agarose gel-electrophoresis (PCR-AGE and RT-PCR-AGE). No cross-reaction to each other and to other pathogenic viruses that can infect pigs were observed according to analytical specificity tests. The duplex LAMP-LFD method for the simultaneous detection of PEDV and PCV2 co-infection could be completed within approximately 1.5 h, and only a simple heating block was required for isothermal amplification. The preliminary validation using 50 swine clinical samples with positive and negative PEDV and/or PCV2 revealed that the sensitivity, specificity, and accuracy of duplex LAMP-LFD were all 100% in comparison to conventional PCR and RT-PCR. Hence, this study suggests that duplex LAMP-LFD is a promising tool for the early detection and initial screening of PEDV and PCV2, which could be beneficial for prevention, planning, and epidemiological surveys of these diseases.

Successful Treatment of Subclavian Artery Thrombosis by Catheter-Based Thrombus Aspiration and Thrombolytic Therapy: A Case Report

Clinical Information Patient Initials or Identifier Number: BP4****/22 Relevant Clinical History and Physical Exam: A 55 Y / Female C/C : Pain, numbness, cold sensation & weakness of left upper limb for 2 hours. Risk Factor : Hypertension, diabetes mellitus O/E : Pale, cold and absent of radial, ulnar, brachial pulse of left upper limb. Muscle power 3/5 left side. So2 86%, BP undetectable. Right upper limb were normal. BP 160/90 mm of hg, pules : 112 b/min, RR : 26/min. Body Temperature 37.5 C [Formula presented] [Formula presented] Relevant Test Results Prior to Catheterization: CBC : WBC 7450, HB % 10.8 g/dl, ESR 20mm in 1st hour, Platelets : 262000, SARS Cov2 Antigen : Negative PT 14.3 sec, INR : 1.07 APTT : 32.4 sec. blood group: O positive Serum Creatinine : 1.1 mg/dl Plasma glucose 9.7 mmmol/l HIV Ab : Negative HBs Ag : Negative Anti-HCV : Negative Urine R/E : Normal lipid profile : Cholesterol 280mg/dl Vascular duplex ultrasound of left upper limb : A dilated echogenic thrombus had blocked the left subclaviav artery lumen. Relevant Catheterization Findings: Conventional angiography with the lowest amount of contrast agent through the right femoral artery, revealed that left subclavian artery thrombosis with total occlusion distal to Left internal mammary artery. [Formula presented] [Formula presented] [Formula presented] Interventional Management Procedural Step: A5Fr MPA catheter with side holes was negotiated through a right femoral sheath and was placed in the left subclavian artery. Initially thrombus aspiration was done with Eliminate aspiration catheter (TERUMO) with no success. Then suction was done with the MPA catheter itself with partial removal of thrombus. Then a 5Fr Pigtail catheter was placed inside the thrombus and kept in situ. For residual thrombus 250,000u of Inj. Streptokinase as a thrombolytic drug was given through the Pigtail catheter as bolus over 30 min. The maintenance dose 100,000 u per hour was given over 24 hours through the Pigtail catheter via infusion pump. After 24 hours of thrombolytic therapy, her pain was reduced, the left hand became slightly warm, and distal pulses were feebly palpable. Moreover, the skin colour returned to near normal with improvement of pallor. Bleeding was well controlled at the catheter site. Doppler sounds revealed partial improvement of arterial flow. After evaluation of partial improvement, a low dose 1000 iu per hour of heparin (UFH)was infused intravenously for 24 hours. After 48 hours, repeat angiography via the inserted catheter at the site did not reveal any atherosclerotic plaque and confirm the thrombosis-dissolution. The latter practice demonstrated a good blood flowto the left upper distal limb leaving a little thrombus in the superficial palmer arch. [Formula presented] [Formula presented] [Formula presented] Conclusion(s): Catheter-based thrombus aspiration and thrombolytic therapy is primarily reserved for patients with acute viable limb ischemia. As observed in the presented case, thrombus aspiration and thrombolytic therapy is recommended to be considered as an alternative therapeutic method for patients with arterial thrombosis due to the rapid response, shorter treatment time and lower cost, compared to common and sometimes unsuccessful therapies.Copyright © 2023

Herpes Zoster Duplex Unilateralis as a manifestation of severe lymphopenia in COVID-19

SARS-CoV2 (severe acute respiratory syndrome coronavirus-2), a novel coronavirus was first reported in December of 2019 from Wuhan, China as an aetiological agent causing a new infectious respiratory disease (coronavirus disease 2019- COVID-19). The main clinical manifestations of COVID-19 are fever, cough, fatigue, dyspnoea and muscle aches. Herpes zoster is characterized by several groups of painful vesicles on an erythematous base with a distribution in a unilateral dermatome involving the skin and/or mucosa. Herpes zoster is caused by varicella zoster virus which remains dormant in the sensory root ganglion contained by the immune system particularly CD lymphocytes. (PsycInfo Database Record (c) 2023 APA, all rights reserved)

