Getting the measure of syphilis: qPCR to better understand early infection.
Ontology highlight
ABSTRACT: Objectives
Until recently, PCR had been used to detect but not quantify Treponema pallidum. To understand infection kinetics of this uncultivable organism, a real-time PCR assay was developed to quantify 47 kDa membrane lipoprotein gene DNA (tpp47).Methods
Assay specificity was determined against DNA from humans, skin organisms and sexually transmitted pathogens. tpp47 DNA (Nichols strain) was used to construct a standard curve for T pallidum quantification. Blood and ulcer samples were obtained from 99 patients being investigated or screened for syphilis and tpp47 was quantified.Results
The assay was specific, not cross-reactive with other organisms tested and sensitive, with a detection limit of a single copy of tpp47 DNA. For ulcer samples, the assay was 100% sensitive and 97.14% specific. Sensitivity fell to 34.1% for blood samples but specificity remained high (100%). tpp47 DNA was more commonly detected, and at a higher copy number, in blood of patients with secondary infection (sensitivity 57.89%) compared with primary infection. Quantity of tpp47 DNA was higher in primary infection ulcers, especially in HIV-1-positive patients, than in ulcers persisting into secondary disease.Conclusions
Quantifying T pallidum provides insight into syphilis infection kinetics: Ulcers of primary disease in HIV-1-positive patients are perhaps more infectious and the presence and load of T pallidum bacteraemia is variable, with a peak in the secondary stage. Quantitative PCR has the potential to map T pallidum infection and to highlight the impact of HIV on syphilis.
SUBMITTER: Tipple C
PROVIDER: S-EPMC3252622 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Transcription factors modulate gene expression by distinct, barely understood mechanisms. The crystal structure of a bacterial transcription subcomplex comprising the effector domain of factor PhoB, its target DNA and the ?4 domain of the RNA polymerase ?70 subunit supports the notion that a stronger grip on the promoter-factor complex results in an enhanced RNAP architecture.
| S-EPMC3654763 | biostudies-literature
Project description:Some studies have shown that manual asymmetries decrease in older age. These results have often been explained with reference to models of reduced hemispheric specialisation. An alternative explanation, however, is that hand differences are subtle, and capturing them requires tasks that yield optimal performance with both hands. Whereas the hemispheric specialisation account implies that reduced manual asymmetries should be reliably observed in older adults, the 'measurement difficulty' account suggests that manual asymmetries will be hard to detect unless a task has just the right level of difficulty--i.e. within the 'Goldilocks Zone', where it is not too easy or too hard, but just right. Experiment One tested this hypothesis and found that manual asymmetries were only detected when participants performed in this zone; specifically, performance on a tracing task was only superior in the preferred hand when task constraints were high (i.e. fast speed tracing). Experiment Two used three different tasks to examine age differences in manual asymmetries; one task produced no asymmetries, whilst two tasks revealed asymmetries in both younger and older groups (with poorer overall performance in the old group across all tasks). Experiment Three revealed task-dependent asymmetries in both age groups, but highlighted further detection difficulties linked with the metric of performance and compensatory strategies used by participants. Results are discussed with reference to structural learning theory, whereby we suggest that the processes of inter-manual transfer lead to relatively small performance differences between the hands (despite a strong phenomenological sense of performance disparities).
| S-EPMC4449126 | biostudies-literature
Project description:The standard approach for quantitative estimation of genetic materials with qPCR is calibration with known concentrations for the target substance, in which estimates of the quantification cycle (Cq ) are fitted to a straight-line function of log(N 0), where N 0 is the initial number of target molecules. The location of Cq for the unknown on this line then yields its N 0. The most widely used definition for Cq is an absolute threshold that falls in the early growth cycles. This usage is flawed as commonly implemented: threshold set very close to the baseline level, which is estimated separately, from designated "baseline cycles." The absolute threshold is especially poor for dealing with the scale variability often observed for growth profiles. Scale-independent markers, like the first derivative maximum (FDM) and a relative threshold (Cr ) avoid this problem. We describe improved methods for estimating these and other Cq markers and their standard errors, from a nonlinear algorithm that fits growth profiles to a 4-parameter log-logistic function plus a baseline function. Further, by examining six multidilution, multireplicate qPCR data sets, we find that nonlinear expressions are often preferred statistically for the dependence of Cq on log(N 0). This means that the amplification efficiency E depends on N 0, in violation of another tenet of qPCR analysis. Neglect of calibration nonlinearity leads to biased estimates of the unknown. By logic, E estimates from calibration fitting pertain to the earliest baseline cycles, not the early growth cycles used to estimate E from growth profiles for single reactions. This raises concern about the use of the latter in lengthy extrapolations to estimate N 0. Finally, we observe that replicate ensemble standard deviations greatly exceed predictions, implying that much better results can be achieved from qPCR through better experimental procedures, which likely include reducing pipette volume uncertainty.
