Rapid Polymerase Chain Reaction Test Research Articles

Impact of a Rapid Point of Care Test for Influenza on Guideline Consistent Care and Antibiotic Use.

Rapid influenza diagnostic tests that detect the presence of viral antigens are currently used throughout the United States but have poor sensitivity. The objective of this study was to identify if the use of a new highly accurate rapid point of care test would significantly increase the likelihood of guideline consistent care. We prospectively recruited 300 students at a university health clinic who presented with cough and 1 influenza-like illness symptom between December 2016 and February 2017 to receive care guided by a rapid polymerase chain reaction (PCR) test. Of the 300 patients receiving the PCR test, 264 had complete medical records and were compared to 771 who received usual care. We used a logistic regression model to identify whether PCR guided care was associated with guideline consistent care, based on the appropriate use of oseltamivir and antibiotics. We also assessed whether PCR guided care decreased the likelihood of return visits within 2 weeks by patients. Logistic regression revealed that the odds of receiving guideline supported care did not significantly increase for patients who received PCR guided care (adjusted odds ratio [aOR], 1.24; 95% CI, 0.83-1.88). It significantly decreased the likelihood of an antibiotic prescription (aOR, 0.61; 95% CI, 0.40-0.94), increased the likelihood of receiving oseltamivir (aOR, 1.57; 95% CI, 1.09-2.28), and decreased the likelihood of return visit within 2 weeks (aOR, 0.19; 95% CI, 0.04-0.81). The use of a rapid PCR test did not significantly improve the likelihood of guideline consistent care. However, independent of test outcome, patients who received the test were more likely to receive an antiviral and less likely to receive an antibiotic or have a return visit within 2 weeks.

The new frontier of diagnostics: Molecular assays and their role in infection prevention and control.

Recent advances in technology over the last decade have propelled the microbiology laboratory into a pivotal role in infection prevention and control. The rapid adaptation of molecular technologies to the field of clinical microbiology now greatly influences infectious disease management and significantly impacts infection control practices. This review discusses recent developments in molecular techniques in the diagnosis of infectious diseases. It describes the basic concepts of molecular assays, discusses their advantages and limitations, and characterizes currently available commercial assays with respect to cost, interpretive requirements, and clinical utility.

Impact of Clinical Pathway and Rapid Direct Influenza Polymerase Chain Reaction Test Introduction on Readmissions Among Non-hospitalized Children with Influenza-Like Illness

BackgroundDiagnosing influenza is challenging in children as influenza-like illness (ILI) symptoms are nonspecific. A clinical pathway was introduced in the emergency department (ED) of a large pediatric hospital to screen and treat patients at high risk of influenza-related complications. A new highly accurate, rapid polymerase chain reaction (PCR) influenza test was introduced the next year to provide more timely results. We aimed to measure the impact of the pathway and new PCR test on rates of all-cause ED readmissions and subsequent hospital admissions in all children with ILI discharged from the ED.MethodsWe conducted a retrospective cohort study of non-hospitalized children≤18 years presenting to the ED with ILI over 3 winter seasons. ILI cases were identified using syndromic surveillance definitions. Outcomes included any return ED encounter or hospital admission within 2 weeks of initial ED visit. We compared difference in outcomes between the years before pathway introduction, after pathway introduction, and after rapid PCR test introduction. Multivariate logistic regression adjusted for pertinent sociodemographic and clinical covariates.ResultsAmong 10799 children with ILI, 6.1% had an ED readmission and 2.5% had a hospital admission within 2 weeks of an initial ED visit. Overall rates of ED readmission or hospitalization did not differ significantly by study year (5.7% vs 6.5% vs 6.1%, P = 0.41, and 2.9% vs 2.4% vs 2.3%, P = 0.36). In multivariate analysis, pathway introduction alone was not associated with likelihood of ED readmission or hospitalizations. However, rapid PCR test introduction with the pathway was associated with lower odds of hospitalization (aOR 0.70, 95% CI 0.51–0.97). High-risk status was associated with higher odds of ED readmission (aOR 1.42, 95% CI 1.21–1.68). High-risk status and severity of illness at the initial ED visit were associated with higher odds of hospitalization (aOR 1.51, 95% CI 1.13–2.03; aOR 10.66, 95% CI 6.05–18.79).ConclusionClinical pathway and rapid PCR test introduction does not decrease all-cause ED readmissions but does decrease all-cause hospital admissions in children with ILI. Clinical factors such as severity of illness and high-risk status play a large role in determining outcomes.Disclosures All authors: No reported disclosures.

