Infection prevention in medical education - results of a descriptive cross-sectional study in Germany.

Objective: The aim of the study was to assess the current curricular status of content on infection prevention in hospitals during medical education prior to the development of a serious game on infection prevention in hospitals. In addition, the data collected was to be contrasted with the training for a specialist nurse in hygiene and infection prevention (FKHI). Methodology: In an online survey, persons in charge of medical degree programs and continuing education centers for FKHI, SkillsLabs and professional associations in Germany were asked to answer 28 questions on framework conditions, teaching, examinations, and gamification. Results: Data was collected for 22 medical degree programs and 5 FKHI continuing education centers. Due to the low response rate, the data for the FKHI was only analyzed in summary form. On average, 13.5 teaching units (median) are available in medical studies. Six degree programs have a longitudinal curriculum. In 7 of the 22 degree programs, teaching is based on the National Competency-Based Learning Objectives Catalogue (NKLM). Almost all locations teach this content in lectures (n=18) and/or in internships (n=13). Teaching and examinations are most common in the third year of study (n=12). In addition to practical OSCE examinations (n=5), written (n=12) and computer-based (n=8) examinations are used in particular. Gamification is known as a didactic approach to some extent but is not used for teaching infection prevention. Conclusions: Infection prevention in hospitals is given relatively low priority in medical education. Teaching and examinations are based on traditional knowledge-oriented formats, although practical teaching and practical examinations are established at some locations. In contrast to the FKHI, learning objectives currently appear to be less standardized. Further interprofessional development of teaching would be desirable in the future.

Rapid Isothermal Detection of Pathogenic Clostridioides difficile Using Recombinase Polymerase Amplification.

Nosocomial-associated diarrhea due to Clostridioides difficile infection (CDI) is diagnosed after sample precultivation by the detection of the toxins in enzyme immunoassays or via toxin gene nucleic acid amplification. Rapid and direct diagnosis is important for targeted treatment to prevent severe cases and recurrence. We developed two singleplex and a one-pot duplex fluorescent 15 min isothermal recombinase polymerase amplification (RPA) assays targeting the toxin genes A and B (tcdA and tcdB). Furthermore, we adapted the singleplex RPA to a 3D-printed microreactor device. Analytical sensitivity was determined using a DNA standard and DNA extracts of 20 C. difficile strains with different toxinotypes. Nineteen clostridial and gastrointestinal bacteria strains were used to determine analytical specificity. Adaptation of singleplex assays to duplex assays in a 50 µL volume required optimized primer and probe concentrations. A volume reduction by one-fourth (12.4 µL) was established for the 3D-printed microreactor. Mixing of RPA was confirmed as essential for optimal analytical sensitivity. Detection limits (LOD) ranging from 119 to 1411 DNA molecules detected were similar in the duplex tube format and in the singleplex 3D-printed microreactor format. The duplex RPA allows the simultaneous detection of both toxins important for the timely and reliable diagnosis of CDI. The 3D-printed reaction chamber can be developed into a microfluidic lab-on-a-chip system use at the point of care.

Did the prevalence of depressive symptoms change during the COVID-19 pandemic? A multilevel analysis on longitudinal data from healthcare workers.

BACKGROUND: Healthcare workers (HCW) are at high risk to develop mental health problems during the COVID-19 pandemic because of additional work load, perceived stress, and exposure to patients with COVID-19. Currently, there are few studies on change over time in the prevalence of depressive symptoms during pandemic start among HCW. Thus, the aims of the current study were to examine whether depressive symptoms increased during the pandemic and were associated with perceived stress and own COVID-19 infection and workplace exposure to virus-infected patients. METHODS: The cohort study used longitudinal data from HCW collected monthly (July 2020 till December 2020) during the first year of the pandemic before vaccination became available. The sample of n = 166 was drawn from a German hospital and included medical (e.g. nurses, therapists, and physicians) and administrative staff. Using multilevel models, we analyzed the change in depressive symptoms [assessed with General Depression Scale (GDS), a validated German version of the Center for Epidemiological Studies Depression Scale (CES-D)] and its association with perceived stress across the study period. Laboratory-confirmed own infection was tested as a potential moderator in this context. Subscales of the GDS were used to examine change over time of depressive symptom modalities (e.g. emotional, somatic, and social interactions (ß, 95% confidence interval). RESULTS: Depression scores increased significantly during the study period (ß = .03, 95% CI [0.02, 0.05]). Perceived stress was associated with depressive symptoms (ß = .12, 95% CI [0.10, 0.14]) but did not change over time. Exposure to COVID-19 infection was associated with a higher increase of depressive symptoms (ß = .12, 95% CI [0.10, .14]). Somatic symptoms of depression increased among medical HCW with workplace exposure to COVID-19 (ß = .25, 95% CI [0.13, 0.38]), but not in administrators (ß = .03, 95% CI [-0.04, 0.11]). CONCLUSION: Research is needed to identify factors that promote the reduction of depressive symptoms in medical HCW with exposition to COVID-19 patients. Awareness of infection protection measures should be increased.

