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Evaluation of the Modified Two-Tiered Testing Method for Diagnosis of Lyme Disease in Children.

ABSTRACT: Conventional two-tiered testing (CTTT) for Lyme disease includes a first-tier enzyme immunoassay (EIA) followed by a supplemental immunoblot, and modified two-tiered testing (MTTT) relies on two different sequential EIAs without the inclusion of an immunoblot. MTTT has shown promising results as an alternative strategy for the diagnosis of Lyme disease in adults but has not yet been evaluated in children. We performed a cross-sectional study of children and adolescents ≤21 years of age undergoing clinical investigation for suspected Lyme disease. Serum specimens were analyzed with both a whole-cell sonicate (WCS) and a C6 EIA, with a supplemental immunoblot if either EIA was positive or equivocal. We compared CTTT (WCS EIA followed by supplemental immunoblot) to MTTT (WCS EIA followed by C6 EIA) using McNemar's test to evaluate for agreement beyond chance alone. We then used a kappa statistic to measure level of the agreement between testing strategies. We included 1,066 serum specimens, of which 156 (14.6%) had a positive CTTT and 165 (15.5%) had a positive MTTT. There were no significant differences between MTTT and CTTT (P = 0.16). Although the overall agreement between MTTT and CTTT was high (kappa, 0.88; 95% confidence interval, 0.84 to 0.92), 33 children had discordant test results. In a cohort of children and adolescents undergoing investigation for suspected Lyme disease, CTTT and MTTT results agreed in most cases. Since immunoblots are time-consuming, laborious, and challenging to interpret, MTTT provides a promising alternate Lyme disease testing strategy for children.

SUBMITTER: Lipsett SC

PROVIDER: S-EPMC6760964 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Evaluation of Modified Two-Tiered Testing Algorithms for Lyme Disease Laboratory Diagnosis Using Well-Characterized Serum Samples.

Project description:Standard two-tiered testing (STTT) is the recommended algorithm for laboratory diagnosis of Lyme disease (LD). Several limitations are associated with STTT that include low sensitivity in the early stages of disease, as well as technical complexity and subjectivity associated with second-tier immunoblotting; therefore, modified two-tiered testing (MTTT) algorithms that utilize two sequential first-tier tests and eliminate immunoblotting have been evaluated. Recently, a novel MTTT that uses a VlsE chemiluminescence immunoassay followed by a C6 enzyme immunoassay has been proposed. The purpose of this study was to evaluate the performance of the VlsE/C6 MTTT using well-characterized serum samples. Serum samples from the CDC Lyme Serum Repository were tested using three MTTTs, VlsE/C6, whole-cell sonicate (WCS)/C6, and WCS/VlsE, and three STTTs (immunoblotting preceded by three different first-tier assays: VlsE, C6, and WCS). Significant differences were not observed between the results of the MTTTs assessed; however, the VlsE/C6 MTTT resulted in the highest specificity (100%) when other diseases were tested and the lowest sensitivity (75%) for LD samples. Significant differences were present between the results for various MTTTs and STTTs evaluated. Specifically, all MTTTs resulted in higher sensitivities than the STTTs for all LD groups combined and were significantly more accurate (i.e., higher proportion of correct classifications) for this group, with the exception of the WCS/ViraStripe STTT. Additionally, when other diseases were tested, only the results of the VlsE/C6 MTTT differed significantly from those of the WCS/ViraStripe STTT, with the VlsE/C6 MTTT resulting in a 6.2% higher accuracy. Overall, the VlsE/C6 MTTT offers an additional laboratory testing algorithm for LD with equivalent or enhanced performance compared to that of the other MTTTs and STTTs evaluated in this study.

| S-EPMC6062810 | biostudies-other

Performance of a Modified Two-Tiered Testing Enzyme Immunoassay Algorithm for Serologic Diagnosis of Lyme Disease in Nova Scotia.

Project description:Compared to the standard two-tiered testing (STTT) algorithm for Lyme disease serology using an enzyme immunoassay (EIA) followed by Western blotting, data from the United States suggest that a modified two-tiered testing (MTTT) algorithm employing two EIAs has improved sensitivity to detect early localized Borrelia burgdorferi infections without compromising specificity. From 2011 to 2014, in the Canadian province of Nova Scotia, where Lyme disease is hyperendemic, sera submitted for Lyme disease testing were subjected to a whole-cell EIA, followed by C6 EIA and subsequently IgM and/or IgG immunoblots on sera with EIA-positive or equivocal results. Here, we evaluate the effectiveness of the MTTT algorithm compared to the STTT approach in a Nova Scotian population. Retrospective chart reviews were performed on patients testing positive with the whole-cell and C6 EIAs (i.e., the MTTT algorithm). Patients were classified as having Lyme disease if they had a positive STTT result, a negative STTT result but symptoms consistent with Lyme disease, or evidence of seroconversion on paired specimens. Of the 10,253 specimens tested for Lyme disease serology, 9,806 (95.6%) were negative. Of 447 patients who tested positive, 271 charts were available for review, and 227 were classified as patients with Lyme disease. The MTTT algorithm detected 25% more early infections with a specificity of 99.56% (99.41 to 99.68%) compared to the STTT. These are the first Canadian data to show that serology using a whole-cell sonicate EIA followed by a C6 EIA (MTTT) had improved sensitivity for detecting early B. burgdorferi infection with specificity similar to that of two-tiered testing using Western blots.

| S-EPMC7315017 | biostudies-literature

Diagnostic parameters of modified two-tier testing in European patients with early Lyme disease.