Geolocalization of Influenza Outbreak Within an Acute Care Population: A Layered-Surveillance Approach

We seek to use a novel layered-surveillance approach to localize influenza clusters within an acute care population. The first layer of this system is a syndromic surveillance screen to guide rapid polymerase chain reaction testing. The second layer is geolocalization and cluster analysis of these patients. We posit that any identified clusters could represent at-risk populations who could serve as high-yield targets for preventive medical interventions. This was a prospective observational surveillance study. Patients were screened with a previously derived clinical decision guideline that has a 90% sensitivity and 30% specificity for influenza. Patients received points for the following signs and symptoms within the past 7 days: cough (2 points), headache (1 point), subjective fever (1 point), and documented fever at triage (temperature >38°C [100.4°F]) (1 point). Patients scoring 3 points or higher were indicated for influenza testing. Patients were tested with Xpert Flu (Cepheid, Sunnyvale, CA), a rapid polymerase chain reaction test. Positive results were mapped with ArcGIS (ESRI, Redlands, CA) and analyzed with kernel density estimation to create heat maps. There were 1,360 patients tested with Xpert Flu with retrievable addresses within the greater Phoenix metro area. One hundred sixty-seven (12%) of them tested positive for influenza A and 23 (2%) tested positive for influenza B. The influenza A virus exhibited a clear cluster pattern within this patient population. The densest cluster was located in an approximately 1-square-mile region southeast of our hospital. Our layered-surveillance approach was effective in localizing a cluster of influenza A outbreak. This region may house a high-yield target population for public health intervention. Further collaborative efforts will be made between our hospital and the Maricopa County Department of Public Health to perform a series of community vaccination events before the next influenza season. We hope these efforts will ultimately serve to reduce the burden of this disease on our patient population, and that this system will serve as a framework for future investigations locating at-risk populations.

A Randomized Controlled Trial Comparing the Treatment of Patients Tested for Chlamydia and Gonorrhea After a Rapid Polymerase Chain Reaction Test Versus Standard of Care Testing.

We tested the effect of a rapid molecular test for Chlamydia trachomatis (CT)/Neisseria gonorrhoeae (NG) diagnosis on clinical emergency department decision making compared with standard care. The new test presents an opportunity to improve antibiotic management and patient outcomes. We conducted a randomized controlled trial of 70 consenting patients 18 years or older presenting to an urban emergency department with sexually transmitted infections complaints (vaginal/penile discharge, dysuria, vaginal/penile itching/pain, dyspareunia). Participants were randomized to rapid testing or standard care if a sexually transmitted infection was suspected. Follow-up phone calls were performed 7 to 10 days postdischarge. The primary outcomes included: antibiotic overtreatment rates, partner notification, and health care utilization. A total of 12.9% tested positive for CT or NG and received antibiotics. Test patients with negative results were less likely to receive empirical antibiotic treatment than control patients, absolute risk difference [RD], 33.4 (95% confidence interval [CI], 7.9%-58.9%), risk ratio [RR], 0.39 (95% CI, 0.19-0.82). Thirty-seven participants (53%) were contacted for follow-up 7 to 10 days postdischarge. Test patients were less likely to report missed antibiotic doses (RD, -51.3%; 95% CI, -84.4% to -18.2%; RR, 0.23; 95% CI, 0.06-0.88). Test patients were more likely to be notified of their results (RD, 50.6%; 95% CI, 22.7%-78.5%; RR, 2.72; 95% CI, 1.26-5.86). There were no significant differences in charges or health care utilization measures. We found a significant reduction in unnecessary antibiotic treatment for CT/NG in subjects receiving the rapid molecular test compared with those receiving nucleic acid amplification test.

Diagnosis, treatment, and outcomes of coccidioidomycosis in allogeneic stem cell transplantation.

Allogeneic hematopoietic stem cell transplant (allo-HSCT) recipients have multiple risk factors for coccidioidomycosis, and previous reports of coccidioidomycosis in this patient population describe severe infections with poor outcomes. We performed a retrospective chart review of allo-HSCT recipients with active coccidioidomycosis to characterize the utility of diagnostic tests for coccidioidomycosis and to determine treatment outcomes. Eleven of 426 (2.6%) allo-HSCT recipients experienced active coccidioidomycosis after transplantation. Of these 11 patients, 1 (9%) had extrapulmonary infection, 9 (82%) patients were hospitalized, and 5 (45%) died. Culture or histology was positive in 33% (3/9) of the patients tested. Most (64% [7/11]) had at least 1 positive serologic test result, and the enzyme immunoassay immunoglobulin G test was positive most often (overall 55% [6/11]). Chest radiographs and chest computed tomography scans showed miliary or multifocal nodular infiltrates or consolidations, consistent with coccidioidomycosis, in 80% (8/10) and 100% (9/9), respectively, of patients tested throughout the course of active illness. Rapid polymerase chain reaction testing was positive in 71% (5/7) of the patients tested. Peripheral eosinophilia was present in 18% (2/11) of patients. Coccidioidomycosis is associated with high morbidity and mortality in allo-HSCT recipients in an area endemic for Coccidioides. Diagnosis of this infection can be difficult and often requires multiple and frequently invasive tests. Antifungal prophylaxis should be considered for patients at highest risk.