Examination of blood samples using deep learning and mobile microscopy.

BACKGROUND: Microscopic examination of human blood samples is an excellent opportunity to assess general health status and diagnose diseases. Conventional blood tests are performed in medical laboratories by specialized professionals and are time and labor intensive. The development of a point-of-care system based on a mobile microscope and powerful algorithms would be beneficial for providing care directly at the patient's bedside. For this purpose human blood samples were visualized using a low-cost mobile microscope, an ocular camera and a smartphone. Training and optimisation of different deep learning methods for instance segmentation are used to detect and count the different blood cells. The accuracy of the results is assessed using quantitative and qualitative evaluation standards. RESULTS: Instance segmentation models such as Mask R-CNN, Mask Scoring R-CNN, D2Det and YOLACT were trained and optimised for the detection and classification of all blood cell types. These networks were not designed to detect very small objects in large numbers, so extensive modifications were necessary. Thus, segmentation of all blood cell types and their classification was feasible with great accuracy: qualitatively evaluated, mean average precision of 0.57 and mean average recall of 0.61 are achieved for all blood cell types. Quantitatively, 93% of ground truth blood cells can be detected. CONCLUSIONS: Mobile blood testing as a point-of-care system can be performed with diagnostic accuracy using deep learning methods. In the future, this application could enable very fast, cheap, location- and knowledge-independent patient care.

COVID-19 among Healthcare Workers: A Prospective Serological-Epidemiological Cohort Study in a Standard Care Hospital in Rural Germany.

Healthcare workers (HCW) play a vital role in the SARS-CoV-2 pandemic control. The aim of this study was to assess the prevalence of SARS-CoV-2 antibodies and the risk of COVID-19 infections in a cohort of HCW from four different risk groups (from intensive care unit to administration) of a hospital of a primary care level in rural Germany. The outcomes were monthly measures of antibody seroprevalence over a period of 6 months. Overall, a seroprevalence of 13.41% was determined, with significantly higher prevalence rates among HCW working in areas with more frequent contact to confirmed or suspected cases (30.30%, p = 0.003). The group specific differences in the risk of infection from COVID-19 were detected, as HCW groups with frequent exposure seemed to have an increased risk (RR = 3.18, p = 0.02; CI95 1.09-9.24). The findings contribute to the epidemiological understanding of the virus spread in an unvaccinated population group, which is highly relevant for the pandemic management.

Does Being Ill Improve Acceptance of Medical Technology?-A Patient Survey with the Technology Usage Inventory.

Acceptance of new medical technology may be influenced by social conditions and an individual's background and particular situation. We studied this acceptance by hypothesizing that current and former COVID-19 patients would be more likely to accept an electrocardiogram (ECG) "patch" (attached to the chest) that allows continuous monitoring of the heart than individuals who did not have the disease and thus the respective experience. Currently infected COVID-19 patients, individuals who had recovered from COVID-19, and a control group were recruited online through Facebook (and Instagram) and through general practitioners (GPs). Demographic information and questions tailored to the problem were collected via an online questionnaire. An online survey was chosen in part because of the pandemic conditions, and Facebook was chosen because of the widespread discussions of health topics on that platform. The results confirmed the central hypothesis that people who had experienced a disease are more willing to accept new medical technologies and showed that curiosity about new technologies and willingness to use them were significantly higher in the two groups currently or previously affected by COVID-19, whereas fears of being "monitored" (in the sense of surveillance) were significantly higher among people who had not experienced the disease and threat. Experiencing a serious disease ("patient experience") promotes acceptance of new medical technologies.

Protection From COVID-19­The Efficacy of Face Masks.