| S-EPMC6554483 | biostudies-literature
Project description:An important data gap in our understanding of the phyllosphere surrounds the origin of the many microbes described as phyllosphere communities. Most sampling in phyllosphere research has focused on the collection of microbiota without the use of a control, so the opportunity to determine which taxa are actually driven by the biology and physiology of plants as opposed to introduced by environmental forces has yet to be fully realized. To address this data gap, we used plastic plants as inanimate controls adjacent to live tomato plants (phyllosphere) in the field with the hope of distinguishing between bacterial microbiota that may be endemic to plants as opposed to introduced by environmental forces. Using 16S rRNA gene amplicons to study bacterial membership at four time points, we found that the vast majority of all species-level operational taxonomic units were shared at all time-points. Very few taxa were unique to phyllosphere samples. A higher taxonomic diversity was consistently observed in the control samples. The high level of shared taxonomy suggests that environmental forces likely play a very important role in the introduction of microbes to plant surfaces. The observation that very few taxa were unique to the plants compared to the number that were unique to controls was surprising and further suggests that a subset of environmentally introduced taxa thrive on plants. This finding has important implications for improving our approach to the description of core phytobiomes as well as potentially helping us better understand how foodborne pathogens may become associated with plant surfaces.
| S-EPMC5036865 | biostudies-literature
Project description:When we are fatigued, we feel that our performance is worse than when we are fresh. Yet, for over 100 years, researchers have been unable to identify an objective, behavioral measure that covaries with the subjective experience of fatigue. Previous work suggests that the metrics of signal detection theory (SDT)-response bias (criterion) and perceptual certainty (d')-may change as a function of fatigue, but no work has yet been done to examine whether these metrics covary with fatigue. Here, we investigated cognitive fatigue using SDT. We induced fatigue through repetitive performance of the n-back working memory task, while functional magnetic resonance imaging (fMRI) data was acquired. We also assessed cognitive fatigue at intervals throughout. This enabled us to assess not only whether criterion and d' covary with cognitive fatigue but also whether similar patterns of brain activation underlie cognitive fatigue and SDT measures. Our results show that both criterion and d' were correlated with changes in cognitive fatigue: as fatigue increased, subjects became more conservative in their response bias and their perceptual certainty declined. Furthermore, activation in the striatum of the basal ganglia was also related to cognitive fatigue, criterion, and d'. These results suggest that SDT measures represent an objective measure of cognitive fatigue. Additionally, the overlap and difference in the fMRI results between cognitive fatigue and SDT measures indicate that these measures are related while also separate. In sum, we show the relevance of SDT measures in the understanding of fatigue, thus providing researchers with a new set of tools with which to better understand the nature and consequences of cognitive fatigue.
| S-EPMC7844088 | biostudies-literature
Project description:Treponema pallidum infections can have severe complications if not diagnosed and treated at an early stage. Screening and diagnosis of syphilis require assays with high specificity and sensitivity. The Elecsys Syphilis assay is an automated treponemal immunoassay for the detection of antibodies against T. pallidum The performance of this assay was investigated previously in a multicenter study. The current study expands on that evaluation in a variety of diagnostic settings and patient populations, at seven independent laboratories. The samples included routine diagnostic samples, blood donation samples, samples from patients with confirmed HIV infections, samples from living organ or bone marrow donors, and banked samples, including samples previously confirmed as syphilis positive. This study also investigated the seroconversion sensitivity of the assay. With a total of 1,965 syphilis-negative routine diagnostic samples and 5,792 syphilis-negative samples collected from blood donations, the Elecsys Syphilis assay had specificity values of 99.85% and 99.86%, respectively. With 333 samples previously identified as syphilis positive, the sensitivity was 100% regardless of disease stage. The assay also showed 100% sensitivity and specificity with samples from 69 patients coinfected with HIV. The Elecsys Syphilis assay detected infection in the same bleed or earlier, compared with comparator assays, in a set of sequential samples from a patient with primary syphilis. In archived serial blood samples collected from 14 patients with direct diagnoses of primary syphilis, the Elecsys Syphilis assay detected T. pallidum antibodies for 3 patients for whom antibodies were not detected with the Architect Syphilis TP assay, indicating a trend for earlier detection of infection, which may have the potential to shorten the time between infection and reactive screening test results.