A Persistent Nightmare: A Case of Recurrent Brachial Artery Thrombosis

Background Brachial artery thrombosis can be seen with thromboembolism, hypercoagulability, and arterial thoracic outlet syndrome. Case A 33-year-old healthy female construction worker presented with right hand discoloration and pain. She suffered a COVID-19 infection 8 weeks prior with hand symptoms developing shortly thereafter. She could no longer work due to the pain. Duplex ultrasound and CTA of the right upper extremity (Figure) demonstrated localized thrombosis of the right brachial artery. The workup yielded no aortic or intracardiac thrombus, and cardiac event monitor showed no atrial arrhythmia. She underwent thrombectomy with brachial artery stenting and was found, during surgery, to have distal ulnar artery occlusion. Two days post-op, she had recurrent pain and was found to have brachial artery recurrent thrombosis. She underwent urgent brachial-brachial bypass. Arm pain continued despite graft patency, so ulnarpalmar bypass was performed. Decision-making Hypercoagulability workup, including antiphospholipid antibody, protein C, protein S, homocysteine, and Lp(a), was negative. Neither central thrombus on TEE nor evidence of thoracic outlet syndrome was found. As a diagnosis of exclusion, brachial artery thrombosis was ascribed to COVID infection. Despite rivaroxaban, the patient developed gangrene (Panel C) requiring partial digit amputation. Conclusion We present a case of COVID-19-induced recurrent brachial artery thrombosis despite surgical intervention. [Formula presented]Copyright © 2023 American College of Cardiology Foundation

Efficacy and safety of sonographer discretion to terminate a venous duplex ultrasound for diagnosis of deep vein thrombosis in coronavirus disease 2019 patients.

OBJECTIVE: Sonographers performing venous duplex ultrasound (VDUS) of patients with coronavirus disease 2019 (COVID-19) have an increased risk of exposure owing to their close contact with these patients for an extended period. The objective of the present study was to evaluate the efficacy of a modified COVID-19 VDUS protocol to reduce sonographer exposure to COVID-19 patients. METHODS: We performed a single-center retrospective review. Patients who had undergone VDUS under the modified COVID-19 protocol between March 1, 2020, and June 30, 2020, with a confirmed or presumed COVID-19 diagnosis at the VDUS were included. The modified COVID-19 protocol was defined as the ability of the sonographer to terminate the examination on detection of an acute deep vein thrombosis (DVT). The primary outcome measures were the number of anatomic deep venous segments recorded by the sonographer, which was used as a surrogate measure for sonographer exposure time, and the number of acute DVTs found on follow-up examinations in segments not visualized at the index VDUS. RESULTS: A total of 160 lower extremity VDUS (LEVDUS) scans and 72 upper extremity VDUS (UEVDUS) scans were performed using the modified COVID-19 protocol. The index VDUS had found an acute DVT for 44 of 160 patients (27.5%) who had undergone LEVDUS and 26 of 72 (36.6%) who had undergone UEVDUS. On follow-up imaging, 7 of 38 LEVDUS scans (17.9%) and 1 of 10 UEVDUS scans (10%) had demonstrated a new acute DVT. Malignancy and surgery 30 days before imaging were significantly associated with acute lower extremity DVT, and mechanical ventilation and extracorporeal membrane oxygenation were associated with acute upper extremity DVT. On the index VDUS, the average was 10.6 of 12 total visualized segments on LEVDUS and 6.4 of 10 total segments on UEVDUS. Of the index VDUS scans, 35.6% of the LEVDUS and 78.6% of the UEVDUS scans had been abbreviated. The index VDUS scans that were positive for acute DVT had had significantly fewer visualized segments for both lower (8.4 vs 11.5; P < .0001) and upper (4.2 vs 7.6) extremities (P < .0001). On the follow-up examinations, only one of eight new acute DVTs had been found in a patient whose index VDUS had been abbreviated and the corresponding segment not assessed. These findings did not affect the patient's clinical course. CONCLUSIONS: The modified COVID-19 VDUS protocol reduced sonographers' potential exposure time to COVID-19. Additionally, the clinical efficacy was maintained, with no missed DVTs, despite the abbreviation of the VDUS examinations.

Development of a duplex q-PCR for the simultaneous detection of Parachlamydia acanthamoebae and Simkania negevensis in environmental and clinical samples.

Parachlamydia acanthamoebae and Simkania negevensis, two Chlamydia-like bacteria, have been recently recognized as emerging human respiratory pathogens. The prevalence and frequency of these bacteria in the environment and among atypical pneumonia patients are still underestimated by classical cultures, immunohistochemistry and serology which are non-specific, long and tedious methods. This study aims to develop a new duplex probe-based q-PCR assay for the simultaneous detection and quantification of P. acanthamoebae and S. negevensis. The selected hydrolysis probes displayed no cross-reaction with the closely related Chlamydia or the other tested waterborne pathogens. The assay achieved a large dynamic range for quantification (from 5 × 106 to 5 DNA copies/reaction). Efficiencies of FAM and JOE label probes weren't affected when they were combined. They were close to 100%, indicating the linear amplification. The application of this diagnostic tool resulted in 9/47 (19%) and 4/47 (8.5%) positive water samples for P. acanthamoebae and S. negevensis, respectively. P. acanthamoebae was also covered from 2/78 (2.5%) respiratory specimens and only one case (1/200 = 0.5%) of P. acanthamoebae and SARS-CoV-2 co-infection was noticed. While S. negevensis wasn't detected in clinical samples, the developed duplex q-PCR was shown to be an accurate, highly sensitive, and robust diagnostic tool for the detection and quantification of P. acanthamoebae and S. negevensis.

