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At least 127 records · Page 7Linked to original sources
PubMed2001

Optimization of random amplification of polymorphic DNA analysis for molecular subtyping of Escherichia coli O157.

Random amplification of polymorphic DNA (RAPD) analysis using the polymerase chain reaction has proved to be a useful technique in the epidemiological investigation of micro-organisms but may suffer from a lack of reproducibility in poorly optimized protocols. In this study a method of obtaining reproducible genomic fingerprints using RAPD analysis of Escherichia coli O157 is described. By systematic optimization of reaction conditions and selection of suitable primers, reproducible and discriminatory profiles could be obtained from all E. coli O157 strains tested. In addition, two other methods of obtaining reproducible profiles from E. coli O157 strains without the need to purify genomic DNA are described.

Buffers↗
PubMed1997

Surveillance for Escherichia coli O157:H7 infections in Minnesota by molecular subtyping.

BACKGROUND: Escherichia coli O157:H7 is a leading cause of diarrhea and the hemolytic-uremic syndrome. Current public health surveillance for E. coli O157:H7 requires considerable resources; traditional methods lack the sensitivity and specificity to detect outbreaks effectively. METHODS: During 1994 and 1995, the Minnesota Department of Health requested that all clinical isolates of E. coli O157:H7 be submitted to our laboratory. Isolates were subtyped by pulsed-field gel electrophoresis (PFGE), and patients were interviewed about potential sources of infection. RESULTS: In 1994 and 1995, 344 cases of E. coli O157:H7 infection were reported to the Minnesota Department of Health; 317 (92 percent) were subtyped by PFGE, and 143 distinct PFGE patterns were identified. Ten outbreaks of E. coli O157:H7 were identified; these accounted for 56 (18 percent) of the 317 subtyped cases. Four outbreaks were detected solely as a result of subtype-specific surveillance. In 11 two-week periods, the number of reported cases of E. coli O157:H7 doubled from the previous two weeks. In eight of these instances, the patterns identified were dissimilar and there were no outbreaks. Two of the remaining three increases resulted from multiple simultaneous outbreaks. CONCLUSIONS: Subtype-specific surveillance for E. coli O157:H7 can identify outbreaks that are not detected by traditional methods and can ascertain whether sudden increases in reported cases are due to sporadic isolated cases or to one or more outbreaks.

Adolescent↗
PubMed2025

Development and validation of a novel risk stratification signature derived from migrasome and tumor microenvironment-related genes for molecular subtyping and improving clinical outcomes in head and neck squamous cell carcinoma.

BACKGROUND: The tumor microenvironment (TME) and migrasomes released by tumor cells significantly influence carcinogenesis and immune evasion. However, our understanding of the prognostic and therapeutic implications of migrasome and tumor microenvironment-related genes (mtmRGs) in head and neck squamous cell carcinoma (HNSCC) remains limited. METHODS: We explored the relationship between mtmRGs and HNSCC prognosis by utilizing The Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO) databases. Subsequently, we developed an innovative prognostic signature, and assessed its prognostic significance using the Kaplan-Meier method, time-dependent receiver operating characteristic (ROC), and Cox regression analyses. To explore the underlying mechanisms, we conducted gene set variation analysis (GSVA), gene set enrichment analysis (GESA), and immune infiltration analysis. A nomogram was developed to estimate the overall survival (OS) rates for HNSCC patients. Lastly, we chose P4HA1, which was part of the signature, for additional experimental validation in vitro and in vivo. RESULTS: The mtmRGs signature effectively classifies HNSCC patients into two distinct risk subgroups, with the high-risk cohort demonstrating significantly poorer OS. The risk score serves as an independent prognostic factor for HNSCC patients; those with lower risk scores are more likely to exhibit favorable responses to immunotherapy, particularly with CTLA4 inhibitors. Furthermore, a lower risk score is significantly correlated with the sensitivity of HNSCC patients to cyclophosphamide, gemcitabine, and axitinib. CONCLUSION: This study presents an innovative gene signature associated with mtmRGs, which may be utilized both for predicting survival and directing personalized chemotherapy and immunotherapy regiments for patients with HNSCC.

