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Rapid screening of Veillonella by ultraviolet fluorescence.

Among 51 strains of anaerobic gram-negative cocci belonging to the family Veillonellaceae, all strains of Veillonella (V. parvula and V. alcalescens) displayed red fluorescence under long-wave (366 nm) ultraviolet light, whereas no Acidaminococcus or Megasphaera demonstrated fluorescence. In contrast to Bacteroides melaninogenicus, growth of Veillonella does not require hemin and menadione, and flourescence is rapidly lost upon exposure to air. The fluorescent component of a strain of V. parvula examined could not be extracted in solution with water, ether, methanol, or chloroform, but was readily extracted with 0.4 N NaOH. Spectrophotofluorometrically, the fluorescence maximum of this extract was 660 nm with an excitation maximum of 300 nm, when measured at pH 7.2 and 25 C. Coupled with the Gram stain, ultraviolet fluorescence may be a useful tool for rapid screening of Veillonella and is particularly helpful for detection and, isolation of this organism from mixed culture.

Air↗

Rapid screening for freshwater bacterial groups by using reverse line blot hybridization.

The identification of phylogenetic clusters of bacteria that are common in freshwater has provided a basis for probe design to target important freshwater groups. We present a set of 16S ribosomal RNA gene-based oligonucleotide probes specific for 15 of these freshwater clusters. The probes were applied in reverse line blot hybridization, a simple method that enables the rapid screening of PCR products from many samples against an array of probes. The optimized assay was made stringent to discriminate at approximately the single-mismatch level. This made 10 of the probes highly specific, with at least two mismatches to the closest noncluster member in the global database. Screening of PCR products from bacterioplankton of 81 diverse lakes from Belgium, The Netherlands, Denmark, Sweden, and Norway showed that the respective probes were reactive against 5 to 100% of the lake samples. Positive reactivity of six highly specific probes showed that bacteria from actinobacterial clusters ACK-M1 and Sta2-30 and from verrucomicrobial cluster CLO-14 occurred in at least 90% of the investigated lakes. Furthermore, bacteria from alpha-proteobacterial cluster LD12 (closely related to the marine SAR11 cluster), beta-proteobacterial cluster LD28 and cyanobacterial cluster Synechococcus 6b occurred in more than 70% of the lakes. Reverse line blot hybridization is a new tool in microbial ecology that will facilitate research on distribution and habitat specificity of target species at relatively low costs.

Animals↗

A rapid screening enzyme linked immunosorbent assay for autoantibodies to extractable nuclear antigens.

An enzyme linked immunosorbent assay (ELISA) has been developed to detect antinuclear antibodies to extractable nuclear antigens (ENA)--a rabbit thymus nuclear derivative. A second ELISA has also been developed (ENA RNA-ase ELISA) to analyse the relative contribution of the components of ENA (nucleoprotein and RNA antigens) to antibody binding as detected by the ELISA assays. Both ELISA assays provide a rapid screening method for antibodies to soluble, diffusable nuclear antigens that could otherwise go undetected by immunodiffusion or the indirect immunofluorescent method that uses unfixed frozen sections of rat liver. The considerable improvement that these ELISA assays offer in efficiency and both diagnostic and serological sensitivity over the traditional immunodiffusion method used for specificity characterisation of ENA autoantibodies is discussed.

Antibody Specificity↗

Dideoxy fingerprinting for rapid screening of rpoB gene mutations in clinical isolates of Mycobacterium tuberculosis.