SARS‐CoV‐2 N Gene‐Targeted Anodic Stripping Voltammetry Sensor Using a Novel CoS‐NGQD/Pt@Pd Platform and Au‐DNA‐CdTe QD Probe

Rapid screening of individuals infected with severe acute respiratory syndrome‐coronavirus‐2 (SARS‐CoV‐2) is necessary to contain contagion in a large population. Nucleic acid‐based gold standard assays are time‐consuming, and nucleic acid amplification is mandatory and expensive, impeding the containment of the coronavirus disease 2019 (COVID‐19) outbreak. To overcome the aforementioned disadvantages, this study deals with a specially designed gold (Au)‐deoxyribonucleic acid (DNA)‐cadmium telluride (CdTe) quantum dot (QD) probe to target two sections of the nucleocapsid (N) gene of SARS‐CoV‐2 ribonucleic acid (RNA) of three variants (B.1.1.529, B.1.617.2, and B.1.351). A duplex‐specific nuclease (DSN)‐assisted highly selective release of signaling probes enable higher specificity, and an Au‐supported DNA probe is incorporated to carry many CdTe QD signaling probes. After dissolution, the generated Cd2+ ions are quantified at the novel cobalt sulfide (CoS)‐nitrogen‐doped graphene QD (NGQD)/platinum (Pt)@palladium (Pd) electrode with extraordinary sensitivity through square wave anodic stripping voltammetry (SWASV). The developed sensor exhibits a wide range of detection (10 to 108 copies µL−1) and a lower detection limit (0.12 copies µL−1), without any amplification. The selectivity of the sensor is tested against MERS and HCoV‐NL63, and real‐time detection is performed on heat‐inactivated viral samples, which show excellent selectivity. [ FROM AUTHOR]

Venous thromboembolism in hospitalized patients with COVID-19

Background: The aim of the study was to analyze the results of treatment and the factors of risk of mortality among the patients with COVID-19 and venous thromboembolism (VTE). Method(s): Retrospective analysis of 87 patients with COVID-19 and VTE treated fromApril 2020 to March 2021 in the City HospitalNo40 of Yekaterinburg was performed. Demographic, clinical and laboratory data, including duplex ultrasound, multispiral CT pulmonary angiograms, Charlson index were retrieved. Comparisons were made between two groups: Survivors (n=39) and deceased (n=48). Statistical processing was performed using EZR v. 4.1.2. Result(s): Among 87 patients hospitalized with COVID- 19 (average age - 68 years, 48(55,2%) women) intensive care was required for 61(70,1%) patients. Duplex ultrasound showed 74 deep vein thrombosis (DVT);53(71,6%) - distal DVT, 13(17,6%) - femoropopliteal DVT, 1(1,3%) - iliofemoral DVT, 3(4,1%) - surface veins of extremities, 3(4,1%) - inferior vena cava, 1(1,3%) - right heart chambers. Bilateral lower extremity DVT developed in 29(39,7%) patients. Isolated venous thrombosis was observed in 47(54,1%) patients. 27(31,0%) patients with venous thrombosis had pulmonary embolism (PE), in 13(14,9%) patients with PE the source was not found. In 17(23%) cases DVT had free flotation. Before the identification of VTE, all patients were receiving direct anticoagulants: Prophylactic doses - 31(35,6%), intermediate doses - 33(37,9%), therapeutic doses - 23(26,4%). Deceased patients were older than survivors (average age - 71,0 years (IQR 61,0;81,2);r=0,012). Severe COVID-19, requirement of intensive therapy and mechanical lung ventilation prevailed in the group of deceased (p<0,001). Charlson index was higher in the group of deceased (6,0 (IQR 4,0;6,2) vs 4,0 (IQR 2,0;5,0);p=0,007). The index of 5 points and more showed that the probability of death increased by 3 times (OR 2,98, 95% SI 1,2-7,1;p=0,01). In the structure of VTE in the group of deceased, DVT was mainly in combination with PE of 20(47,1%);p=0,027. In both groups, distal DVT was more frequent: 21(61,8%) among the survivors, 32(80,0%) among the deceased. The localization of venous thrombosis, flotation, bilateral DVT of the lower extremities were not statistically different in the studied groups. Laboratory indicators: D-dimer (3955.0 ng/ml (IQR 2550,0;5325,0) - survivors, 4600,0 ng/ml (IQR 2745,0;38300,0) - deceased;p=0,291);hyperfibrinogenemy (5,2 g/l (IQR 4,0;6,5) - deceased;r=0,033). The dose of direct anticoagulants before the identification VTE was not statistically significant. Conclusion(s): The factors of unfavorable outcome in patients of COVID-19 and VTE were: Age, Charlson index higher than 4 points, intensive therapy, mechanical lung ventilation, DVT in combination with PE. The dose of direct anticoagulants before to the detection of VTE did not affect the unfavorable outcome in the studied category of patients.

