PCR Test Research Articles

Suspected Cases of Chlamydia-Induced Fertility Problems in Sows: What Is the Approach of Austrian Practitioners?

Although it is generally accepted that Chlamydia spp. can induce fertility problems in sows, many questions concerning diagnostic, therapeutic, and prophylactic approaches remain unanswered. The aim of this study was to gather information about how swine practitioners deal with these knowledge gaps. Austrian swine practitioners were anonymously asked for their assessment of chlamydiosis using the Lime Survey program. PCR tests are used for molecular chlamydia detection from abortion material (64.3%), vaginal swabs (40.5%), and the genital tract (postmortem slaughterhouse samples) (9.5%). Treatment mainly includes tetracyclines (95.2%), occasionally macrolides (2.4%) or other (not mentioned) antibiotic groups (7.1%). Treatment periods vary greatly, from 5 to 10 days (23.8%), from 11 to 15 days (47.6%), from 16 to 21 days (11.9%), and >21 days (9.5%). Prophylactic measures include hygiene improvement, rodent control, and herd treatments with tetracyclines. Serological methods are perceived as inadequate, and there is a general need for better guidance on interpreting results. Knowledge about the increased incidence of tetracycline-resistant Chlamydia suis strains is limited. Weighing up economic interests, feasibility, and animal welfare plays an important role. Further research to improve diagnosis and prevention of chlamydia-related fertility diseases in sows is urgently needed.

Presence of Enterococci in Cats with Urinary Tract Infections and Antibiotic Resistance Profiles

Background: Bacterial urinary tract infection (UTI) is one of the most common diseases in cats caused by an imbalance between the natural host defence mechanisms of the urinary tract and virulent bacteria. This study investigated the prevalence and antimicrobial susceptibility of enterococci isolated from cats presenting urinary tract problems to veterinary clinics in Istanbul. Methods: 100 urine samples were collected from cats by cystocentesis and plated on blood agar. Isolates found catalase and oxidase negative plated on Enterococcocel agar. After incubation, the isolates formed black colonies were tested by PCR using E. faecium and E. faecalis-specific primers. Then the antibiotic susceptibility of the isolates was determined. Result: Eight of the 100 isolates that formed black colonies on Enterococcosel agar were identified as Enterococcus spp. After PCR tests, six isolates were classified as E. faecalis, one isolate was classified as E. faecium and one isolate that did not form a band was classified as Enterococcus spp. As a result of the Kirby-Bauer disc diffusion method used to determine the antibiotic susceptibility of the isolates, most resistance was found to cephalothin (100%) and penicillin, erythromycin, clindamycin and tetracycline (87.5%). While vancomycin resistance was not observed in any isolate, the lowest resistance was to enrofloxacin and ciprofloxacin (33.3%). Multiple antibiotic resistance was observed in all (100%) isolates. The presence of multi-resistant enterococci in cats represents a significant health threat. Bacterial identification and antimicrobial susceptibility testing should always be performed in urinary tract infections to guide definitive diagnosis and treatment.

Illustration of the Financial Impact of Usage of Initial Specimen Diversion Devices (ISDD) or Continued Educational Initiatives on the Costs from Blood Culture Contamination

Abstract Introduction/Objective Blood culture contamination (BCC) has the potential to lead to additional healthcare system costs and harm to patient. Two ways to reduce BCC have been explored in the literature and are economically modeled including initial specimen diversion devices (ISDD) and continued education on phlebotomy. Methods/Case Report A literature review on the economic costs of BCC, ISDD efficacy, and phlebotomy education to reduce BCC rates was undertaken. The reported average BCC at a veteran affairs medical center (VAMC), its reported rate during a month of phlebotomy training, the cost of an ISDD device at VAMC, and the average wage for 4 hours of phlebotomy manager time for training was noted. An illustrative economic model involving 100 patients was used at BCC rates of 1%, 2%, and 3% along with what had been observed at the VAMC was used for economic modeling. Results (if a Case Study enter NA) It was previously reported by Skoglund et al that in laboratories with rapid diagnostic testing modalities (such as MALDI-TOF or PCR testing), the cost per BCC event is $13,026 versus $8,287 in costs for a negative blood culture. ISDD efficacy in the literature reviewed (74.1% to 87.5%), continuous education efficacy (16.2% to 57.8%), ISDD device cost ($17.70 per device), and the hourly wage of a phlebotomy supervisor ($29.34) were noted. The reported average BCC rate from a regional VAMC was 1.33%, and during the month with education, the rate was 0.5%. From a model of 100 patients, ISDD device savings ranged from $1,741.60-$2,376.63 (1% BCR), $5,253.20-$6,523.25 (2% BCR), and $8,764.79-$10,669.88 (3% BCR). Education savings ranged from $755.98-$2,727.41 (1% BCR), $1,523.70-5,466.55 (2% BCR), and $2,291.42-$8,205.69 (3% BCR). The observed impact of education at the regional VAMC would have resulted in savings of $650.83 per 100 patients. Conclusion Both education and ISDD devices can result in significant savings from the costs of BCC.