BACKGROUND: Since the beginning of 2020 the SARS-CoV-2 virus has spread to nearly every country in the world. The mainly airborne pathogen has led to large numbers of deaths, principally in elderly and vulnerable segments of the population. Protective vaccines have recently become available, but it is not yet clear whether and when population-wide immunity will be achieved. The existence of evidence for the protective effect of masks covering the mouth and nose is a topic of public debate. METHODS: A selective literature search was carried out in PubMed. Data from the German Robert Koch Institute and the Centers for Disease Control and Prevention were also taken into account. RESULTS: When talking, as many as 20 000 droplets ranging in size from 20 to 500 µM are released every second. According to PCR tests, the amount of virus exhaled is highest immediately before the onset of symptoms. No randomized trials have been conducted on the effect of masks covering the mouth and nose. A metaanalysis of 29 studies on infection with SARS-CoV-2, SARS, or MERS revealed that type N-95 masks (corresponding approximately to FFP-2), surgical masks, or similar multilayer cotton masks can greatly reduce the infection risk for the wearers (RR 0.34 [0.26; 0.45], with moderate heterogeneity [I2 = 48%]). Model experiments and case reports suggest that masks covering the mouth and nose afford considerable protection against transmission of SARS-CoV-2 and other airborne diseases by reducing release of and exposure to potentially infectious droplets; in addition, infections that do occur take a milder course. A limitation of the studies analyzed is that in most cases, this effect cannot be viewed in isolation from the protective impact of other measures (distancing, hygiene precautions). CONCLUSION: It can plausibly be assumed that consistent use of masks covering the mouth and nose can play an important role in containing the spread of SARSCoV- 2.

Mental health and psychosocial support strategies in highly contagious emerging disease outbreaks of substantial public concern: A systematic scoping review.

BACKGROUND: Acute disease outbreaks such as the COVID-19 pandemic cause a high burden of psychological distress in people worldwide. Interventions to enable people to better cope with such distress should be based on the best available evidence. We therefore performed a scoping review to systematically identify and summarize the available literature of interventions that target the distress of people in the face of highly contagious disease outbreaks. METHODS: MEDLINE, Cochrane CENTRAL, Web of Science (January 2000 to May 7, 2020), and reference lists were systematically searched and screened by two independent reviewers. Quantitative and qualitative studies investigating the effects of psychological interventions before, during, and after outbreaks of highly contagious emerging infectious diseases, such as SARS, MERS, Ebola, or COVID-19 were included. Study effects were grouped (e.g. for healthcare professionals, community members, people at risk) and intervention contents at the individual and organizational level summarized. We assessed the level of evidence using a modified scheme from the Oxford Centre for Evidence-based Medicine and the Australian National Health and Medical Research Council. RESULTS: Of 4030 records found, 19 studies were included (two RCTs). Most interventions were delivered during-exposure and face-to-face, focused on healthcare workers and crisis personnel, and combined psychoeducation with training of coping strategies. Based on two high-quality studies, beneficial effects were reported for resilience factors (e.g. positive cognitive appraisal) and professional attitudes of healthcare workers, with mixed findings for mental health (e.g. depression). Across all studies, there was positive qualitative feedback from participants and facilitators. We identified seven ongoing studies mostly using online- and mobile-based deliveries. CONCLUSIONS: There is preliminary evidence for beneficial effects of interventions to enable people to better cope with the distress of highly contagious emerging disease outbreaks. Besides the need for more high-quality studies, the summarized evidence may inform decision makers to plan interventions during the current pandemic and to develop pandemic preparedness plans.

Suitcase Lab for Rapid Detection of SARS-CoV-2 Based on Recombinase Polymerase Amplification Assay.

In March 2020, the SARS-CoV-2 virus outbreak was declared as a world pandemic by the World Health Organization (WHO). The only measures for controlling the outbreak are testing and isolation of infected cases. Molecular real-time polymerase chain reaction (PCR) assays are very sensitive but require highly equipped laboratories and well-trained personnel. In this study, a rapid point-of-need detection method was developed to detect the RNA-dependent RNA polymerase (RdRP), envelope protein (E), and nucleocapsid protein (N) genes of SARS-CoV-2 based on the reverse transcription recombinase polymerase amplification (RT-RPA) assay. RdRP, E, and N RT-RPA assays required approximately 15 min to amplify 2, 15, and 15 RNA molecules of molecular standard/reaction, respectively. RdRP and E RT-RPA assays detected SARS-CoV-1 and 2 genomic RNA, whereas the N RT-RPA assay identified only SARS-CoV-2 RNA. All established assays did not cross-react with nucleic acids of other respiratory pathogens. The RT-RPA assay's clinical sensitivity and specificity in comparison to real-time RT-PCR (n = 36) were 94 and 100% for RdRP; 65 and 77% for E; and 83 and 94% for the N RT-RPA assay. The assays were deployed to the field, where the RdRP RT-RPA assays confirmed to produce the most accurate results in three different laboratories in Africa (n = 89). The RPA assays were run in a mobile suitcase laboratory to facilitate the deployment at point of need. The assays can contribute to speed up the control measures as well as assist in the detection of COVID-19 cases in low-resource settings.