THAT BLOODY CLOT DOES NOT ALWAYS NEED A TUNNEL TO MIGRATE: A CASE OF MULTIPLE ARTERIAL AND VENOUS THROMBOSIS IN A COVID-19 PATIENT

SESSION TITLE: Rare Cases of Nervous System and Thrombotic Complication Posters SESSION TYPE: Case Report Posters PRESENTED ON: 10/17/2022 12:15 pm - 01:15 pm INTRODUCTION: Covid 19 virus has impacted nearly 450 million people across the globe;ranging from an asymptomatic carrier state to respiratory symptoms, cardiovascular symptoms, hematologic manifestations and multiorgan failure to death. Thrombotic events are one of its devastating complications. CASE PRESENTATION: A 66 year old man with a history of diabetes mellitus, hypertension and 30 pack years smoking history presented to the emergency room with hypoxia and altered mental status. On exam, his GCS was 8/15 and oxygen saturation was 85% on room air. He was subsequently intubated. CTA chest demonstrated bilateral diffuse ground glass opacities and left pulmonary embolism (PE). CT abdomen and pelvis showed multifocal infarcts in the right kidney with findings suggestive of renal artery thrombosis. Initial platelet count was 80,000/ul with creatinine of 3.9 mg/dl and creatine kinase (CK) of 3977 u/l. His INR was 1.4. Patient was not a candidate for thrombolysis given his thrombocytopenia. He was started on intravenous (IV) heparin and given IV hydration. On day 3 of his admission, he developed dry gangrene of the toes. Ankle brachial index of the right lower extremity (LE) was 1.16 and left LE was 0. Duplex ultrasonography of left LE showed mid to distal popliteal artery thrombus occluding below knee popliteal and tibial arteries. Echocardiogram showed ejection fraction of 55% and bubble study was negative for any intra atrial or pulmonary shunting. On day 4 of his admission, he developed oliguria and his gangrene got worse. His platelet counts decreased to 36,000/ul. Other pertinent labs showed INR 1.2, PT 15.3, PTT 34, D dimer 14.82, fibrinogen 498, CK 6434 mg/dl, hemoglobin 13.2 g/dl, haptoglobin 243 mg/dl and LDH 1041 U/l. Given his poor prognosis in the setting of ventilator dependent respiratory failure, multiple thrombosis and kidney failure requiring hemodialysis, the family decided to withdraw care. DISCUSSION: There are multiple hypotheses of thrombus formation in Covid 19 infection such as interleukin 6 and other cytokines induced endothelial injury, angiogenesis and elevated prothrombotic factors such as factor VIII and fibrinogen. Our patient had PE, renal artery thrombosis and popliteal artery thrombosis. Despite being on full dose anticoagulation, he developed gangrene of the toes. His lab results were not consistent with disseminated intravascular coagulation, thrombotic thrombocytopenic purpura and he was not known to have any baseline hypercoagulable disorder. He did not have any intra cardiac shunts. Hence, it is most likely Covid 19 induced multiple arterial and venous thrombosis. CONCLUSIONS: The treatment of Covid 19 related thrombosis has become very challenging especially in the setting of multiple clots. It is crucial to have large multicenter studies to investigate vascular complications of Covid-19 and to formulate management strategies to ensure good patient outcomes. Reference #1: https://www.nejm.org/doi/full/10.1056/nejmoa2015432 Reference #2: https://journal.chestnet.org/article/S0012-3692(21)01126-0/fulltext DISCLOSURES: No relevant relationships by Devashish Desai No relevant relationships by Swe Swe Hlaing no disclosure on file for Jean Marie Koka;No relevant relationships by Hui Chong Lau No relevant relationships by Subha Saeed No relevant relationships by Anupam Sharma No relevant relationships by Muhammad Moiz Tahir

Development of a duplex real-time PCR assay for simultaneous detection and differentiation of Theileria equi and Babesia caballi.

Equine Piroplasmosis (EP) is a tick-borne disease caused by three apicomplexan protozoan parasites, Theileria equi (T. equi), Babesia caballi (B. caballi) and T. haneyi, which can cause similar clinical symptoms. There are five known 18S rRNA genotypes of T. equi group (including T. haneyi) and three of B. caballi. Real-time PCR methods for detecting EP based on 18S rRNA analysis have been developed, but these methods cannot detect all genotypes of EP in China, especially genotype A of T. equi. In this study, a duplex real-time PCR detection method was developed for the simultaneous detection and differentiation of T. equi and B. caballi. The primers and probes for this duplex real-time PCR assay were designed based on the conserved 18S rRNA gene sequences of all genotypes of T. equi and B. caballi including Chinese strain. Double-quenched probes were used in this method, which provide less background and more signal to decrease the number of false positives relative to single-quenched probes. The newly developed real-time PCR assays exhibited good specificity, sensitivity, repeatability and reproducibility. The real-time PCR assays were further validated by comparison with a nested PCR assay and a previous developed real-time PCR for EP and sequencing results in the analysis of 506 clinical samples collected from 2019 to 2020 in eleven provinces and regions of China. Based on clinical performance, the agreements between the duplex real-time PCR assay and the nPCR assay or the previous developed real-time PCR assay were 92.5% (T. equi) and 99.4% (B. caballi) or 87.4% (T. equi) and 97.2% (B. caballi). The detection results showed that the positivity rate of T. equi was 43.87% (222/506) (10 genotype A, 1 genotype B, 4 genotype C, 207 genotype E), while that of B. caballi was 5.10% (26/506) (26 genotype A), and the rate of T. equi and B. caballi co-infection was 2.40% (12/506). The established method could contribute to the accurate diagnosis, pathogenic surveillance and epidemiological investigation of T. equi and B. caballi infections in horses.