Evaluation of Loopamp Leishmania detection kit for the diagnosis of cutaneous leishmaniasis in Ethiopia

BackgroundCutaneous leishmaniasis (CL) in Ethiopia and some parts of Kenya is predominantly caused by Leishmania aethiopica. While skin-slit (SS) microscopy is routinely used for CL diagnosis, more sensitive molecular tests are available. The Loopamp™ Leishmania detection kit (Loopamp) is a robust loop-mediated isothermal amplification (LAMP) assay with the potential for implementation in primary healthcare facilities. In this study, we comparatively assessed the diagnostic accuracy of four methods currently used to diagnose CL: Loopamp, kinetoplast DNA (kDNA) PCR, spliced leader RNA (SL-RNA) PCR and SS microscopy.MethodsA study on 122 stored tape disc samples of suspected CL patients was conducted in Gondar, northwestern Ethiopia. Routine SS microscopy results were obtained from all patients. Total nucleic acids were extracted from the tapes and subjected to PCR testing targeting kDNA and SL-RNA, and Loopamp. Diagnostic accuracy was calculated with SS microscopy as a reference test. The limit of detection (LoD) of Loopamp and kDNA PCR were determined for cultured L. aethiopica and Leishmania donovani.ResultsOf the 122 patients, 64 (52.5%) were identified as CL cases based on SS microscopy. Although the PCR tests showed a sensitivity of 95.3% (95% confidence interval [CI] 91.6–99.1), Loopamp only had 48.4% (95% CI 39.6–57.3) sensitivity and 87.9% (95% CI 82.1–93.7) specificity. The LoD of Loopamp for L. donovani was 100-fold lower (20 fg/µl) than that for L. aethiopica (2 pg/µl).ConclusionsThe Loopamp™ Leishmania detection kit is not suitable for the diagnosis of CL in Ethiopia, presumably due to a primer mismatch with the L. aethiopica 18S rRNA target. Further research is needed to develop a simple and sensitive point-of-care test that allows the decentralization of CL diagnosis in Ethiopia.Graphical

Adopting Covid-19 and Flu Testing Strategies In An Underserved Community

Abstract Introduction/Objective The COVID-19 pandemic has had a significant impact on communities worldwide, while little is known about how medically underserved minority communities perceive and experience COVID-19 and/or COVID- 19/Flu combined testing. This research target at a medically underserved community in east Harris County, Texas, and was designed to 1) understand the perception of the single COVID-19 test as well as the COVID-19 and flu combined test, 2) implement different testing strategies in collaboration with the local community. Methods/Case Report Residents in the eligible communities who are age ≥ 18 years or older are recruited through community engagement groups and community originations. Participant’s demographic information, perception and histories of vaccination and testing were collected through a cross-sectional questionnaire. Participants willing to undergo testing were assisted by community health workers, who then collected feedback on test acceptability via follow-up questionnaires. Results (if a Case Study enter NA) A total of 325 individuals participated in the study from Sept 2022-June 2023. 52% of participants are minority ethnicity, including 35% African American and 17% Latino. 11% of participants are uninsured and 32% of participants are from households with income lower than 25,000 a year. We surveyed our participants on their opinion of COVID-19 and Flu testing and vaccination, majority of our participants have had more experience in COVID-19 (92% had been tested and 93% had been vaccinated) than Flu (64% had been tested and 85% had been vaccinated). 96% consider COVID-19 testing tolerable, with major barrier of being tested is fear of painful sampling (49%). PCR testing is the dominant testing method in the community (> 40%) for COVID-19, while only 16% received antigen testing. Conclusion Our findings suggest that while participants recognized the role of COVID-19 testing, there were significant barriers to access and uptake of testing, including concerns about painful sampling, inconvenience, and a lack of trust in the healthcare system. Participants also expressed a desire for clearer information about testing and more accessible testing sites. Participants have less experience with Flu testing and many of whom had never been tested in the past. Our study highlights the need for targeted community education and outreach efforts to increase testing uptake. Development and implementation of these efforts would better prepare us for combating other commutable diseases of public health significance in the future.

