SARS-CoV-2 in asymptomatic pregnant women in South Brazil: RT-PCR and serological detection
Objectives: This study aims to detect the SARS-CoV-2 infection prevalence in asymptomatic pregnant women.
Methods: A group of 195 asymptomatic pregnant women who attended the prenatal care outclinic and to the obstetric emergency department was tested concomitantly for SARS-CoV-2 by RT-PCR and serological tests.
Results: The virus was detected by RT-PCR in two (1.02%) cases and 17 (8.71%) patients had antibodies detected by immunochromatographic tests.
Conclusions: Due to the high risk of this emerging infection in the health of pregnant women, fetuses and newborns, we suggest the universal screening of all pregnant women admitted to hospital through the combined method RT-PCR and serological.
SaQuant: a real-time PCR assay for quantitative assessment of Staphylococcus aureus
Background: Molecular assays are important tools for pathogen detection but need to be periodically re-evaluated with the discovery of additional genetic diversity that may cause assays to exclude target taxa or include non-target taxa. A single well-developed assay can find broad application across research, clinical, and industrial settings. Pathogen prevalence within a population is estimated using such assays and accurate results are critical for formulating effective public health policies and guiding future research. A variety of assays for the detection of Staphylococcus aureus are currently available. The utility of commercial assays for research is limited, given proprietary signatures and lack of transparent validation.
Results: In silico testing of existing peer-reviewed assays show that most suffer from a lack of sensitivity and specificity. We found no assays that were specifically designed and validated for quantitative use. Here we present a qPCR assay, SaQuant, for the detection and quantification of S. aureus as might be collected on sampling swabs. Sensitivity and specificity of the assay was 95.6 and 99.9 %, respectively, with a limit of detection of between 3 and 5 genome equivalents and a limit of quantification of 8.27 genome equivalents. The presence of DNA from non-target species likely to be found in a swab sample, did not impact qualitative or quantitative abilities of the assay.
Conclusions: This assay has the potential to serve as a valuable tool for the accurate detection and quantification of S. aureus collected from human body sites in order to better understand the dynamics of prevalence and transmission in community settings.
Molecular typing, biofilm production, and detection of carbapenemase genes in multidrug-resistant Acinetobacter baumannii isolated from different infection sites using ERIC-PCR in Hamadan, west of Iran
Background: Acinetobacter baumannii is an opportunistic pathogen that can cause several kinds of nosocomial infections. Increasing antibiotic resistance as well as identifying genetic diversity and factors associated with pathogenicity and prevalence of this bacterium is important. The aim of this study was the investigation of molecular typing, biofilm production, and detection of carbapenemase genes in multidrug-resistant Acinetobacter baumannii isolated from different infection sites using ERIC-PCR in Iran.
Methods: Forty isolates of A. baumannii were obtained from various wards of the central hospital, in the west of Iran. Phenotypic identification and genetic diversity, biofilm production assay, and detection of Carbapenemase genes carried out.
Results: Tracheal samples 26 (61.9 %) are the most frequent isolates, and 95 % of isolates were identified as MDR. 32.5 % of all A. baumannii strains were capable to form a strong biofilm. It was founded that antimicrobial resistance patterns had a significant relationship with strong biofilm formation (P = 0.001). Most frequencies of the studied genes were in the order of VIM (81 %), SPM (45.2 %), and IMP (35.7 %) genes. The VIM gene was the most frequent in all isolates which were significant (P = 0.006). 14 different ERIC-types were observed including 7 common types and 7 unique or single types. F type is the largest common type consisting of nine isolates and B, D, and E types contain two isolates separately.
Conclusions: ERIC-PCR technique was used to genetically classify A. baumannii isolates as one of the most common microorganisms in nosocomial infections.
Development of a reverse transcription droplet digital PCR assay for sensitive detection of peach latent mosaic viroid
Peach latent mosaic viroid (PLMVd) represents a continuing threat to peach tree production worldwide. In this study, a sensitive and accurate quantification of PLMVd in peach leaves was established using a reverse transcription droplet digital polymerase chain reaction (RT-ddPCR) assay. The quantitative linearity, accuracy, and sensitivity of RT-ddPCR for the detection of PLMVd were comparatively assessed to those of reverse-transcription real-time quantitative polymerase chain reaction (RT-qPCR) assay. The specificity assay shows no amplification in major peach viruses, apple chlorotic leaf spot virus and prunus necrotic ring spot virus and negative control.
Furthermore, the levels of PLMVd transcripts determined using RT-ddPCR and RT-qPCR showed a high degree of linearity and quantitative correlation. Our results also indicated that the RT-ddPCR assay is at least two-fold more sensitive than qPCR and could therefore, be used to detect PLMVd in field samples. Moreover, optimization of RT-ddPCR was found to enhance the sensitivity of PLMVd detection in the peach leaf samples with low viral loads. In summary, the established RT-ddPCR assay represents a promising alternative method for the precise quantitative detection of PLMVd; it would be particularly applicable for diagnosing PLMVd infections in plant quarantine inspection and PLMVd-free certification program.
Prospective Evaluation of Xpert Xpress Strep A Automated PCR Assay vs. Solana Group A Streptococcal Nucleic Acid Amplification Testing vs. Conventional Throat Culture
In our laboratory, the negative rapid group A streptococcal (GAS) antigen assays are backed up by the Solana® GAS Assay by Quidel instead of a Group A streptococcal throat culture. Another FDA cleared RT-PCR assay is the Xpert® Xpress Strep A, which detects Streptococcus pyogenes DNA, and is performed on the Cepheid GeneXpert instrument. Three hundred seventy-five positive and negative specimens were randomly selected from 5489 throat specimens that had been tested by the Solana® GAS Assay during January 2018 and were tested with the Xpress Strep A assay. A throat culture was also set up (sheep blood agar at 35 °C in 5% CO2).
All beta-hemolytic streptococci were purified and identified by MALDI-TOF mass spectrometry. Of the 375 samples, 185 were positive by Solana® GAS Assay, and 187 were positive by the Xpress Strep A. The total agreement between the Solana® GAS Assay and the Xpert® Xpress Strep A was 99.5%. The agreement of the Xpert® Xpress Strep A assay with culture was 90.1%. The sensitivity and specificity for Xpress Strep A versus culture were 100% and 83.5%, respectively. The Xpert® Xpress Strep A assay’s performance was equivalent to the Solana® GAS Assay, and was highly sensitive. The lower specificity was likely due to the Xpress Strep A assay having higher sensitivity as compared to throat culture.