Successful Treatment of Paraplegia Caused by Venous Hypertension Secondary to Extensive Iliocaval Deep Venous Thrombosis in the Absence of Phlegmasia Cerulea Dolens

Objectives: Paraplegia is known to complicate extensive iliocaval and lower extremity deep vein thrombosis (DVT) in rare instances. The most common pathophysiology is ischemia from severe venous hypertension in phlegmasia cerulea dolens. Less understood, however, is paresis or paraplegia in the absence of ischemia. We present a case of paraplegia in extensive iliocaval and lower extremity DVT without ischemia, which was successfully treated by percutaneous pharmacomechanical therapy. Methods: A 46-year-old African American woman with a history of hypertension, insulin-dependent diabetes mellitus, indwelling inferior vena cava filter since 2005, and recent coronavirus disease 2019 diagnosis, presented with acute abdominal pain with severe bilateral lower extremity edema, pain, and paresis. She was found to have bilateral iliocaval to tibial DVT (Fig 1). The patient was noted to have multiphasic arterial waveforms on ankle-brachial index and duplex ultrasound examination. Paresis quickly progressed to flaccid bilateral lower extremity paralysis. Neurologic workup was unrevealing. Despite her symptoms, thrombolytic therapy was delayed due to severe menstrual bleeding requiring a blood transfusion. Therapeutic anticoagulation was initiated. Results: On hospital day 10, the patient underwent 24-hour catheter-directed thrombolysis via bilateral popliteal vein access. Bilateral mechanical thrombectomy was then performed, achieving recanalization of the bilateral lower extremities, iliac veins, and inferior vena cava with minimal residual thrombus (Fig 2). The patient's edema and sensorimotor function immediately improved and never incurred lower extremity tissue ischemia. She was discharged on lifelong rivaroxaban. With physical therapy, the patient ambulated independently at 1 month postoperatively. Venous duplex ultrasound examination revealed continued iliocaval and lower extremity patency at 6 months postoperatively. Conclusions: We postulate that this patient suffered lower extremity paralysis secondary to cauda equina syndrome. Pharmacomechanical thrombectomy is a pragmatic means that reestablishes venous patency and relieves venous hypertension. This pathophysiology and its treatment should be considered in extensive iliocaval DVT and lower extremity neurologic compromise despite duration of paralysis. [Formula presented] [Formula presented]

Effect of the COVID-19 Pandemic on Infra-Inguinal Vein Bypass Graft Surveillance DUS Scans: Single Unit Experience

Introduction: Vein bypass graft surveillance has been a controversy in the literature. The pros are that it allows early detecting of graft complications and intervention;however, the cons are its time and cost burden, without significant benefit. The established practice in our unit is to offer duplex ultrasound (DUS) scan for patients who had infra-inguinal bypass surgeries using autogenous veins for the first two years. Scans should be done at six weeks, three months, six months, one year, and two year intervals. Our aim was to compare graft surveillance scan practice during the COVID-19 pandemic to practice in previous years. Methods and data collection: We collected the data from electronic patients records completed by the unit for two years on 2016 – 2017 and from March 2020 until the end of February 2021. Patients who are scheduled for scans but have not yet reached the scan date were counted in the non-applicable group. We included only the first six months of graft surveillance. Results: Thirty-four patients had bypass at this time;four patients were excluded as two of them had predischarge occlusion, one was done using synthetic graft, and a patient passed away after discharge. The six week scan compliance was as follows: 17 scans were successfully done;11 scans were not done;and five patients were non-applicable. The three month scan compliance was as follows: 11 scans were successfully done, 17 scans were not done, and five scans were non-applicable. The six month scan compliance was distributed as follows: one scan was done;19 scans were not done;and 12 scans were non-applicable. Graft surveillance compliance percentages between 2016 and 2017 to the COVID-19 pandemic year (March 2020 – March 2021) were as follows: six week scan compliance was 86% – 60%;three month scan compliance was 79% – 42%;and six month compliance was 76% – 39%. Six interventions were offered to bypass patients based on the findings of DUS graft surveillance: five patients had graft angioplasty and one had graft thrombolysis. A predischarge surveillance scan was done for 12 patients, representing 36% of the total number of patients. The findings detected three patent grafts, one graft had mild stenosis, five grafts had moderate stenosis, one graft had severe stenosis, and two grafts had complete occlusion. Conclusion and recommendations: The COVID-19 pandemic has negatively affected the number of graft surveillance scans. A predischarge graft surveillance scan was found useful as it detected significant findings that could be missed upon discharge. Therefore, when there are constraints on getting surveillance scans during the COVID-19 pandemic, it is reasonable to do predischarge scans to identify patients who would be at a higher risk of developing graft complications. [Formula presented]