Schnellnachweis von SARS-CoV-2 mit recombinase polymerase amplification.

The COVID-19 pandemic highlights the need for fast and simple assays for nucleic acid detection. As an isothermal alternative to RT-qPCR, we outline the development of a detection scheme for SARS-CoV-2 RNA based on reverse transcription recombinase polymerase amplification (RT-RPA) technology. RPA uses recombination proteins in combination with a DNA polymerase for rapid amplification of target DNA at a constant temperature (39-42 °C) within 10 to 20 minutes and can be monitored in real-time with fluorescent probes.

3D Printed Monolithic Microreactors for Real-Time Detection of Klebsiella pneumoniae and the Resistance Gene blaNDM-1 by Recombinase Polymerase Amplification.

We investigate the compatibility of three 3D printing materials towards real-time recombinase polymerase amplification (rtRPA). Both the general ability of the rtRPA reaction to occur while in contact with the cured 3D printing materials as well as the residual autofluorescence and fluorescence drift in dependence on post curing of the materials is characterized. We 3D printed monolithic rtRPA microreactors and subjected the devices to different post curing protocols. Residual autofluorescence and drift, as well as rtRPA kinetics, were then measured in a custom-made mobile temperature-controlled fluorescence reader (mTFR). Furthermore, we investigated the effects of storage on the devices over a 30-day period. Finally, we present the single- and duplex rtRPA detection of both the organism-specific Klebsiella haemolysin (khe) gene and the New Delhi metallo-ß-lactamase 1 (blaNDM-1) gene from Klebsiella pneumoniae. Results: No combination of 3D printing resin and post curing protocol completely inhibited the rtRPA reaction. The autofluorescence and fluorescence drift measured were found to be highly dependent on printing material and wavelength. Storage had the effect of decreasing the autofluorescence of the investigated materials. Both khe and blaNDM-1 were successfully detected by single- and duplex-rtRPA inside monolithic rtRPA microreactors printed from NextDent Ortho Clear (NXOC). The reaction kinetics were found to be close to those observed for rtRPA performed in a microcentrifuge tube without the need for mixing during amplification. Singleplex assays for both khe and blaNDM-1 achieved a limit of detection of 2.5 × 101 DNA copies while the duplex assay achieved 2.5 × 101 DNA copies for khe and 2.5 × 102 DNA copies for blaNDM-1. Impact: We expand on the state of the art by demonstrating a technology that can manufacture monolithic microfluidic devices that are readily suitable for rtRPA. The devices exhibit very low autofluorescence and fluorescence drift and are compatible with RPA chemistry without the need for any surface pre-treatment such as blocking with, e.g., BSA or PEG.

Rapid Detection of SARS-CoV-2 by Low Volume Real-Time Single Tube Reverse Transcription Recombinase Polymerase Amplification Using an Exo Probe with an Internally Linked Quencher (Exo-IQ).

BACKGROUND: The current outbreak of SARS-CoV-2 has spread to almost every country with more than 5 million confirmed cases and over 300,000 deaths as of May 26, 2020. Rapid first-line testing protocols are needed for outbreak control and surveillance. METHODS: We used computational and manual designs to generate a suitable set of reverse transcription recombinase polymerase amplification (RT-RPA) primer and exonuclease probe, internally quenched (exo-IQ), sequences targeting the SARS-CoV-2 N gene. RT-RPA sensitivity was determined by amplification of in vitro transcribed RNA standards. Assay selectivity was demonstrated with a selectivity panel of 32 nucleic acid samples derived from common respiratory viruses. To validate the assay against full-length SARS-CoV-2 RNA, total viral RNA derived from cell culture supernatant and 19 nasopharyngeal swab samples (8 positive and 11 negative for SARS-CoV-2) were screened. All results were compared to established RT-qPCR assays. RESULTS: The 95% detection probability of the RT-RPA assay was determined to be 7.74 (95% CI: 2.87-27.39) RNA copies per reaction. The assay showed no cross-reactivity to any other screened coronaviruses or respiratory viruses of clinical significance. The developed RT-RPA assay produced 100% diagnostic sensitivity and specificity when compared to RT-qPCR (n = 20). CONCLUSIONS: With a run time of 15 to 20 minutes and first results being available in under 7 minutes for high RNA concentrations, the reported assay constitutes one of the fastest nucleic acid based detection methods for SARS-CoV-2 to date and may provide a simple-to-use alternative to RT-qPCR for first-line screening at the point of need.