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| S-EPMC7561539 | biostudies-literature

High Throughput Newborn Screening for Sickle Cell Disease - Application of Two-Tiered Testing with a qPCR-Based Primary screen.

Project description:BackgroundSickle cell disease (SCD) is a group of hemoglobinopathies with a common point mutation causing the production of sickle cell hemoglobin (HbS). In high-throughput newborn screening (NBS) for SCD, a two-step procedure is suitable, in which qPCR first pre-selects relevant samples that are differentiated by a second method.MethodsThree NBS centers using qPCR-based primary screening for SCD performed a laboratory comparison. Methods using tandem MS or HPLC were used for differentiation.ResultsIn a benchmarking test, 450 dried blood samples were analyzed. Samples containing HbS were detected as reliably by qPCR as by methods established for hemoglobinopathy testing. In a two-step screening approach, the 2nd-tier-analyses have to distinguish the carrier status from pathological variants. In nine months of regular screening, a total of 353,219 samples were analyzed using two-stage NBS procedures. The 1st-tier screening by qPCR reduced the number of samples for subsequent differentiation by>99.5%. Cases with carrier status or other variants were identified as inconspicuous while 78 cases with SCD were revealed. The derived incidence of 1:4,773, is in good agreement with previously published incidences.ConclusionIn high-throughput NBS for SCD, qPCR is suitable to focus 2nd-tier analyses on samples containing HbS, while being unaffected by factors such as prematurity or transfusions. The substantial reduction of samples numbers positively impacts resource conservation, sustainability, and cost-effectiveness. No false negative cases came to attention.

| S-EPMC10635756 | biostudies-literature

Evaluation of fecal occult blood testing for rapid diagnosis of invasive diarrhea in young children.

Project description:Antimicrobials are only indicated in acute childhood diarrhea if it is invasive or persistent. Rapid screening for invasive diarrhea can therefore inform treatment decisions but pathogen identification by culture is slow, expensive and cumbersome. This study aimed to assess the diagnostic utility of stool microscopy and immunochromatographic fecal occult blood test (FOBT) kits for identifying invasive or potentially invasive diarrhea in Ibadan, Nigeria. Fecal specimens from 46 children under 5 years old with diarrhea, collected as part of ongoing case-control studies, were subjected to stool microscopy for erythrocytes and leucocytes, and FOBT using the innovator's product and four locally procurable generic immunochromatographic kits, each according to manufacturers' instructions. Stool specimens were cultured for enteric bacterial pathogens using standard procedures. Presumptive pathogen isolates were identified biochemically and by PCR, and then confirmed by whole genome sequencing. Shigella, enteroinvasive Escherichia coli and Yersinia, pathogens that invariably cause invasive diarrhea, were detected in five of 46 specimens. Occult blood detection by microscopy was 55.6% sensitive and 78.4% specific, while the innovator's FOBT product was respectively 62.5% and 81.6% sensitive and specific compared to strict invasive pathogen recovery. Microscopy and FOBT testing were less sensitive in identifying specimens that contained pathogens that do not always elicit invasive diarrhea. Generic FOBT tests compared well with the innovator's product. Microscopy and FOBT testing have some value for delineating likely invasive diarrheas. They could inform treatment and serve as early warning indicators for dysentery outbreaks in resource limited settings. Inexpensive, generic FOBT kits that are locally procurable in Nigeria performed as well as the innovator's product.

| S-EPMC10358913 | biostudies-literature

Evaluation of Patient IgM and IgG Reactivity Against Multiple Antigens for Improvement of Serodiagnostic Testing for Early Lyme Disease.

Project description:Serologic testing is the standard for laboratory diagnosis and confirmation of Lyme disease. Serodiagnostic assays to detect antibodies against Borrelia burgdorferi, the agent of Lyme borreliosis, are used for detection of infection. However, serologic testing within the first month of infection is less sensitive as patients' antibody responses continue to develop. Previously, we screened several B. burgdorferi in vivo expressed antigens for candidates that elicit early antibody responses in patients with Stage 1 and 2 Lyme disease. We evaluated patient IgM seroreactivity against 6 antigens and found an increase in sensitivity without compromising specificity when compared to current IgM second-tier immunoblot scoring. In this study, we continued the evaluation using a multi-antigen panel to measure IgM plus IgG seroreactivity in these early Lyme disease patients' serum samples. Using two statistical methods for calculating positivity cutoff values, sensitivity was 70 and 84-87%, for early acute and early convalescent Lyme disease patients, respectively. Specificity was 98-100% for healthy non-endemic control patients, and 96-100% for healthy endemic controls depending on the statistical analysis. We conclude that improved serologic testing for early Lyme disease may be achieved by the addition of multiple borrelial antigens that elicit IgM and IgG antibodies early in infection.

| S-EPMC6906137 | biostudies-literature

Diagnosis of Kawasaki Disease in children using host RNA expression

Project description:This SuperSeries is composed of the SubSeries listed below.