Venous Thrombo-Embolism in an Outpatient Vascular Ultrasound Laboratory All-Comers Cohort with Recent COVID-19 Infection

Introduction: Inflammatory mechanisms triggered and supported by SARS-CoV-2 infection can increase venous thromb-oembolism (VTE) risk. Aim: The aim of the present comparative study was to report on the incidence of VTE in a prospective consecutive series of COVID-19 negative outpatients referred to our vascular ultrasound laboratory for suspected VTE with (COVID+) or without (COVID–) recent SARS-CoV-2 infection during the COVID-19 pandemic. Methods: All patients included in the present study were assessed by duplex ultrasound for the detection of VTE. The following data were collected for each patient: time from first negative swab after COVID-19 infection and VTE diagnosis in COVID+ patients;administration of heparin prophylaxis during COVID-19 infection;presence of venous risk factors (previous VTE episode, chronic venous insufficiency, thrombophilia, recent surgery with prolonged immobilisation, history of malignancy, need for high dose steroid therapy, and dehydration during infection). Rate of VTE was detected and compared between the two groups of COVID+ or COVID– patients by chi square test for categorical data. The presence of risk factors for VTE were analysed as related to VTE occurrence in both groups. Results: From 1 February 2021 to 31 March 2021, 34 patients were included in the study. Among them eight had previous SARS-CoV-2 infection and were negative at the time of investigation. In COVID+ patients, time from first negative swab after COVID-19 infection and VTE diagnosis ranged between 3 and 50 days (mean 17 ± 14.39 days), and 12.5% (n = 1/8) had heparin prophylaxis during infection. Risk factors for VTE were detected in all COVID+ patients and 80.7% (n = 21/26) of COVID– patients. Rate of VTE was 87.5% (n = 7/8) in COVID+ patients and 11.5% (n = 3/26) in COVID– patients (odds ratio 53.66, 95% confidence interval 4.79 – 601.23;p <.001). In the COVID+ population only one patient receiving heparin prophylaxis during infection did not present with VTE. One COVID+ patient presented with both arterial and venous popliteal thrombosis. By comparing directly COVID+ patients with no heparin prophylaxis and venous risk factors (n = 7) to COVID– patients with venous risk factors (n = 21), VTE risk was strongly associated with the presence of previous SARS-CoV-2 infection without proper heparin prophylactic administration (p <.001). Conclusion: In this preliminary series presence of risk factors for VTE and recent SARS-CoV-2 infection with no heparin prophylaxis is strongly associated with VTE occurrence. COVID-19 outpatients should be treated by prophylactic heparin whenever VTE risk factors are detected and duplex ultrasound cannot be performed to exclude a VTE episode, so that physicians treating outpatients should be aware of the VTE risk in those patients. Both arterial and venous conditions prone to thrombosis should be fully assessed in patients when diagnosing a new SARS-CoV-2 infection.

UPPER EXTREMITY DEEP VENOUS THROMBOSIS AFTER BNT162B2 MRNA COVID-19 VACCINE

CASE: A 60-year-old woman with past medical history including hypertension, nephrolithiasis, and Covid-19 4 months prior presented to the emergency department with 3 days of substernal chest pain radiating toward the back and shoulders 6 days after receiving her second dose of the BNT162b2 mRNA Covid-19 vaccine (Pfizer/BioNTech) in her left deltoid. The patient tested negative for Covid-19 and denied shortness of breath, cough, fever, or dyspnea on exertion. Her ECG was notable for more pronounced t-wave inversions in III and aVF, but further cardiac workup was unremarkable, and she was discharged the next day. The patient re-presented to care 6 days later with left arm pain, erythema, edema, and warmth. Her left bicep circumference was 31cm versus 28cm on the right. Upper extremity duplex ultrasound (US) was remarkable for deep venous thrombosis (DVT) of the left internal jugular, subclavian, axillary, and basilic veins. MRI angiogram was confirmatory. Other than her occupation as a hairdresser, the patient did not have known risk factors for DVT: no personal or family history of thromboembolism, no tobacco use, took no prescription medications, and had received all ageappropriate cancer screening. Her thrombophilia workup was negative. The patient was discharged on apixaban. Eleven days later, a venogram showed persistent clot burden in the left axillary, mid-subclavian, and brachiocephalic veins. Thrombectomy, overnight tPA infusion, and left subclavian vein stenting were performed and the patient was discharged on daily apixaban and aspirin. IMPACT/DISCUSSION: As of December 10th, 2021, the vaccine adverse event reporting system yielded 464 reports of “thrombosis” after the Pfizer/ BioNTech vaccine in individuals with no reported current illness, 32 of which occurred in the upper extremity. To our knowledge, our patient represents the first report of upper extremity deep venous thrombosis (UEDVT) shortly after receipt of the Pfizer vaccine in an otherwise healthy person. UEDVT is relatively rare: it occurs in about 0.4 to 1 per 10,000 people per year and less than 20% of incidents are idiopathic. Given the scarcity of potential causes, our case may simply reflect expected background incidence. Nevertheless, the literature includes multiple case reports of DVT after mRNA Covid-19 vaccination, including cases of lower extremity DVT and DVT with pulmonary embolism (PE) after the Pfizer vaccine, and cases of lower extremity DVT, PE, and UEDVT after the mRNA-1273 (Moderna) vaccine. Given the similar mechanism of action between the Moderna and Pfizer vaccines, it is possible that the same pathophysiology underlies the reports of DVT in these vaccine recipients. CONCLUSION: Upper extremity deep venous thrombosis after Covid-19 mRNA vaccination should remain on the differential as clinicians assess chest and arm pain following vaccination. There is a possible association between the BNT162b2 mRNA Covid-19 vaccine and upper extremity deep venous thrombosis that requires further research.