Humans↗
PubMed2026

Circadian-rhythm-based dynamics of the secretome in molecular subtypes of pancreatic ductal adenocarcinoma.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) lacks a reliable diagnostic biomarker, largely due to its asymptomatic onset and frequent late-stage detection, resulting in a poor 5-year survival rate. Identifying biomarkers for timely diagnosis is critical. Cancer cells display distinct protein regulation changes that drive disease hallmarks, and characterizing these across PDAC subtypes may provide insights into disease progression. RESEARCH DESIGN AND METHODS: In this study, a label-free quantitative (LFQ) proteomics approach using mass spectrometry (MS) was employed to profile circadian rhythm-regulated proteins in the secretome of PDAC cell lines. RESULTS: LFQ analysis revealed rhythmic protein regulation patterns, reflecting temporal control of biological processes in PDAC. Upregulated pathways included signal transduction, glycolysis, angiogenesis, and protein synthesis, indicating enhanced metabolic and proliferative activity. Downregulated immune pathways suggested potential immune modulation. Comparative analysis revealed subtype-specific patterns: the quasi-mesenchymal subtype exhibited higher levels of metabolic and extracellular matrix (ECM) remodeling proteins, while the classical subtype showed higher levels of ECM-degrading proteins, consistent with known phenotypic differences. CONCLUSION: These findings highlight rhythmically regulated proteins as potential subtype-specific markers in PDAC and provide a basis for future validation studies. Mass spectrometry proteomics data are available via the ProteomeXchange Consortium (PRIDE: PXD054693).

Humans↗
PubMed1996

Molecular subtypes and antifungal susceptibilities of serial Cryptococcus neoformans isolates in human immunodeficiency virus-associated Cryptococcosis. Cryptococcal Disease Active Surveillance Group.

Serial isolates of Cryptococcus neoformans from 33 human immunodeficiency virus-infected patients with cryptococcosis were analyzed to determine whether persistence might result from reinfection with a new cryptococcal strain or acquisition of antifungal resistance. Isolates were subtyped by multilocus enzyme electrophoresis (MEE), electrophoretic karyotyping (EK), random-amplified polymorphic DNA (RAPD), and the CNRE-1 DNA probe, MICs of amphotericin B, fluconazole, and 5-fluorocytosine were determined. No changes in MEE or RAPD subtypes were detected in serial isolates from any patient. Isolates from 8 patients (24%) showed alterations in EK only (mobility change in two or more bands) but not with any other subtyping method. MICs did not change significantly in isolates from 30 patients. In 1 case, the fluconazole MIC increased stepwise over 18 months, suggesting development of resistance. These overall invariant subtyping and MIC results confirm previous studies suggesting that persistent cryptococcal infection is due to relapse rather than reinfection or antifungal drug resistance.

AIDS-Related Opportunistic Infections↗
PubMed2002

Molecular subtype analyses of Campylobacter spp. from Arkansas and California poultry operations.

Campylobacter isolates from diverse samples within broiler production and processing environments were typed by using flaA short variable region DNA sequence analysis. Sixteen flocks from four different farms representing two broiler producers in Arkansas and California were analyzed. Fourteen of the flocks (87.5%) were Campylobacter-positive; two remained negative throughout the 6-week rearing period. In general, multiple clones were present within a flock. Additionally, clones found within a flock were also present on the final product, although the diversity of Campylobacter spp. on the final product appeared to be reduced relative to that observed within the flock. Comparison of clones between flocks on the same farm revealed that some clones of Campylobacter persisted in multiple flocks. Furthermore, some clones were identified across the two farms that were under the same management. In two sampling periods, environmental isolates were positive for Campylobacter prior to flock shedding. Environmental samples associated with five additional flocks were positive for Campylobacter concomitantly with recovery of Campylobacter from the birds. Analysis of the environmental isolates that were positive prior to flock shedding demonstrated that in some instances the environmental isolates possessed genotypes identical to those of isolates originating from the flock, while in other cases the environmental isolates possessed genotypes that were distantly related to isolates obtained from the flock. Analyses of environmental isolates that tested positive concurrently with the positive isolates from the flocks demonstrated varied results; in some instances the environmental isolates possessed genotypes identical to those of isolates originating from the flock, while in other cases the environmental isolates possessed genotypes that were distantly related to isolates obtained from the flock. These data suggest that the external environment may contribute to Campylobacter contamination during poultry production and processing. However, environmental contamination with Campylobacter does not appear to be the sole contributing factor.