Rifampin is a key component of therapeutic regimens for tuberculosis control, and a marker for multidrug resistance of Mycobacterium tuberculosis. Mutations responsible for conferring rifampin resistance in M. tuberculosis are known to occur in a 69-bp region of the rpoB gene. In this study, we assessed the accuracy of dideoxy fingerprinting (ddF), a hybrid technique employing elements of dideoxy sequencing and single-strand polymorphism analysis, for rapid screening of rifampin resistance in clinical isolates of M. tuberculosis. This technique was used to analyze 72 M. tuberculosis isolates. The results were compared with those of automated dideoxy sequencing and the antibiotic resistance profile (determined with the BACTEC system). Of the 72 isolates, 50 were rifampin resistant. The ddF findings were completely consistent with those of dideoxy sequencing in all isolates. In 68 (94%) isolates, the ddF findings were consistent with the rifampin resistance status determined with the BACTEC system; all four isolates with inconsistent results had no mutation in the 69-bp region, but were resistant to rifampin. Our findings suggest that ddF accurately detects mutations in the rifampin resistance-associated 69-bp region of the rpoB gene in clinical isolates of M. tuberculosis, and may be a valuable screening tool for rifampin resistance.

DNA Fingerprinting↗

Emergence of protease inhibitor resistance mutations in human immunodeficiency virus type 1 isolates from patients and rapid screening procedure for their detection.

Patient human immunodeficiency virus type 1 (HIV-1) isolates that are resistant to protease inhibitors may contain amino acid substitutions L10I/V, M46L/I, G-48V, L63P, V82A/F/T, I84V, and L90M in the protease gene. Substitutions at positions 82 and/or 90 occur in variants that display high levels of resistance to certain protease inhibitors. Nucleotide substitutions at these two sites also lead to the loss of two HindII restriction enzyme digestion sites, and these changes make possible a rapid procedure for the detection of drug-resistant variants in patients on protease inhibitor therapy. This procedure was used to detect the emergence of mutated viruses at various times after the initiation of therapy with the HIV-1 protease inhibitor indinavir. The method includes viral RNA isolation from plasma and reverse transcription PCR amplification of the protease gene with fluorescence-tagged primers. The PCR product is digested with HindII, the cleavage products are separated on a urea-acrylamide gel in a DNA sequencer, and the extent of cleavage is automatically analyzed with commercially available software. In viruses from 34 blood samples from four patients, mutations leading to an amino acid change at residue 82 appeared as early as 6 weeks after the start of therapy and persisted throughout the course of the study period (48 weeks). Mutations leading to double substitutions at residues 82 and 90 were seen at a lower frequency and appeared later than the change at position 82. The changes detected by restriction enzyme cleavage were confirmed by DNA sequencing of the cloned protease genes by reverse transcription PCR amplification of viral RNA from isolates in plasma. In addition to the changes at positions 82 and 90, we have identified M46L/I, G48V, and I54V substitutions in isolates derived from indinavir-treated patients. HindII analysis of uncloned, PCR-amplified DNA offers a rapid screening procedure for the detection of virus isolates containing mutations at amino acid residues 82 and 90 in the HIV-1 protease gene. By using other restriction enzymes, the same method can be used to detect additional protease drug-resistant variants and is generally applicable for the detection of mutations.

Amino Acid Sequence↗

Highly sensitive and selective fluorescence assays for rapid screening of endothelin-converting enzyme inhibitors.

The highly potent vasoconstrictor peptide endothelin (ET) is generated from an inactive precursor, big endothelin (bET), by endothelin-converting enzyme (ECE). ECE is a phosphoramidon-sensitive zinc metallopeptidase, which is closely related to neprilysin (neutral endopeptidase). It is possible that compounds which inhibit the formation of ET may be used as new drugs for the treatment of cardiovascular diseases. Such an approach requires a fast, simple and selective assay to measure ECE activity, allowing rapid screening of inhibitors. We describe here two new ECE substrates based on the concept of 'intramolecularly quenched fluorescence' which may fulfill this aim. One, S(1) [Pya(21)-Nop(22)-bET-1(19--35)], is the (19--35) fragment of the natural peptide big-ET-1(1--38), which is modified by introducing the fluorescent amino acid, pyrenylalanine (Pya), in position 21 and a quencher, p-nitrophenylalanine (Nop), in position 22. The second substrate (S(2)) is a small peptide, Ac-Ser-Gly-Pya-Lys-Ala-Phe-Ala-Nop-Gly-Lys-NH(2), from a biased substrate peptide library. The recombinant, hECE-1c, cleaved both Pya(21)-Nop(22)-bET-1(19--35) and the natural substrate selectively between residues 21 and 22, whereas cleavage occurred between alanine and phenylalanine in the small peptide. In both cases, this generated intense fluorescence emission. The synthesis and kinetic parameters of these substrates are described. These assays, which can be used directly on tissue homogenates, are the most sensitive and selective described to date for ECE, and are easily automated for a high-throughput screening of inhibitors.