Comparative retrospective analysis of the prevalence of deep vein thrombosis detected by duplex ultrasound in primary health care facilities.

Aim. To identify the incidence of deep vein thrombosis (DVT) as determined by compressive duplex ultrasound in primary health care facilities before and during the pandemic of a coronavirus disease 2019 (COVID-19). Material and methods. This retrospective analysis of 1492 duplex ultrasound examinations in 2018 and 1710 examinations in 2020-2021 in patients with suspected lower limb DVT was performed. The studies were carried out in 4 primary health care institutions in different cities of the Republic of Uzbekistan. Each patient underwent duplex ultrasound of lower limb deep veins. Most patients had complaints (swelling, leg pain) (symptomatic patients), while some individuals underwent preventive screening before various surgical interventions and had no complaints (asymptomatic patients). Results. According to the data obtained, the prevalence of DVT increased significantly during the pandemic — from 3,9 to 15,1% (χ2 =113,23, p<0,001). During the pandemic, there was a trend towards a relative increase in lower limb DVT among the elderly and young people — by 8,7 and 3,8%, respectively (χ2 =1,66, p=0,19 and χ2 =0,64, p=0,42, respectively). No dependence of the prevalence of lower limb DVT on the sex was found (χ2 =0,02, r=0,9). Conclusion. The study results showed an increase in the prevalence of lower limb DVT during the COVID-19 pandemic. Further clinical studies are needed to analyze the lower limb DVT during this period. (English) [ FROM AUTHOR] Цель. Выявить частоту тромбоза глубоких вен (ТГВ), установленного методом компрессионного дуплексного сканирования в учреждениях первичного звена здравоохранения в предпандемийный период и во время пандемии новой коронавирусной инфекции (COVID-19 (COronaVIrus Disease 2019)). Материал и методы. Проведен ретроспективный анализ результатов 1492 дуплексных ультразвуковых исследований в 2018г и 1710 исследований в 2020-2021гг у пациентов с подозрением на ТГВ нижних конечностей (НК). Исследования проводились в 4-х медицинских учреждениях первичного звена здравоохранения в разных городах Республики Узбекистан. Каждому пациенту проводилось дуплексное сканирование глубоких вен НК. Большинство пациентов имели жалобы (отек, боль в ногах) (симптомные пациенты), некоторые прошли профилактический скрининг перед различными оперативными вмешательствами и не имели жалоб (бессимптомные пациенты). Результаты. По полученным данным частота ТГВ статистически значимо возросла во время пандемии — от 3,9 до 15,1% (χ2 =113,23, p<0,001). В период пандемии отмечалась тенденция к относительному увеличению частоты ТГВ НК среди пожилых и молодых людей — на 8,7 и 3,8%, соответственно (χ2 =1,66, р=0,19 и χ2 =0,64, р=0,42, соответственно). Зависимости частоты ТГВ НК от пола пациента не обнаружено (χ2 =0,02, r=0,9). Заключение. Результаты проведенного исследования показали увеличение частоты выявления ТГВ НК в период пандемии COVID-19. Необходимы дальнейшие клинические исследования для анализа частоты ТГВ НК в этот период. (Russian) [ FROM AUTHOR] Copyright of Cardiovascular Therapy & Prevention is the property of Silicea-Poligraf LLC and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full . (Copyright applies to all s.)

COVID-19 ASSOCIATED REDUCTION IN HYPERTENSION-RELATED DIAGNOSTIC AND THERAPEUTIC PROCEDURES IN EXCELLENCE CENTERS OF THE EUROPEAN SOCIETY OF HYPERTENSION