2018-08-09 | GSE73464 | GEO

Improved Serodiagnostic Performance for Lyme Disease by Use of Two Recombinant Proteins in Enzyme-Linked Immunosorbent Assay Compared to Standardized Two-Tier Testing.

Project description:The most reliable test method for the serological confirmation of Lyme disease (LD) is a 2-tier method recommended by the CDC in 1995. The first-tier test is a low-specificity enzyme-linked immunosorbent assay (ELISA), and the second-tier tests are higher-specificity IgG and IgM Western blots. This study describes the selection of two Borrelia burgdorferi recombinant proteins and evaluation of their performance in a simple 1-tier test for the serological confirmation of LD. These two proteins were generated from (i) the full-length dbpA gene combined with the invariable region 6 of the vlsE gene (DbpA/C6) and (b) the full-length ospC gene (OspC). The expressed DbpA/C6 and OspC proteins were useful in detecting anti-Borrelia IgG and IgM antibodies, respectively. A blind study was conducted on a well-characterized panel of 279 human sera from the CDC, comparing ELISAs using these two recombinant antigens with the 2-tier test method. The two methods (DbpA/C6-OspC versus 2-tier test) were equivalent in identifying sera from negative-control subjects (99% and 100% specificity, respectively) and in detecting stage II and III LD patient sera (100% and 100% sensitivity). However, the DbpA/C6-OspC ELISA was markedly better (80% versus 63%) than the 2-tier test method in detecting anti-Borrelia antibodies in stage I LD patients. The findings suggest that these antigens could be used in a simple 1-tier ELISA that is faster to perform, easier to interpret, and less expensive than the 2-tier test method and which is better at detecting Borrelia-specific antibodies in sera from patients with stage I LD.

| S-EPMC5625391 | biostudies-literature

Lyme disease testing by large commercial laboratories in the United States.

Project description:BackgroundLaboratory testing is helpful when evaluating patients with suspected Lyme disease (LD). A 2-tiered antibody testing approach is recommended, but single-tier and nonvalidated tests are also used. We conducted a survey of large commercial laboratories in the United States to assess laboratory practices. We used these data to estimate the cost of testing and number of infections among patients from whom specimens were submitted.MethodsLarge commercial laboratories were asked to report the type and volume of testing conducted nationwide in 2008, as well as the percentage of positive tests for 4 LD-endemic states. The total direct cost of testing was calculated for each test type. These data and test-specific performance parameters available in published literature were used to estimate the number of infections among source patients.ResultsSeven participating laboratories performed approximately 3.4 million LD tests on approximately 2.4 million specimens nationwide at an estimated cost of $492 million. Two-tiered testing accounted for at least 62% of assays performed; alternative testing accounted for <3% of assays. The estimated frequency of infection among patients from whom specimens were submitted ranged from 10% to 18.5%. Applied to the total numbers of specimens, this yielded an estimated 240 000 to 444 000 infected source patients in 2008.DiscussionLD testing is common and costly, with most testing in accordance with diagnostic recommendations. These results highlight the importance of considering clinical and exposure history when interpreting laboratory results for diagnostic and surveillance purposes.

| S-EPMC4646413 | biostudies-literature

Testing Raman spectroscopy as a diagnostic approach for Lyme disease patients.

Project description:Lyme disease (LD), the leading tick-borne disease in the Northern hemisphere, is caused by spirochetes of several genospecies of the Borreliella burgdorferi sensu lato complex. LD is a multi-systemic and highly debilitating illness that is notoriously challenging to diagnose. The main drawbacks of the two-tiered serology, the only approved diagnostic test in the United States, include poor sensitivity, background seropositivity, and cross-reactivity. Recently, Raman spectroscopy (RS) was examined for its LD diagnostic utility by our earlier proof-of-concept study. The previous investigation analyzed the blood from mice that were infected with 297 and B31 strains of Borreliella burgdorferi sensu stricto (s.s.). The selected strains represented two out of the three major clades of B. burgdorferi s.s. isolates found in the United States. The obtained results were encouraging and prompted us to further investigate the RS diagnostic capacity for LD in this study. The present investigation has analyzed blood of mice infected with European genospecies, Borreliella afzelii or Borreliella garinii, or B. burgdorferi N40, a strain of the third major class of B. burgdorferi s.s. in the United States. Moreover, 90 human serum samples that originated from LD-confirmed, LD-negative, and LD-probable human patients were also analyzed by RS. The overall results demonstrated that blood samples from Borreliella-infected mice were identified with 96% accuracy, 94% sensitivity, and 100% specificity. Furthermore, human blood samples were analyzed with 88% accuracy, 85% sensitivity, and 90% specificity. Together, the current data indicate that RS should be further explored as a potential diagnostic test for LD patients.

| S-EPMC9647194 | biostudies-literature

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