Clinical Utility of Broad Range PCR for Infectious Organisms

Abstract Broad range PCR is used to identify infectious organisms when bacterial, fungal, and mycobacterial cultures are negative despite clinical evidence of infection. We performed a 6-month retrospective chart review to assess the actionability of broad range PCR and identify opportunities to improve the utility of this referral test at our institution. One hundred eleven broad range PCR tests were ordered from July to December 2023. Seventy-five (67.6%) results did not affect patient management, 18 (16.2%) results affected patient management, and the treatment impact of 18 (16.2%) results is unknown because they were from outside consult cases. There were no instances in which antibiotic therapy was completely discontinued due to a negative broad range PCR result. Concurrent in-house microbiology testing yielded a positive result in 15 (13.5%) cases, of which 8 (53%) had a negative broad range PCR result and 2 (13.3%) had a conflicting broad range PCR result. Thirty-eight (34.2%) specimens were formalin fixed paraffin embedded tissue, 29 (26.1%) were fresh tissue, 13 (11.7%) were ESWABS, and the remaining 31 specimens were various fluid types (27.9%). While fresh tissue is the preferred specimen, broad range PCR detected an organism in 20 (53%) formalin fixed paraffin embedded tissue specimens, 11 (37.9%) fresh tissue specimens, 3 (23.1%) ESWAB specimens, and 2 (6.5%) fluid specimens (abscess aspirate and cerebrospinal fluid). Improvements in physician communication and laboratory workflow are necessary to improve the specimen quality and thus the utility of this referral test at our institution.

SARS-CoV-2 infection and the risk of depressive symptoms: a retrospective longitudinal study from the population-based CONSTANCES cohort.

Should COVID-19 have a direct impact on the risk of depression, it would suggest specific pathways for prevention and treatment. In this retrospective population-based study, we aimed to examine the association of prior SARS-CoV-2 infection with depressive symptoms, distinguishing self-reported v. biologically confirmed COVID-19. 32 007 participants from the SAPRIS survey nested in the French CONSTANCES cohort were included. COVID-19 was measured as followed: ad hoc serologic testing, self-reported PCR or serology positive test results, and self-reported COVID-19. Depressive symptoms were measured with the Center of Epidemiologic Studies-Depression Scale (CES-D). Outcomes were depressive symptoms (total CES-D score, its four dimensions, and clinically significant depressive symptoms) and exposure was prior COVID-19 (no COVID-19/self-reported unconfirmed COVID-19/biologically confirmed COVID-19). In comparison to participants without COVID-19, participants with self-reported unconfirmed COVID-19 and biologically confirmed COVID-19 had higher CES-D scores (β for one interquartile range increase [95% CI]: 0.15 [0.08-0.22] and 0.09 [0.05-0.13], respectively) and somatic complaints dimension scores (0.15 [0.09-0.21] and 0.10 [0.07-0.13]). Only those with self-reported but unconfirmed COVID-19 had higher depressed affect dimension scores (0.08 [0.01-0.14]). Accounting for ad hoc serologic testing only, the CES-D score and the somatic complaints dimension were only associated with the combination of self-reported COVID-19 and negative serology test results. The association between COVID-19 and depressive symptoms was merely driven by somatic symptoms of depression and did not follow a gradient consistent with the hypothesis of a direct impact of SARS-CoV-2 infection on the risk of depression.

Advances in clinical diagnosis and treatment of microsporidial keratitis

Microsporidia, a unicellular eukaryotic microorganism, poses a risk of infecting the eyes, precipitating microsporidia keratitis (MK). This condition typically manifests in two forms: microsporidian keratoconjunctivitis (MKC) or stromal keratitis (MSK). While MKC often resolves spontaneously, it can progress to MSK, a vision-threatening condition that, in severe instances, may lead to corneal perforation. Epidemiological studies reveal that MK is prevalent in Southeast Asia, particularly during the rainy season. Diagnosis encompasses a range of methods, including corneal scraping for microbiological analysis, PCR testing, and advanced imaging techniques such as AS-OCT and IVCM. Therapeutic approaches vary, with MKC typically managed through local and systemic drug therapy, while MSK may necessitate more aggressive interventions, including corneal transplantation. In China, MK case reports are scarce, and physicians still grapple with a lack of comprehensive understanding regarding its clinical presentation, diagnostic strategies, and treatment options. This deficit can lead to missed or misdiagnoses, as well as overtreatment. Consequently, this review endeavors to comprehensively outline the epidemiology, etiology, clinical features, diagnostic modalities, and therapeutic interventions for MK, thereby offering valuable insights and guidance for clinical practice.

Pneumocystis jirovecii pneumonia in hemato-oncological patients; 10 years of experience at Hospital Infantil Teletón de Oncología