A selected screening programme was less effective in the detection of methicillin-resistant Staphylococcus aureus colonisation in an orthopaedic unit

Our unit has used a selective screening policy for methicillin-resistant Staphylococcus aureus (MRSA) colonisation using standard chromogenic growth media, based upon risk stratification. The aim of this study was to examine the effectiveness of this selective screening policy. A cohort of 429 patients was assessed for their risk status for MRSA colonisation using both rapid polymerase chain reaction (PCR) swabs and traditional culture and sensitivity analysis. The sensitivity, specificity, positive predictive values and negative predictive values of the traditional selective approach were calculated compared to universal rapid screening. One hundred eighteen patients were considered high risk and would traditionally be further screened with standard culture of swabs. The prevalence of MRSA was 15/429 (3.5%). The sensitivity of selective screening was 53% identifying eight of 15 cases. The false-negative rate was therefore 47% and seven would have been missed. PCR results were available within four to six hours, whereas culture results were only available at 24 hours for the media showing no growth and not until 72 hours for positive MRSA cases. We now advocate universal screening prior to, or on admission, using this rapid PCR test, as we consider this identifies MRSA colonisation more effectively and facilitates "ring-fencing" of orthopaedic beds.

Impact of Rapid Methicillin-Resistant Staphylococcus aureus Polymerase Chain Reaction Testing on Mortality and Cost Effectiveness in Hospitalized Patients with Bacteraemia

Patients hospitalized with Staphylococcus aureus bacteraemia have an unacceptably high mortality rate. Literature available to date has shown that timely selection of the most appropriate antibacterial may reduce mortality. One tool that may help with this selection is a polymerase chain reaction (PCR) assay that distinguishes methicillin (meticillin)-resistant S. aureus (MRSA) from methicillin-susceptible S. aureus (MSSA) in less than 1 hour. To date, no information is available evaluating the impact of this PCR technique on clinical or economic outcomes. To evaluate the effect of a rapid PCR assay on mortality and economics compared with traditional empiric therapy, using a literature-derived model. A literature search for peer-reviewed European (EU) and US publications regarding treatment regimens, outcomes and costs was conducted. Information detailing the rates of infection, as well as the specificity and sensitivity of a rapid PCR assay (Xpert MRSA/SA Blood Culture PCR) were obtained from the peer-reviewed literature. Sensitivity analysis varied the prevalence rate of MRSA from 5% to 80%, while threshold analysis was applied to the cost of the PCR test. Hospital and testing resource consumption were valued with direct medical costs, adjusted to year 2009 values. Adjusted life-years were determined using US and WHO life tables. The cost-effectiveness ratio was defined as the cost per life-year saved. Incremental cost-effectiveness ratios (ICERs) were calculated to determine the additional cost necessary to produce additional effectiveness. All analyses were performed using TreeAge Software (2008). The mean mortality rates were 23% for patients receiving empiric vancomycin subsequently switched to semi-synthetic penicillin (SSP) for MSSA, 36% for patients receiving empiric vancomycin treatment for MRSA, 59% for patients receiving empiric SSP subsequently switched to vancomycin for MRSA and 12% for patients receiving empiric SSP for MSSA. Furthermore, with an MRSA prevalence of 30%, the numbers of patients needed to test in order to save one life were 14 and 16 compared with empiric vancomycin and SSP, respectively. The absolute mortality difference for MRSA prevalence rates of 80% and 5% favoured the PCR testing group at 2% and 10%, respectively, compared with empiric vancomycin and 18% and 1%, respectively, compared with empiric SSP. In the EU, the cost-effectiveness ratios for empiric vancomycin- and SSP-treated patients were Euro 695 and Euro 687 per life-year saved, respectively, compared with Euro 636 per life-year saved for rapid PCR testing. In the US, the cost-effectiveness ratio was $US 898 per life-year saved for empiric vancomycin and $US 820 per life-year saved for rapid PCR testing. ICERs demonstrated dominance of the PCR test in all instances. Threshold analysis revealed that PCR testing would be less costly overall, even at greatly inflated assay prices. Rapid PCR testing for MRSA appears to have the potential to reduce mortality rates while being less costly than empiric therapy in the EU and US, across a wide range of MRSA prevalence rates and PCR test costs.