Amino Acid Sequence↗

[The development of rapid screening of colorectal cancer by mutation detection in fecal DNA].

Colorectal cancer is one of the leading causes of death in the world. Screening high-risk populations for colorectal cancer can lower both the incidence of and deaths from the disease. Mutation detection in fecal DNA is a new approach of screening patients for colorectal cancer. Compared with colonoscopy and fecal occult-blood test, this approach has the advantages of high specificity, high sensitivity and general acceptability. This paper systematically reviewed genes associated with fecal DNA testing, methods for extracting tumor-specific DNA, and methods for detecting gene mutations. Moreover, the application of mutation detection using fecal DNA for rapid screening of colorectal cancer was briefly discussed.

Colorectal Neoplasms↗

EMTscore infers divergent EMT pathways from omics data and enables rapid screening for EMT-associated gene sets.

MOTIVATION: Quantitative analyses of epithelial-mesenchymal transition (EMT) have been widely used in several areas of biomedical sciences due to its importance in development and cancer progression, but its multi-contextual nature requires standardization and implementation of gene set scoring methods beyond capacities of conventional tools. RESULTS: We developed EMTscore, a package that provides an efficient implementation of unbiased scoring methods for multiple EMT pathways using individual single-cell or bulk omics data, and the package allows rapid screening for cellular processes correlated with EMT. AVAILABILITY AND IMPLEMENTATION: EMTscore is available from GitHub https://github.com/wenmm/EMTscore under the GNU General Public License, and is uploaded on Zenodo with a DOI 10.5281/zenodo.19487376.

Epithelial-Mesenchymal Transition↗

Rapid screening of mtDNA coding region SNPs for the identification of west European Caucasian haplogroups.

This work presents a selection of 16 SNPs from the coding region of the human mitochondrial DNA. The selected markers are used for the assignment of individuals to one of the nine major European Caucasian mitochondrial haplogroups. The selected SNPs are targeted in two multiplex systems, via the application of the SNaPshot kit, a multiplex method based on the dideoxy single-base extension of unlabeled oligonucleotide primers. The method is conceived as a rapid screening technique prior to sequencing analysis, in order to eliminate multiple suspects from an inquiry or to discriminate between stains in a high volume casework example. Moreover, the ability to assign an unknown sample to an mtDNA type of known Caucasian origin could be of probative value in some investigations. A database of 277 Austrian Caucasians has been constructed, and the probability of a chance match between two unrelated individuals is calculated as 11.4%. This novel multiplex PCR amplification and typing system for mtDNA coding region SNPs promises to be a convenient and informative new DNA profiling system in the forensic field.

DNA, Mitochondrial↗

A rapid screening method to detect nonsense and frameshift mutations: identification of disease-causing APC alleles.

A functional screen for nonsense and frameshift mutations has been devised that allows genes of interest to be scanned in segments. This assay is based on the cloning of these segments in-frame with a colorimetric marker gene (lacZ) followed by screening for the level of functional activity from the marker polypeptide (beta-galactosidase). Individuals at risk for any one of a number of genetic diseases, in particular familial adenomatous polyposis coli (APC), can be quickly screened for chain-terminating mutations introduced by stops and frameshifts. At present, scanning of the APC gene for mutation requires significant effort because it is a large gene and most APC mutations are unique. Therefore, this assay offers a powerful option for the diagnosis of this and other genetic diseases, as well as great potential for the development of a similar rapid screen to detect APC mutations in colorectal adenomas and carcinomas.

Alleles↗

The rapid screen of concussion: an evaluation of the non-word repetition test for use in mTBI research.