Abstract Background Pneumocystis jirovecii pneumonia is an opportunistic infection that most frequently affects immunocompromised people, especially patients with HIV; clinical presentation is nonspecific and mortality rate in patients without HIV ranges are between 30% and 75%. In the international literature, there are few case reports of pediatric oncology patients. A series of cases is presented with the objective of describing the epidemiological characteristics, clinical presentation, treatment, and outcomes of Pneumocystis jirovecii pneumonia in this population group. Methods An observational, descriptive, and retrospective study was carried out in cancer patients under 18 years, with clinically compatible symptoms, an imaging study suggestive of the pneumonia, and identification of P. jirovecii using a PCR test of bronchial aspirate at Hospital Infantil Teletón de Oncología in Querétaro Mexico, since November 2014 to June 2023. Results A total of 17 events were reported with a median age of 9.8 years, 14/17 (82%) with a diagnosis of acute leukemias, 1/17 (6%) Ewing sarcoma, 1/17 (6%) lymphoma large cell anaplastic and 1/17 (6%) neuroblastoma. Regarding their therapeutic phases, 7/17 (41%) were in consolidation, 4/17 (23%) induction, 3/17 (18%) allogeneic HSCT (Hematopoietic stem-cell transplantation), 2/17 (12%) in surveillance, and 1/17 (6%) in maintenance. The clinical presentation in our patients was characterized by the presence of fever in 15/17 (88%), dry cough in 10/17 (59%) and 17/17 (100%) developed acute respiratory distress syndrome, which was classified as mild in 7/17 (41%), moderate in 5/17 (29%) and severe in 5/17 (29%). 13/17 (75%) presented distributive shock. All our patients underwent chest x-rays, 12/17 (70.5%) presented bilateral interstitial infiltrate, 4/17 (23.5%) consolidation, and 1/17 (6%) pleural effusion. Lung CT (computed tomography) was performed in 4/17 (23.5%), and 4/4 (100%) presented ground glass opacities. All of our patients were admitted to the PICU (Pediatric intensive care unit) and 16/17 (94%) required invasive mechanical ventilation. All receive treatment with TMP-SMX (Trimethoprim-sulfamethoxazole). 4/17 patients died (23%) of whom none received prophylaxis. 3/17 patients (18%) received prophylaxis with trimethoprim-sulfamethoxazole (TMP-SMX). Conclusion The present study is the first study in Mexico that compiles cases of Pneumocystis Jirovecii pneumonia in pediatric oncology patients. P. jirovecii pneumonia remains a rare cause, but represents a potentially fatal cause in oncology patients, so it is important to have a high clinical suspicion during the evaluation of an immunocompromised patient with evidence of pneumonia during all treatment phases with chemotherapy, in order to start treatment promptly. Thus, we also emphasize the importance of adequate prophylaxis as the most important preventive measure.

Prevalence of SARS-CoV-2 Among Deported Guatemalan Migrants in Tecun Uman, Guatemala

Abstract Background Migrant populations in Central America face complex disease burdens, including respiratory viruses. A frequently overlooked sub-group within the migrant populations is the returned migrants. Assessment of COVID-19 disease burden and associated risk factors are important for identifying high-risk groups, as well as for informing disease reduction efforts, including vaccination programs. Methods As a sub-study within a larger (AGRI-CASA) cohort study, we screened and offered enrollment to returned migrants on a weekly basis in El Centro de Retornados de Ayutla, located in the Tecun Uman community along the Guatemala-Mexico border. The existing screening program includes symptomatic screening of all returnees for cough or fever; we then performed SARS-CoV-2 rapid antigen testing from nasopharyngeal swab on all symptomatic and 10% of asymptomatic individuals. In our sub-study of this screened population, we collected additional clinical and exposure data and a second mid-turbinate nasal swab on all consenting adults available with COVID-like illness (CLI). The samples underwent PCR testing at the Trifinio Center for Human Development (CHD) laboratory. We did not enroll minors due to uncertainty of legal guardians. Results From January-September 2023, 10,570 Guatemalans were deported through El Centro de Salud de Ayutla, of which 5,373 (50.8%) were tested for SARS-CoV-2 and 42 (0.8%) were positive, a decrease from 2021 (n=40,927 migrants, 12,181 tested, and 714 [5.9%) positive) and 2022 (n=36,865 migrants, 12,406 tested, 403 [3.2%] positive). Our sub-study enrolled 197 (30.1%) of 655 symptomatic participants with CLI from January-September 2023. Of those, median age (SD) was 26 (7.87) years and 174 (88%) were male. The most commonly reported symptoms were cough (34.5%) nasal congestion (27.9%), sore throat (21.3%), and headache (19.3%). Of 197 participants, 163 samples were available for testing; 4 (2.5%) of those were SARS-CoV-2 antigen positive via POC testing and 5 (3.1%) of those samples were PCR-positive at the Trifinio Center for Human Development (CHD) laboratory. The PCR-confirmed SARS-CoV-2 positive samples contained XBB1.15, XBB1.5.11, and XBB.1.5 variants. According to Guatemala’s Laboratorio Nacional de Salud, the main variant in the country in February and June 2023 was Omicron, especially the XBB variant. Conclusion Despite the crowding conditions for deported migrants returning from Mexico, SARS-CoV-2 positivity for symptomatic individuals was relatively low. Further studies should evaluate additional pathogens and migrants traveling in both directions to characterize the full disease burden of this vulnerable population. Combined with additional information on the vaccination status of the deported migrants, these results will be crucial in calculating the number-needed-to-vaccinate (NNV) among the departed migrant populations and the local Guatemalans as well as assessing the cost effectiveness of vaccination programs targeting these populations.