An Evaluation of a Rapid Real Time Polymerase Chain Reaction Assay for Detection of Group B Streptococcus as Part of a Neonatal Group B Streptococcus Prevention Strategy

ObjectiveTo evaluate the sensitivity, specificity, and feasibility of a rapid real-time polymerase chain reaction (PCR) test for group B streptococcus (GBS) completed during labour, compared with the standard culture test performed at 35 to 37 weeks’ gestation. MethodsWomen presenting to the maternity unit for term vaginal delivery had two vaginal/rectal samples collected. One swab was tested using a rapid PCR method (IDI-Strep B, Infectio Diagnostic [IDI] Inc., Sainte-Foy QC), and the other was cultured after enrichment (intrapartum culture). Comparisons were made between these results and those of a culture-based screen at 35 to 37 weeks’ gestation. ResultsOf the 190 women enrolled, 85% had results of the standard screen at 35 to 37 weeks available for comparison. The sensitivity and specificity of the standard 35- to 37-week screen were 84.3% (95% confidence interval [CI], 71.4–93.0) and 93.2% (95% CI 86.5–97.2) respectively, whereas the sensitivity and specificity of the rapid PCR were 90.7% (95% CI 79.7–96.9) and 97.6% (95% CI 93.1–99.5), respectively. The median reporting time for the rapid PCR test was 99 minutes (range 50–255). Results were available more than four hours before delivery in 81% of cases. ConclusionIn this Canadian centre, a rapid PCR test done at the time of labour (IDI-Strep B) demonstrated high sensitivity and specificity, comparable to the 35- to 37-week screen. The time to reporting results was acceptably short, allowing for timely administration of intrapartum prophylactic antibiotics.

Acceptance of a Rapid Herpes Test in Labour: Survey of Attitudes of Patients and Health Care Providers

ObjectiveTo determine the acceptability to pregnant women and their health care providers of a rapid test for genital herpes simplex virus (HSV) in labour. MethodsA cross-sectional survey was conducted with outpatient pregnant women and their health care providers (obstetricians, family physicians and midwives) at BC Women’s Hospital and Health Centre. ResultsOf pregnant women approached, 207 (92%) completed the survey; 90% reported no history of genital herpes. Rapid HSV testing in labour was acceptable to 85% of pregnant women. Among the 133 women who were planning a vaginal delivery, 63% were willing to consider delivery by Caesarean section and 53% were willing to consider intrapartum anti-viral medications if HSV was present in the genital tract. Of 51 health care providers surveyed, 98% indicated interest in knowing if their patient had a newly acquired HSV infection, while 84% indicated interest in knowing if the patient had a reactivation of infection. If HSV was detected in their patient’s genital tract, 36% indicated they would recommend a Caesarean section, and 25% would consider antiviral medication as an investigational intrapartum treatment. Interestingly, both of these proportions increased if the patient had ruptured membranes for more than four hours. ConclusionMost pregnant women and their health care providers are receptive to the use of a rapid polymerase chain reaction test to detect genital HSV shedding in labour. This supports the development of HSV rapid testing and antiviral therapy trials in the labour setting.

Amplification of ST50 gene using dry-reagent–based polymerase chain reaction for the detection of Salmonella typhi

Conventional polymerase chain reaction (PCR) testing requires many pipetting steps and has to be transported and stored in cold chain. To overcome these limitations, we designed a ready-to-use PCR test for Salmonella typhi using PCR reagents, primers against the ST50 gene of S. typhi, a built-in internal amplification control (IAC), and gel loading dye mixed and freeze-dried in a single tube. The 2-step dry-reagent–based assay was used to amplify a 1238-bp target gene and an 810-bp IAC gene from 73 BACTEC blood culture broths (33 true positives for S. typhi and 40 true negatives for non- S. typhi). The sensitivity, specificity, positive predictive value, and negative predictive value of the PCR assay were 87.9%, 100%, 100%, and 90.9%, respectively. We suggest that this rapid 2-step PCR test could be used for the rapid diagnosis of typhoid fever.