| S-EPMC5005501 | biostudies-literature
Project description:High rates of syphilis have been reported among men who have sex with men (MSM) internationally. Guidelines recommend presumptive treatment of sexual contacts of individuals with syphilis at the point of care. The aim of this study was to determine the proportion who were infected with syphilis and the factors predictive of infection among men reporting contact with a man with syphilis.Contacts who were syphilis infected (cases) were compared with those who were uninfected (controls).This study was conducted at the main public sexually transmitted diseases clinic in Victoria, Australia.One hundred and seventy-two MSM presenting as sexual contacts of men with syphilis at a sexual health service in Melbourne, Australia, between July 2007 and October 2011 were assessed for syphilis.Proportion of MSM who are infected with syphilis and factors associated with infection.Of the 172 men who presented reporting contact with syphilis, 26 (15%, 95% CI 10 to 20%) had syphilis. One man had primary syphilis, 4 had secondary syphilis, while the remaining 21 had early latent syphilis. Infection was associated with unprotected anal sex over the prior 3 months (adjusted OR 6.1, 95% CI 1.4 to 26.8).One in seven men presenting as contacts of syphilis had syphilis infection, most of whom were latently infected. Contacts reporting recent unprotected anal sex were more likely to have syphilis.
| S-EPMC3425903 | biostudies-other
Project description:In order to understand the developmental trajectories of early lymphocyte development it is crucial to prospectively isolate stage and lineage specific cells. It has become clear that early lymphoid progenitor compartments in the bone marrow are molecularly and functionally heterogeneous which warrants further investigation to refine the marker combinations used to isolate these progenitors. An initial antibody screen revealed extensive surface marker heterogeneity amongst early lymphoid progenitors. This heterogeneity was resolved using single cell gene expression assays and single cell in vitro differentiation assays, identifying marker combinations that isolate functionally distinct populations. In addition, using reporter transgenic mice we were able to identify a set of surface markers that can be used alone or in combination with classical targets to identify specific stages of B-cell development. B-cell stages based on transgene expression which were used for screening purposes were verified by RNASeq. The data provides a greater resolution of the complexity of the lymphoid progenitor compartment within the bone marrow than has been understood to date and provides novel tools for the further identification of cell populations in B-lineage development. qPCR gene expression profiling of mouse common lymphoid progenitors (CLPs). Four 96-well plates with pre-amplified single CLP bone marrow cell cDNA (with 10 and 20 cell controls) were loaded on to a 96.96 Dynamic Array Chip for Gene Expression. Samples Ids follow where SS<X> denotes well ID, and P<X>denoted the plate. Controls include well where no RT is performed (noRT) and 10 and 20 cell controls.
2016-10-23 | E-GEOD-74606 | biostudies-arrayexpress
Project description:In order to understand the developmental trajectories of early lymphocyte development it is crucial to prospectively isolate stage and lineage specific cells. It has become clear that early lymphoid progenitor compartments in the bone marrow are molecularly and functionally heterogeneous which warrants further investigation to refine the marker combinations used to isolate these progenitors. An initial antibody screen revealed extensive surface marker heterogeneity amongst early lymphoid progenitors. This heterogeneity was resolved using single cell gene expression assays and single cell in vitro differentiation assays, identifying marker combinations that isolate functionally distinct populations. In addition, using reporter transgenic mice we were able to identify a set of surface markers that can be used alone or in combination with classical targets to identify specific stages of B-cell development. B-cell stages based on transgene expression which were used for screening purposes were verified by RNASeq. The data provides a greater resolution of the complexity of the lymphoid progenitor compartment within the bone marrow than has been understood to date and provides novel tools for the further identification of cell populations in B-lineage development.
2017-10-26 | GSE74606 | GEO
Project description:The purpose of this registry study is to create a database-a collection of information-for better understanding young onset colorectal cancer. Colorectal cancer patients are considered to have young onset colorectal cancer if they are diagnosed with their cancer before the age of 50. Researchers will use the information from this database to learn more about how young onset colorectal cancer may be similar to or different from colorectal cancer that is diagnosed later in life. Researchers will also use information from the database for current and future research on young onset colorectal cancer.
| 2346978 | ecrin-mdr-crc