Cytomegalovirus chronic retinal necrosis with ganciclovir resistance: a case report.

Cytomegalovirus (CMV) chronic retinal necrosis (CRN) is a rare viral retinal infection that occurs in mildly immunocompromised people. It shares some features with both acute retinal necrosis and CMV retinitis. It is typically treated with combination intravitreal and systemic ganciclovir. We discuss the management of a case of CMV CRN with ganciclovir resistance. An 80-year-old female presented with one month of blurry vision in the left eye. She was being treated with abatacept, methotrexate, and prednisone for rheumatoid arthritis. Examination revealed anterior chamber and vitreous cell along with peripheral retinal whitening. Fluorescein angiogram showed diffuse retinal non-perfusion. Aqueous fluid PCR testing returned positive for CMV. The retinitis was initially controlled with oral and intravitreal ganciclovir, but then recurred and progressed despite these therapies. Ganciclovir resistance was suspected and the patient was switched to intravitreal foscarnet injections, along with oral letermovir and leflunomide, which lead to resolution of the retinitis. The patient has now continued with letermovir and leflunomide for approximately 2.5 years without reactivation of the retinitis or need for further intravitreal anti-viral injections and with adequate control of her rheumatoid arthritis. The incidence of CMV CRN may increase in the future as the use of non-cytotoxic immunosuppressive therapies that result in relatively mild immunosuppression also increases. Treatment with ganciclovir is effective but frequently leads to resistance, as in our case. In this situation, combination therapy with letermovir and leflunomide, particularly in the setting of rheumatoid arthritis where leflunomide can also have an anti-inflammatory effect, can be considered.

Detection and characterization of HIV-1 group O and HIV-2 in the Central African Republic.

The Central African Republic (CAR) is characterized by widespread HIV epidemic with notable prevalence and genetic diversity. We herein analysed the genetic diversity of atypical non-M HIV-1 strains. In-house serotyping assays for variants of HIV-1 (M, N, O, P) and HIV-2 were used to test a biological collection of 6092 HIV-seropositive blood samples collected between 2003 and 2014 at the Institut Pasteur de Bangui. Samples indicative of recombinant M/O groups, HIV-2, or those that yield doubtful/negative results underwent further PCR tests and sequencing. We found six atypical HIV strains: specifically, three (0.05%) HIV-1 group O strains (subtype H) detected in samples from 2005, 2008 and 2009, alongside three (0.05%) HIV-2 strains (two group A and one group B) identified in samples from 2007 and 2009. HIV-1/O strains showed a genetic link to Cameroon and Gabon strains. This study highlights the dominance of HIV-1/M in the CAR's HIV epidemic over time and underscores the infrequent occurrence of HIV-1 group O and HIV-2 strains. These findings validate the efficacy of WHO-recommended HIV testing protocols and emphasize the need for adaptive surveillance and management strategies to confront the complexities introduced by the genetic diversity of HIV strains.

8278 Myxedema Coma With COVID-19 Infection

Abstract Disclosure: R. Kalas: None. V. Manohar: None. N.S. Bapat: None. Y. Kim: None. Myxedema coma is a rare medical emergency that occurs in severe hypothyroidism with a mortality rate reaching 60%. The hallmarks include depressed mental status and hypothermia, but hypotension, hyponatremia, bradycardia, and hypoventilation can also occur. This complication can occur in patients with a longstanding history of severe hypothyroidism especially when precipitated by an acute event such as infection, myocardial infarction, or surgery. Prompt recognition and treatment of the condition is important to prevent fatal outcomes. We present a case of myxedema coma in the setting of COVID-19 infection and nonadherence to medication. A 91-year-old woman with history of hypothyroidism, hypertension, coronary artery disease, cardiomyopathy, and chronic obstructive pulmonary disease presented with dyspnea. Her home medications included levothyroxine 75 mcg daily, which she reported not taking for at least 3 weeks. Vital signs revealed a normal heart rate and blood pressure of 84 beats/min and 137/78 mmHg, but the patient was hypothermic to 35.7 °C. On exam, she was somnolent. Labs were notable for markedly elevated TSH level of 111 (0.380 - 4.700 uIU/mL), low free T4 level of 0.57 (0.93 - 1.70 ng/dL), random cortisol level of 25.5 (6.2-18.4 ug/dL), and positive SARS-COV-2 PCR test. Chest x-ray revealed bilateral lower lung parenchymal consolidations and arterial blood gas revealed a pH of 7.29 and pCO2 of 52.5 (35-45 mm Hg), indicating hypercapnia. She was treated for COVID pneumonia and given concern for COVID induced myxedema coma she was given IV levothyroxine 100 mcg for 2 days, after which the patient’s hypothermia and somnolence improved, with an improved repeat free T4 level 0.87 ng/dL. She was subsequently transitioned to oral levothyroxine 112 mcg daily and discharged after improvement of her COVID symptoms. Myxedema coma is a rare complication of decompensated hypothyroidism, frequently triggered by an infection. Thyroid hormone plays an important role in regulating both the innate and adaptive immune systems. As a result, individuals with hypothyroidism are at increased risk of acquiring infections, which can further lead to myxedema coma. COVID-19 infection has been associated with thyrotoxicosis, thyroiditis and hypothyroidism. Given the associated high mortality rate, regardless of the underlying precipitant, prompt diagnosis and early administration of thyroid hormone replacement therapy is crucial to improve outcomes. Presentation: 6/3/2024