Animals↗
PubMed1994

Molecular subtyping of Salmonella enteritidis phage type 8 strains from the United States.

Salmonella enteritidis is now the most common serotype of the genus Salmonella reported in the United States. Bacteriophage typing has been helpful for subdividing S. enteritidis strains from different sources in the United States. Most S. enteritidis outbreaks reported were egg related, and the majority of them were caused by strains of phage type 8. To determine whether restriction fragment length polymorphism of the rRNA genes (ribotyping) and of the genomic DNAs from two lysogenic phages from S. enteritidis could be used to discriminate between S. enteritidis phage type 8 strains, we conducted Southern hybridization studies on 24 isolates from different outbreaks and six non-outbreak-associated strains using DNA probes for 16S and 23S rRNA genes and S. enteritidis typing phages 1 and 2 from the Ward typing system (L. R. Ward, J. D. H. de Sa, and B. Rowe, Epidemiol. Infect. 99:291-294, 1987). Of seven restriction endonucleases screened with the probe for rRNA genes, AccI provided the best discrimination between strains; six distinct patterns were observed. AccI ribosomal DNA patterns 1 to 6 were detected among 76.7, 3.3, 6.7, 3.3, 3.3, and 6.7% of isolates tested, respectively. Strains of AccI ribosomal DNA pattern 3 could be further subdivided into two additional patterns by using SmaI. Epidemiologically related strains had identical patterns. No discrimination between strains was achieved by probes for phages 1 and 2. No sequences homologous to the phage I probe were detected among phage type 8 strains, and all strains tested with six restriction enzymes had the same hybridization pattern with the phage 2 probe. These findings demonstrate that ribotyping with AccI and SmaI provides an additional means of discriminating between some phage type 8 strains; however, ribotyping and the phage 2 hybridization results from egg-related outbreak strains support previous findings that these strains are closely related.

Bacterial Typing Techniques↗
PubMed1995

Comparison of multilocus enzyme electrophoresis and random amplified polymorphic DNA analysis for molecular subtyping of Cryptococcus neoformans. The Cryplococcal Disease Active Surveillance Group.

We evaluated multilocus enzyme electrophoresis (MEE) and random amplified polymorphic DNA (RAPD) for their usefulness in subtyping 344 Cryptococcus neoformans clinical isolates obtained from four U.S. metropolitan areas in 1992 to 1994. MEE and RAPD with five primers both discriminated between the two varieties of C. neofromans. MEE divided C. neoformans var. neoformans isolates into 15 enzyme electrophoretic subtypes (ETs) arranged in three complexes. The predominant ET 1 complex contained 10 ETs, with isolates from 70% of patients in 1 ET. RAPD with five primers further sorted this predominant ET into 19 subtypes, with 60% of isolates sorting into three RAPD types. The ET 8 MEE complex, containing three ETs, could not be divided further by RAPD. The ET 7 complex (two ETs) included isolates from all serotype AD patients. Although both MEE and RAPD identified isolates of C. neoformans var. gattii, neither distinguished between serotypes B and C. These results showed that the two C. neoformans varieties could be identified by MEE or RAPD profile as well as by biochemical methods. RAPD improved the discriminatory power of MEE for isolates within the ET 1 complex but with other ETs offered little additional sensitivity over MEE and was less sensitive than MEE with isolates of C. neoformans var. gattii. This information will be useful in identifying particular environmental sources of disease-causing exposures, in seeking clusters of cases, and in determining whether an infecting strain changes over time.

Cryptococcus neoformans↗
PubMed2001

Correlations between molecular subtyping and serotyping of Listeria monocytogenes.

To define relationships between Listeria monocytogenes genetic lineages, ribotypes, and serotypes, 235 L. monocytogenes isolates were characterized by serotyping and automated EcoRI ribotyping. Genetic lineage predicted the following serovar clusters: lineage I, comprising serotypes 1/2b, 3b, 3c, and 4b; lineage II, comprising serotypes 1/2a, 1/2c, and 3a; and lineage III, comprising serotypes 4a and 4c. Some EcoRI ribotypes contained multiple serotypes; a subset of these isolates was further differentiated with PvuII ribotyping. Of the 12 resultant EcoRI-PvuII combination types, only 4 contained multiple serotypes, demonstrating the potential of ribotyping for serotype prediction.