A lab-on-a-chip for free-flow electrophoretic preconcentration of viruses and gel electrophoretic DNA extraction.

Nucleic acid amplification techniques such as real-time PCR are essential instruments for the identification and quantification of viruses. They are fast, very sensitive and highly specific, but often require elaborate and labor intensive sample preparation to achieve successful amplification of the target sequence. In this work we demonstrate the complete microfluidic preparation of amplifiable virus DNA from dilute specimens. Our approach combines free-flow electrophoretic preconcentration of viral particles with thermal lysis and gel-electrophoretic nucleic acid extraction on a single device. The on-chip preconcentration achieves a capture efficiency of >99% for dilute suspensions of bacteriophage PhiX174. Following preconcentration, phages are thermally lysed and released DNA is recovered after 40 s of on-chip gel-electrophoresis with a recovery rate of ∼73%. Furthermore we demonstrate a detection limit of ∼1 PFU ml-1 (∼0.02 DNA copies per µl) for the detection of bacteriophage PhiX174 by PCR. To simplify operation of the device, we describe the development of a custom-made chip holder as well as a compact peristaltic pump and power supply, which enable user-friendly operation with low risk of cross-contamination and high potential for automation in the field of point-of-care diagnostics.

A lab-on-a-chip for rapid miRNA extraction.

We present a simple to operate microfluidic chip system that allows for the extraction of miRNAs from cells with minimal hands-on time. The chip integrates thermoelectric lysis (TEL) of cells with native gel-electrophoretic elution (GEE) of released nucleic acids and uses non-toxic reagents while requiring a sample volume of only 5 µl. These properties as well as the fast process duration of 180 seconds make the system an ideal candidate to be part of fully integrated point-of-care applications for e.g. the diagnosis of cancerous tissue. GEE was characterized in comparison to state-of-the-art silica column (SC) based RNA recovery using the mirVana kit (Ambion) as a reference. A synthetic miRNA (miR16) as well as a synthetic snoRNA (SNORD48) were subjected to both GEE and SC. Subsequent detection by stem-loop RT-qPCR demonstrated a higher yield for miRNA recovery by GEE. SnoRNA recovery performance was found to be equal for GEE and SC, indicating yield dependence on RNA length. Coupled operation of the chip (TEL + GEE) was characterized using serial dilutions of 5 to 500 MCF7 cancer cells in suspension. Samples were split and cells were subjected to either on-chip extraction or SC. Eluted miRNAs were then detected by stem-loop RT-qPCR without any further pre-processing. The extraction yield from cells was found to be up to ~200-fold higher for the chip system under non-denaturing conditions. The ratio of eluted miRNAs is shown to be dependent on the degree of complexation with miRNA associated proteins by comparing miRNAs purified by GEE from heat-shock and proteinase-K based lysis.

Recombinase polymerase amplification assay for rapid detection of Monkeypox virus.

In this study, a rapid method for the detection of Central and West Africa clades of Monkeypox virus (MPXV) using recombinase polymerase amplification (RPA) assay targeting the G2R gene was developed. MPXV, an Orthopoxvirus, is a zoonotic dsDNA virus, which is listed as a biothreat agent. RPA was operated at a single constant temperature of 42°C and produced results within 3 to 10 minutes. The MPXV-RPA-assay was highly sensitive with a limit of detection of 16 DNA molecules/µl. The clinical performance of the MPXV-RPA-assay was tested using 47 sera and whole blood samples from humans collected during the recent MPXV outbreak in Nigeria as well as 48 plasma samples from monkeys some of which were experimentally infected with MPXV. The specificity of the MPXV-RPA-assay was 100% (50/50), while the sensitivity was 95% (43/45). This new MPXV-RPA-assay is fast and can be easily utilised at low resource settings using a solar powered mobile suitcase laboratory.

Diagnosing Zika virus infection against a background of other flaviviruses: Studies in high resolution serological analysis.