Comparing AI/ML approaches and classical regression for predictive modeling using large population health databases: Applications to COVID-19 case prediction

BackgroundResearch comparing artificial intelligence and machine learning (AI/ML) methods with classical statistical methods applied to large population health databases is limited. ObjectivesThis retrospective cohort study aimed to compare the predictive performance of AI/ML algorithms against conventional multivariate logistic regression models using linked health administrative data. MethodsUsing Ontario's population health databases, we created a cohort of residents of the city of Ottawa, Ontario, who underwent a PCR test for COVID-19 between March 10, 2020, and May 13, 2021. Using demographic, socio-economic and health data (including COVID-19 PCR test results and available, symptom data), we developed predictive models for the purpose of COVID-19 case identification using the following approaches: classical multivariate logistic regression (LR); deep neural network (DNN); random forest (RF); and gradient boosting trees (GBT). Model performance comparisons were made using the area under the curve (AUC) swarm plot for 10-fold cross-validation. ResultsThe cohort consisted of n = 351,248 Ottawa residents tested for COVID-19 during the study period. Among whom, a total of n = 883,879 unique COVID-19 tests were performed (2.6 % positive test results). Inclusion of COVID-19 symptoms data in the analysis improved model performance and variable predictive value across all tested models (p < 0.0001), with the 10-fold cross-validation AUC increasing to near or over 0.7 in all models when symptoms data were included. In various pairwise comparisons, the GBT method had the highest predictive ability (AUC = 0.796 ± 0.017), significantly outperforming multivariate logistic regression and the other AI/ML approaches. ConclusionsConventional multivariate regression-based models are better than some and worse than other machine learning algorithms to provide good predictive accuracy in a moderate dataset with a reasonable number of features. However, whenever possible, the AI/ML GBT approach should be considered.

Digital twin simulation modelling shows that mass testing and local lockdowns effectively controlled COVID-19 in Denmark

BackgroundFollowing the COVID-19 pandemic, it is important to evaluate different mitigation strategies for future preparedness. Mass testing and local lockdowns were employed during the Alpha wave in Denmark, which led to ten times more tests than the typical European member state and incidence-based restrictions at the parish level. This study aims to quantify the effects of these interventions in terms of hospital admissions and societal freedom.MethodsThis study assesses the effectiveness of these strategies via counterfactual scenarios using a detailed, individual-based simulation model that replicates the entire Danish population. The model considers multiple factors, including evolving societal restrictions, vaccination roll-out, seasonal influences, and varying intensities of PCR and antigen testing across different age groups and degree of completed vaccination. It also integrates adaptive human behavior in response to changes in incidences at the municipality and parish levels.ResultsThe simulations show, that without mass testing in Denmark, there would have been a 150% increase in hospital admissions, and additional local lockdowns equivalent to 21 days of strict national lockdown. Without the policy of local lockdowns, hospitalizations would have increased by 50%.ConclusionsIn conclusion, the combination of mass testing and local lockdowns likely prevented a large increase in hospitalizations while increasing overall societal freedom during the Alpha wave in Denmark. In future epidemics, mass testing and local lockdowns can likely prevent overwhelming healthcare systems in phases of high transmission and hospitalization risks.

Nirsevimab Effectiveness Against Severe RSV Infection in the Primary Care Setting.

This study assesses the effectiveness of nirsevimab, a monoclonal antibody, in preventing medically attended respiratory syncytial virus-lower respiratory tract infections (RSV-LRTI) in a large primary care network in Spain, in both overall and catch-up infants under ten months of age. The 2023-24 immunization campaign with nirsevimab in Spain targeted all infants born after April 1st, 2023. Those born after October 1st received it at birth in hospitals, while others received it through a catch-up program. The MEDIPRIM network of primary care centers recruited all infants with LRTI for RSV PCR testing and employed a test-negative design (TND) approach to estimate the effectiveness of nirsevimab. The study included 160 infants, 141 (88%) of them received nirsevimab and 128 belonged to the catch-up group (88% received nirsevimab). Overall, RSV was detected in 44 infants (27.5%). Within the catch-up group, 37 (28.9%) were positive for RSV. The overall effectiveness was 75.8% (95% CI: 40.4-92.7), and 80.2% (95% CI: 44.3-95.4) in infants belonging to the catch-up group. This study underscores the effectiveness of nirsevimab in preventing medically attended LRTI in infants in outpatient settings and emphasizes the importance of a catch-up immunization program to reduce the disease burden in primary care.