Effect on MRSA transmission of rapid PCR testing of patients admitted to critical care

We report a significant reduction in the rate of meticillin-resistant Staphylococcus aureus (MRSA) transmission on a critical care unit when admission screening by culture was replaced with a same-day polymerase chain reaction (PCR) test. This was an observational cohort study, set in a 19-bed mixed medical and surgical adult critical care unit in southwest England. We studied 1305 patients admitted between April 2005 and February 2006. Standard MRSA culture methods were used to screen 612 patients between April 2005 and August 2005, and the IDI MRSA PCR test was used to screen 693 patients between September 2005 and February 2006. Standard infection control precautions were instituted when positive results were obtained by either method. Outcome measures included carriage rate, turnaround time for results and rate of subsequent MRSA transmission on the unit. The overall carriage rate on admission to the unit was 7.0%. Culture results were available in three working days, PCR results within one working day. The mean incidence of MRSA transmission was 13.89/1000 patient days during the culture phase and 4.9/1000 patient days during the PCR phase (relative risk reduction 0.65, 95% CI 0.28-1.07). PCR screening for MRSA on admission to critical care units is feasible in routine clinical practice, provides quicker results than culture-based screening and is associated with a significant reduction in subsequent MRSA transmission.

Recurrent inversion breaking intron 1 of the factor VIII gene is a frequent cause of severe hemophilia A

The messenger RNA (mRNA) from 5 of 69 patients with severe hemophilia A did not support amplification of complementary DNA containing the first few exons of the factor VIII (F8) gene but supported amplification of mRNA containing exon 1 of F8 plus exons of the VBP1 gene. This chimeric mRNA signals an inversion breaking intron 1 of the F8 gene. Using an inversion patient, one deleted for F8 exons 1 to 6, and cosmids mapped 70 to 100 kb telomeric of the F8 gene, this study shows that this break strictly affects a sequence (int1h-1) repeated (int1h-2) about 140 kb more telomerically, between the C6.1A and VBP1 genes. The 1041-base pair repeats differ at a single nucleotide (although int1h-2 also showed one polymorphism) and are in opposite orientation. The results demonstrate that they cause inversions by intrachromosome or intrachromatid homologous recombination. The genomic structure of the inversion region shows that transcription traverses intergenic spaces to produce the 2 chimeric mRNAs containing the F8 sequences and characteristic of the inversion. This observation prompts the suggestion that nature may use such extended transcription to test whether the addition of novel domains from neighboring genes creates desirable new genes. A rapid polymerase chain reaction test was developed for the inversion in both patients and carriers. This has identified 10 inversions, affecting F8 genes with 5 different haplotypes for the BclI, introns 13 and 22 VNTR polymorphism, among 209 unrelated families with severe hemophilia A. This indicates a prevalence of 4.8% and frequent recurrence of the inversion. This should result in absence of F8, and one inversion patient is known to have inhibitors. (Blood. 2002;99:168-174)

Rapid polymerase chain reaction-based detection of the causative agent of cat scratch disease ( Bartonella henselae) in formalin-fixed, paraffin-embedded samples

Bartonella (formerly Rochalimaea) henselae ( Bh) plays a central role in cat scratch disease. A polymerase chain reaction (PCR)-based assay that can detect Bh DNA in formalin-fixed, paraffin-embedded (FF-PE) samples would have utility in the evaluation of processed lymph nodes suggestive of this disorder. Fresh or FF-PE cultures of Bh and related species were analyzed. Thirteen lymph nodes (12 FF-PE and one fresh cell suspension) with necrotizing suppurative granulomatous inflammation and seven FF-PE negative control lymph nodes were also evaluated. PCR was performed using a novel, heminested protocol. Amplified products were analyzed by gel electrophoresis. The fresh and FF-PE Bh cultures showed a specific PCR product with an analytical sensitivity of 0.5 pg bacterial DNA. Seven (54%) of 13 clinical lymph node samples with morphological features suggestive of cat scratch disease also had detectable Bh DNA, whereas none of the seven negative control lymph nodes yielded positive results. We have designed a rapid and sensitive PCR test that can reliably detect Bh DNA in fresh and FF-PE samples. Our findings indicate that this assay has clinical utility in the diagnosis of cat scratch disease.

Development of a polymerase chain reaction-probe test for identification of Alloiococcus otitis.

A rapid polymerase chain reaction test was developed for specific identification of the human middle ear pathogen Alloiococcus otitis. Primers for the enzymatic amplification reaction were designed from highly specific sequences within the 16S rRNA gene. In addition, a confirmatory test based on hybridization of the polymerase chain reaction products to a specific internal probe was developed.