Zika virus (ZIKV) is a mosquito-borne flavivirus. Homologous proteins of different flaviviruses display high degrees of sequence identity, especially within subgroups. This leads to extensive immunological cross-reactivity and corresponding problems for developing a ZIKV-specific serological assay. In this study, peptide microarrays were employed to identify individual ZIKV antibody targets with promise in differential diagnosis. A total of 1643 overlapping oligopeptides were synthesized and printed onto glass slides. Together, they encompass the full amino acid sequences of ZIKV proteomes of African, Brazilian, USA, and French Polynesian origins. The resulting ZIKV scanning microarray chips were used to screen three pools of sera from recent Zika outbreaks in Senegal and Cape Verde, in Brazil, and from overseas travelers returning to the EU. Together with a mixed pool of well characterized, archived sera of patients suffering from infections by dengue, yellow fever, tick-borne encephalitis, and West Nile viruses, a total of 42 sera went into the study. Sixty-eight antibody target regions were identified. Most of which were hitherto unknown. Alignments and sequence comparisons revealed 13 of which could be classified as bona fide ZIKV-specific. These identified antibody target regions constitute a founding set of analytical tools for serological discrimination of ZIKV from other flaviviruses.

Development of Mobile Laboratory for Viral Hemorrhagic Fever Detection in Africa.

Background: A mobile laboratory transportable on commercial flights was developed to enable local response to viral hemorrhagic fever outbreaks. Methods: The development progressed from use of mobile real-time reverse-transcription polymerase chain reaction to mobile real-time recombinase polymerase amplification. In this study, we describe various stages of the mobile laboratory development. Results: A brief overview of mobile laboratory deployments, which culminated in the first on-site detection of Ebola virus disease (EVD) in March 2014, and their successful use in a campaign to roll back EVD cases in Conakry in the West Africa Ebola virus outbreak are described. Conclusions: The developed mobile laboratory successfully enabled local teams to perform rapid disgnostic testing for viral hemorrhagic fever.

Direct DNA and RNA detection from large volumes of whole human blood.

PCR inhibitors in clinical specimens negatively affect the sensitivity of diagnostic PCR and RT-PCR or may even cause false-negative results. To overcome PCR inhibition, increase the sensitivity of the assays and simplify the detection protocols, simple methods based on quantitative nested real-time PCR and RT-PCR were developed to detect exogenous DNA and RNA directly from large volumes of whole human blood (WHB). Thermus thermophilus (Tth) polymerase is resistant to several common PCR inhibitors and exhibits reverse transcriptase activity in the presence of manganese ions. In combination with optimized concentrations of magnesium ions and manganese ions, Tth polymerase enabled efficient detection of DNA and RNA from large volumes of WHB treated with various anticoagulants. The applicability of these methods was further demonstrated by examining WHB specimens collected from different healthy individuals and those stored under a variety of conditions. The detection limit of these methods was determined by detecting exogenous DNA, RNA, and bacteria spiked in WHB. To the best of our knowledge, direct RNA detection from large volumes of WHB has not been reported. The results of the developed methods can be obtained within 4 hours, making them possible for rapid and accurate detection of disease-causing agents from WHB.

Capacity of rTth polymerase to detect RNA in the presence of various inhibitors.

The full potential of the real-time reverse transcription polymerase chain reaction (RT-PCR) as a rapid and accurate diagnostic method is limited by DNA polymerase inhibitors as well as reverse transcriptase inhibitors which are ubiquitous in clinical samples. rTth polymerase has proven to be more resistant to DNA polymerase inhibitors present in clinical samples for DNA detection and also exhibits reverse transcriptase activity in the presence of Mn2+ ions. However, the capacity of rTth polymerase, which acts as DNA polymerase and reverse transcriptase, to detect RNA in the presence of various inhibitors has not been investigated in detail. Herein, the inhibitors originating from various clinical samples such as blood, urine, feces, bodily fluids, tissues and reagents used during nucleic acid extraction were employed to evaluate the capacity of rTth polymerase to detect RNA. The results show that the inhibitors have different inhibitory effects on the real-time RT-PCR reactions by rTth polymerase, and the inhibitory effects are concentration dependent. Additionally, the capacity of rTth polymerase to detect RNA in the presence of various inhibitors is better or at least comparable with its capacity to detect DNA in the presence of various inhibitors. As a consequence, RNA may be directly detected in the presence of co-purified inhibitors or even directly from crude clinical samples by rTth polymerase.