The Recombinase Polymerase Amplification Test for Strongyloides stercoralis Is More Sensitive than Microscopy and Real-Time PCR in High-Risk Communities of Cusco, Peru.

Strongyloidiasis is a neglected, soil-transmitted helminth infection prevalent worldwide. The true burden of strongyloidiasis is unclear due to the lack of sensitive, field-friendly diagnostic tests. PCR tests to detect Strongyloides DNA in stool are sensitive and specific, but the need for expensive equipment limits their use in endemic regions. Isothermal PCR amplification tests are easier to deploy while maintaining sensitivity and specificity. We developed and evaluated a recombinase polymerase amplification lateral flow assay (RPA-LFA) to detect Strongyloides stercoralis in human stool samples. Three hundred stool samples were collected in three communities in the jungle of Cusco, Peru. Samples were tested for S. stercoralis larvae using microscopy (Baermann's, agar plate culture (APC), and rapid sedimentation), real-time PCR, and RPA-LF for Strongyloides DNA. The RPA-LFA showed an analytical limit of detection of 20 pg/µL. The prevalence of S. stercoralis was 27%, 38%, 46.3%, and 46% using microscopy, PCR, microscopy/PCR, and RPA-LFA, respectively. RPA-LFA had a sensitivity and specificity of 59.3% and 58.9%, 66.2% and 71.4%, and 77.2% and 73.1% when microscopy, microscopy/PCR, and real-time PCR were used as the gold standards, respectively. The Strongyloides RPA-LFA is a novel, fast, highly sensitive, and specific molecular method with the potential for deployment in endemic regions.

B-195 Accelerating Molecular Diagnostics: Development of a novel Rapid, Integrated PCR System for Acute Respiratory Infections Detection

Abstract Background The COVID-19 pandemic and acute respiratory infections have underscored the necessity for high-throughput and timely molecular diagnostics. Existing molecular diagnostic methods are labor-intensive, a challenge particularly evident in large-scale testing. This study aims to streamline the nucleic acid testing process by integrating extraction and detection in PCR testing, thereby optimizing efficiency. Methods We developed 'EXP-160R', an integrated PCR system capable of rapid thermal cycling. This innovation employs advanced temperature control technology to markedly reduce heating, cooling, and annealing-extension times. The PCR reagent-kit (Zybio Inc.) was optimized to complement the instrument's capabilities, aiming for pathogen nucleic acid detection in acute respiratory infections within 25 minutes. The precision of in-tube temperature during rapid amplification was validated by using temperature-sensitive molecular probes in system development stage. Performance evaluation used the WHO International Standard for SARS-CoV-2 RNA to assess sensitivity and detection limits. We also analyzed 500 clinical samples from multiple Chinese hospitals, including 295 COVID-19 positives and 205 negatives, comparing the EXP-160R system's performance with an NMPA-approved PCR kit. Results The innovative system significantly improved PCR cycle times, enabling detection of 16 samples in 22 minutes and a throughput of 2400 tests per day, without compromising sensitivity or specificity. Validation confirmed a detection limit of 200 copies/mL, and the system achieved a 100% detection rate for medium and high viral load samples, with a coefficient of variation (CV) for Ct values ≤ 5%. Clinical trial comparisons with clinical standard diagnostics revealed a sensitivity of 98.05%, specificity of 100%, accuracy of 99.2%, and a Kappa coefficient of 0.984. When compared to the established PCR kit, our system showed comparable efficacy, with a sensitivity of 98.50%, specificity of 98.67%, accuracy of 98.6%, and a Kappa of 0.971. Conclusions The rapid PCR system represents a significant advancement in molecular diagnostics, offering rapid, high-throughput, and accurate detection of SARS-CoV-2. This system is particularly beneficial in urgent testing scenarios, such as airports and fever clinics. It addresses current pandemic needs and sets a new benchmark for future respiratory and infectious disease diagnostics, enhancing both efficiency and user experience.