PRIMARY OBJECTIVES: (1) To investigate the Nonword Repetition test (NWR) as an index of sub-vocal rehearsal deficits after mild traumatic brain injury (mTBI); (2) to assess the reliability, validity and sensitivity of the NWR; and (3) to compare the NWR to more sensitive tests of verbal memory. RESEARCH DESIGN: An independent groups design. METHODS AND PROCEDURES: Study 1 administered the NWR to 46 mTBI and 61 uninjured controls with the Rapid Screen of Concussion (RSC). Study 2 compared mTBI, orthopaedic and uninjured participants on the NWR and the Hopkins Verbal Learning Test (HVLT-R). MAIN OUTCOMES AND RESULTS: The NWR did not improve the diagnostic accuracy of the RSC. However, it is reliable and indexes sub-vocal rehearsal speed. These findings provide evidence that although the current form of the NWR lacks sensitivity to the impact of mTBI, the development of a more sensitive test of sub-vocal rehearsal deficits following mTBI is warranted.

Adolescent↗

Sensitive streptavidin-biotin enzyme-linked immunosorbent assay for rapid screening of chloramphenicol residues in swine muscle tissue.

A sensitive, competitive enzyme-linked immunosorbent assay (ELISA) for chloramphenicol (CAP) in swine muscle tissue has been developed. The ELISA is based on an earlier procedure. To improve sensitivity, different optimization procedures were investigated. The introduction of a streptavidin-biotin system and the use of a coating antigen with a lower CAP incorporation resulted in the most sensitive ELISA: the CAP concentration giving 50% inhibition decreased from 125 ng/mL to 3.0 ng/mL. This ELISA procedure was applied for a rapid screening of CAP residues in swine muscle tissue. The tissues were extracted with demineralized water. A concentrated phosphate-buffered saline solution was added to the filtered aqueous extract and this sample solution was directly submitted to the ELISA procedure. The results were compared to values obtained by analysis of a corresponding blank. This blank was prepared by treating a part of the aqueous sample solution with an immobilized monoclonal antibody preparation. This treatment was necessary because aqueous extracts of different swine muscle tissues showed a high variation in dose-response curves, probably caused by the complexity and variability of the matrix. In spiked tissues, the presence of CAP at concentrations of 10 micrograms/kg and higher can be easily demonstrated.

Alkaline Phosphatase↗

Value of rapid screening for acetaminophen in all patients with intentional drug overdose.

We performed a prospective study to determine the incidence of missed, potentially toxic acetaminophen poisoning in all patients with any type of intentional drug ingestion presenting to two large, county hospital emergency departments. Of 486 patients with drug ingestion seen during a five-month period, 114 (23.5%) were suspected of having ingested acetaminophen. Of these, 71 (62.3%) had insignificant acetaminophen levels (false-positive history). Of the 365 patients who were not suspected of having acetaminophen ingestion, seven patients (1.9%) were found to have elevated levels on rapid screening (false-negative history). Of these, only one patient had a potentially hepatotoxic level. We conclude that the incidence of missed, potentially serious acetaminophen overdose is very low in our study population.

Acetaminophen↗

Relative DNA binding affinity of helix 3 homeodomain analogues, major groove binders, can be rapidly screened by displacement of prebound ethidium bromide. A comparative study.

The binding affinity for a 12-bp dsDNA of Antennapedia helix 3 analogues, major groove binders, has been measured by displacement of prebound ethidium bromide, a fluorescent displacement assay proposed for minor groove binders by Boger et al.(J. Am. Chem. Soc., 2000, 122, 6382-6394). Relative binding affinities determined by this method were compared to those obtained by gel mobility shift and footprinting assays for the 12-bp dsDNA and a 178-bp DNA fragment. The present work demonstrates that the fluorescence displacement assay is suitable for rapid screening of major groove binders, even though about 60 to 70% of the prebound ethidium bromide is displaced by these peptides. Total (100%) displacement of ethidium bromide was serendipitously achieved by addition in the peptide sequence, at the N-terminus, of a S-3-nitro-2-pyridinesulfenyl-N-acetyl-cysteine residue. S-3-nitro-2-pyridinesulfenylcysteine was shown to (i) bind to dsDNA with a micromolar affinity and (ii) direct within DNA grooves a peptide with no affinity for dsDNA.