Objective: The Covid-19 pandemic necessitated a decrease in non-Covid-19 related diagnostic and therapeutic procedures in many countries. We explored the impact on hypertension care in the Excellence Center (EC) network of the European Society of Hypertension. Design and method: We conducted an electronic survey regarding 6 key procedures in hypertension care among our ECs. Results: Overall, 54 ECs from 18 European and 3 non-European countries participated. From 2019 to 2020, there was a significant decrease in the median number per center of ambulatory blood pressure monitorings (ABPMs;544 vs 289), duplex ultrasound investigations of renal arteries (DUS RA;88.5 vs 55), computed tomographic investigations of renal arteries (CT RA;66 vs 19.5), percutaneous renal artery angioplasties (PTA RA;5 vs 1), laboratory tests for catecholamines (2019: 116 vs 67.5) and for ennin/aldosterone (146 vs 83.5). All comparisons were statistically significant with p < 0.001, respectively (Figure). While the reduction in all diagnostic and therapeutic procedures was observed in all 3-months period comparisons between 2019 and 2020, the most profound decrease occurred from April to June 2020, which was the period of the first wave and the first lockdown in most countries. In this period, as compared to 2019, the median reduction in 2020 was 50.7% (ABPM), 47.1% (DUS RA), 50% (CT RA), 57.1% (PTA RA), 46.9% (catecholamines) and 41% ( ennin/aldosterone), respectively. Based on Friedman test, overall differences in reduction between 3 months time intervals were statistically highly significant. Conclusions: Diagnostic and therapeutic procedures related to hypertension were dramatically reduced during the first year of the Covid-19 pandemic, with the largest reduction during the first lockdown. The long-term consequences regarding blood pressure control and, ultimately, cardiovascular events remain to be investigated.

Duplex One-Step RT-qPCR Assays for Simultaneous Detection of Genomic and Subgenomic RNAs of SARS-CoV-2 Variants.

A hallmark of severe acute respiratory syndrome virus (SARS-CoV-2) replication is the discontinuous transcription of open reading frames (ORFs) encoding structural virus proteins. Real-time reverse transcription PCR (RT-qPCR) assays in previous publications used either single or multiplex assays for SARS-CoV-2 genomic RNA detection and a singleplex approach for subgenomic RNA detection. Although multiplex approaches often target multiple genomic RNA segments, an assay that concurrently detects genomic and subgenomic targets has been lacking. To bridge this gap, we developed two duplex one-step RT-qPCR assays that detect SARS-CoV-2 genomic ORF1a and either subgenomic spike or subgenomic ORF3a RNAs. All primers and probes for our assays were designed to bind to variants of SARS-CoV-2. In this study, our assays successfully detected SARS-CoV-2 Washington strain and delta variant isolates at various time points during the course of live virus infection in vitro. The ability to quantify subgenomic SARS-CoV-2 RNA is important, as it may indicate the presence of active replication, particularly in samples collected longitudinally. Furthermore, specific detection of genomic and subgenomic RNAs simultaneously in a single reaction increases assay efficiency, potentially leading to expedited lucidity about viral replication and pathogenesis of any variant of SARS-CoV-2.

Intracellular flow cytometry complements RT-qPCR detection of circulating SARS-CoV-2 variants of concern.

Basic and antiviral research on SARS-CoV-2 rely on cellular assays of virus replication in vitro. In addition, accurate detection of virus-infected cells and released virus particles is needed to study virus replication and to profile new candidate antiviral drugs. Here, by flow cytometry, we detect SARS-CoV-2 infection at single cell level and distinguish infected Vero E6 cells from uninfected bystander cells. Furthermore, based on the viral nucleocapsid expression, subpopulations of infected cells that are in an early or late phase of viral replication can be differentiated. Importantly, this flow cytometric technique complements our duplex RT-qPCR detection of viral E and N, and it can be applied to all current SARS-CoV-2 variants of concern, including the highly mutated Omicron variant.

Asymptomatic deep vein thromboses in prolonged hospitalized COVID-19 patients.

High incidence of venous thromboembolic (VTE) events in coronavirus disease 2019 (COVID-19) patients has been reported despite pharmacologic thromboprophylaxis. We performed prospective bilateral lower extremity ultrasound evaluation of prolonged hospitalized COVID-19 ward patients from our institution without clinical suspicion of deep vein thrombosis (DVT).A total of 102 patient were included in the study. All patients were receiving pharmacologic thromboprophylaxis, the majority in intermediate or therapeutic doses. Asymptomatic DVT was detected in 26/102 (25.5%) patients: 22 had distal and four had proximal DVT, six had bilateral leg involvement. Pulmonary embolism was highly prevalent (17/70, 24.3%) but similarly grouped among patients with and without asymptomatic DVT. In total 37.2% of patients included in the study were recognized as having VTE.Asymptomatic DVT events were more common in intensive care unit (ICU) survivors (60% in postmechanically ventilated ICU survivors, 21.2% in ward patients, 22% in high-flow oxygen treated patients; P = 0.031), in patients with higher modified International Medical Prevention Registry on Venous Thromboembolism (IMPROVE) VTE risk-score (median 3 vs. 2 points with and without DVT; P = 0.021) and higher body temperature on admission (median 38.7 °C vs. 37.7 °C with and without DVT; P = 0.001). No clear associations with Padua VTE risk score, demographic and other clinical characteristics, intensity of thromboprophylaxis, severity of other COVID-19 symptoms, degree of systemic inflammation or D­dimers on admission were found (P > 0.05 for all analyses).Systematic ultrasound assessment in prolonged hospitalized severe COVID-19 patients prior to hospital discharge is needed, especially in ICU survivors, to timely recognize and appropriately treat patients with asymptomatic DVT.