B-192 Covid on the PCR Pro: a consumer-friendly PCR test

Abstract Background The emergence of COVID-19 highlighted the need for rapid and accurate diagnostic solutions that can be deployed on a broad scale. The complexity and long turn-around-time of PCR testing typically performed in molecular laboratories is not compatible with community-wide screening. To overcome these challenges, we present the Co-Dx PCR Pro™, a rapid and simple test platform developed for self-collection and at-home use. The instrument is controlled from the user’s smartphone or tablet and performs sample treatment, RT-PCR, and DNA melting analysis. An automatic calling algorithm provides a simple, “Detected,” or, “Not detected,” result. We demonstrate the utility of this platform using a test for the detection of SARS-COV-2. Methods A self-collected anterior nasal swab sample and 2 mLs of water are added to the collection cup. Closing the lid of the cup initiates rehydrating a dried Proteinase K mixture to digest proteins in the sample. After a brief Proteinase K heat-kill step, the instrument mixes the sample with a dried PCR master mix. Primers target two SARS-COV-2 specific genes (RdRp and E gene) and the human RNase P gene as a control. The sample undergoes an RT step followed by 45 cycles of PCR and DNA melting analysis. Detection is by an automatic calling algorithm that identifies a SARS-COV-2 amplicon melting peak at 82°C and RNase P peak at 87°C. The total runtime is less than 30 minutes. The limit of detection for the assay was assessed by contriving intact virus in negative sample matrix, which was then applied to dry swabs. In addition, frozen and banked clinical samples were tested using the Logix Smart®™ Covid-19 test as a comparator assay. A third party performed a usability assessment by having (1) naïve users follow the instructions and self-administer the test (2) expert trained users perform an independent assessment of the assay. Participants were then asked to complete a usability assessment. Results The 95% limit of detection was 2,100 copies/swab using contrived nasal swabs. Clinical samples were blinded to the operator and run on the platform. All samples resulted in a diagnosis concordant with the comparator assay. The usability assessment was performed by six naïve users and resulted in a systems usability scale score of “Best” (84.2 ± 14.3, out of 100). The two expert users determined an 80% limit of detection of 700 genome equivalents and “ease of use,” and “satisfaction,” scores of 4.5/5.0 and 5.0/5.0, respectively. Conclusions Performance of the PCR Pro platform is on par with laboratory-based PCR testing. The benefits of our assay are its speed and the low complexity platform accessible to the lay. This platform could improve access to molecular diagnostic testing, including use at home. Future expansion to include common respiratory pathogens (Flu, RSV, Strep) will increase utility. Additionally, anonymized results can be aggregated and provide a wealth of epidemiological information at the community level to better inform public health initiatives and health outcomes.

B-217 Effective Diagnostic Platform and Method to Differentiate the Infectious From Non-Infectious Viruses With Nucleic Acid Detection Workflow

Abstract Background While qPCR has demonstrated high sensitivity, specificity, and ability to deliver reliable quantitative data in clinical samples, such results do not differentiate between infectious and non-infectious viral particles. In view of the principle of the qPCR method itself, the amplification detects the nucleic acid (DNA/RNA) of the virus, not the virus itself. In many cases, it is still possible to detect DNA/RNA fragments from dead viruses in recovered patients, while they are not infectious anymore. This will lead to prolonged quarantine period or treatment for the patients, which could cause great psychological pressure to the patients and increased healthcare burden. Therefore, an effective detection method that can discriminate viable and inviable pathogens is of great significance. Methods SARS-CoV2 viral RNA, SARS-CoV-2 viruses, or clinical samples with SARS-CoV2 infection were diluted in Delta MTM (Molecular Transport Media) to the respective testing concentrations. The samples were incubated with intercalating dye (PMAxx 100nM) with or without Triton X-100 for 10 min. The mixture was then exposed to the photo-activation device for 5-15 min (@ temperature below 37° C). The samples were then subjected to the standard nucleic acid extraction &amp; purification steps with Qiagen viral RNA extraction kit. The extracted RNA was subjected to amplification with SARS-CoV2 Resolute 2.0 &amp; Fortunate 2.1 assays on BioRAD CFX96. Results The viability PCR (vPCR) efficiency was almost 100% for all SARS-CoV2 RNA input tested suggesting that without viral membrane protection, PMAxx dye effectively bound to the naked RNA after photoactivation process, and the bound RNA was not able to be amplified. The vPCR efficiency was further enhanced with addition of surfactant Triton X-100 suggesting the improved permeability of dye through the viral envelope facilitated the PMAxx binding to the nucleic acid. COVID19 positive clinical samples with viral load ≈ Ct 30 treated with or without PMAxx and photo-activation procedures showed distinctive ΔCt value ranging from 9.27±0.18 to 15.01±0.14. The Ct after dye treatment were all non-numeric (non-detectable) if cut-off Ct sets @ 42.0, indicating the complete inhibition after vPCR pretreatment. Since the clinical samples were undergone heat-inactivation pre-treatment, the result shown here indicating that all samples were non-infectious, which was consistent with the samples tested. Conclusions We developed an effective viability PCR detection method and proprietary vPCR device, which was specifically designed for effective dye photo-activation. In addition, the method takes less than 15 min and could be integrated into the existing standard nucleic acid extraction and amplification workflow. Comparing to the standard PCR test result, this method could provide the information not only on the positivity of the tested pathogen, but also its viability, which could be very essential information for public surveillance and diagnostic decisions for healthcare providers.