Amino Acid Sequence↗

Rapid screening procedures for the hydrolysis products of chemical warfare agents using positive and negative ion liquid chromatography-mass spectrometry with atmospheric pressure chemical ionisation.

Qualitative screening procedures have been developed for the rapid detection and identification of the hydrolysis products of chemical warfare agents in aqueous samples and extracts, using liquid chromatography-mass spectrometry with positive and negative atmospheric pressure chemical ionisation (APCI). Previously reported screening procedures, which used positive APCI or electrospray ionisation (ESI), were modified by using LC conditions that allowed acquisition of positive and negative ion mass spectra. APCI was generally found to be more robust than ESI, probably due to variable adduct ion formation with ESI, depending on the condition of the sample and the system. Negative APCI provided selective detection of acidic analytes and allowed facile differentiation of alkyl alkylphosphonic acids from isomeric dialkyl alkylphosphonates. The combination of positive and negative APCI, using a C18 column and water-methanol mobile phase modified with ammonium formate, provides a rapid screening procedure for chemical warfare agent degradation products, with limits of detectability in the range 10-100 ng/ml. In the case of proficiency test samples, where analyte concentrations are in the range 1-10 ppm, introduction of the sample by infusion may provide an even faster preliminary screening procedure.

Atmospheric Pressure↗

A rapid screening method for acid-volatile sulfide in sediments.

A new, direct method for the determination of acid-volatile sulfide (AVS) in sediments is presented. The method utilizes the direct reaction of Clines reagent with small amounts of sediment followed by colorimetric determination of AVS and offers fast analysis times without the need for specialized glassware or equipment. A comparison between AVS measured by this method and that measured by a purge-and-trap AVS method shows a linear relationship (r = 0.986, p < 0.001), although the method underestimates the AVS concentration measured by the purge-and-trap method. Sample throughput by the direct AVS method is more than 10 times that of purge-and-trap methods. The applicability of the direct AVS method to the rapid screening of AVS concentrations in sediments and for evaluating sediment quality with respect to metal contamination is demonstrated.

Acids↗

Solid-phase microextraction and gas chromatography-mass spectrometry for the rapid screening of triazole residues in wine and strawberries.

A solid-phase microextraction gas chromatography-mass spectrometry method has been developed for the determination of triazole residues, such as triadimefon, propiconazole, myclobutanil and penconazole. The method has been successfully applied to the analysis of strawberries and wine samples. The procedure is solvent-free, simple and highly sensitive. Within-day and day-to-day RSDs ranged between 2-11% and 7-28%, respectively. Detection limits estimated at a signal-to-noise ratio of 3 ranged between 30 (propiconazole) and 100 ng/kg (triadimefon). Since the detection limits achieved by this method are well below the maximum residue levels for wine (or grapes) and strawberries recommended by the European legislation, it can be conveniently used as a low-cost rapid screening method for the contamination of the considered samples.

Fragaria↗

Rapid screening for bacteriuria with a laser nephelometer.

Detection of significant bacteriuria with a laser nephelometer was evaluated in this study and compared with the results obtained by the quantitative loop method. We screened 1002 urine specimens and 220 (21.95%) were found to be positive at greater than or equal to 10(5) colony-forming units (CFU)/mL of urine by the standard method. Of the 220 positive specimens, 210 (95.4%) were detected in 6 h or less and 177 (80.4%) were detected within 3 h. The false-positive rate was 2.3% at 3 h and 19.7% at 6 h. These findings suggest that a 6-h procedure is necessary to detect 95% or more of significant bacteriuria. Laser nephelometer is versatile and can be used for rapid screening of bacteriuria.

Bacteria↗