Animals↗
PubMed2003

Molecular subtyping of Vibrio cholerae O1 and O139 by pulsed-field gel electrophoresis in Hong Kong: correlation with epidemiological events from 1994 to 2002.

Two hundred twenty isolates of Vibrio cholerae O1 and O139 collected from 1994 to 2002 in Hong Kong were analyzed by pulsed-field gel electrophoresis (PFGE). Chromosomal DNAs from all V. cholerae isolates in agarose plugs were digested with the restriction enzyme NotI, resulting in 20 to 27 bands. Sixty distinctive PFGE patterns in the range of 10 to 300 kb were noted among 213 isolates typeable by PFGE. By comparing the common PFGE patterns obtained from four well-defined outbreaks of V. cholerae O1 and O139 with those obtained from other, epidemiologically unrelated isolates during the study period, indistinguishable and similar PFGE patterns were identified, indicating their close relatedness, in agreement with the results of epidemiological investigations. Heterogeneous PFGE patterns (with four to six banding differences), however, were identified among strains that were imported from other parts of Asia, including Indonesia, India, and Pakistan. Correlations with epidemiological information further support the usefulness of PFGE as an epidemiological tool in laboratory investigations of suspected outbreaks. Standardization of PFGE methodology will allow international comparison of fingerprint patterns and will form the basis of a laboratory network for tracking V. cholerae.

Bacterial Typing Techniques↗
PubMed2005

DNA polymorphism and molecular subtyping of the capsular gene cluster of group B streptococcus.

Serotyping and other phenotypic methods are often used to characterize the capsular polysaccharide of group B streptococci (GBS). We describe a capsular genotyping method that utilizes PCR of capsular polysaccharide synthesis genes (cps) and restriction enzyme digestion. This method facilitates the detection of DNA polymorphism in cps genes and correlates well with serotyping.

Bacterial Capsules↗
PubMed2006

Molecular subtyping of poultry-associated type A Clostridium perfringens isolates by repetitive-element PCR.

Clostridium perfringens strains (type A) isolated from an integrated poultry operation were subtyped using repetitive-element PCR with Dt primers. Isolates were obtained from fecal, egg shell, fluff, and carcass rinse samples as part of a previously reported temporally linked epidemiological survey. A total of 48 isolates of C. perfringens were obtained from different stages of the broiler chicken production chain from two separate breeder farms that supplied a single hatchery that in turn provided chicks to a single grow-out farm whose flocks were processed at a single plant. All 48 isolates were typeable (100% typeability) by repetitive-element PCR with Dt primers. This subtyping method was highly reproducible and discriminatory. By repetitive-element PCR with Dt primers, isolates were classified into four major branches with 12 subgroups or clades. The Simpson's index of discrimination was calculated to be 0.96 for groupings of >95% correlation. Toxin gene profiles of the isolates indicated that all of the isolates were C. perfringens alpha-toxin gene positive and 46 of 48 isolates were beta2-toxin gene positive. All strains were negative for beta- and epsilon-toxin genes. Repetitive sequence-based PCR was found to be a technically practical and reproducible means of subtyping C. perfringens libraries from specific epidemiological or production environment settings.

Animals↗
PubMed2006

Molecular subtypes of breast cancer in relation to paclitaxel response and outcomes in women with metastatic disease: results from CALGB 9342.

INTRODUCTION: The response to paclitaxel varies widely in metastatic breast cancer. We analyzed data from CALGB 9342, which tested three doses of paclitaxel in women with advanced disease, to determine whether response and outcomes differed according to HER2, hormone receptor, and p53 status. METHODS: Among 474 women randomly assigned to paclitaxel at a dose of 175, 210, or 250 mg/m2, adequate primary tumor tissue was available from 175. Immunohistochemistry with two antibodies and fluorescence in situ hybridization were performed to evaluate HER2 status; p53 status was determined by immunohistochemistry and sequencing. Hormone receptor status was obtained from pathology reports. RESULTS: Objective response rate was not associated with HER2 or p53 status. There was a trend toward a shorter median time to treatment failure among women with HER2-positive tumors (2.3 versus 4.2 months; P = 0.067). HER2 status was not related to overall survival (OS). Hormone receptor expression was not associated with differences in response but was associated with longer OS (P = 0.003). In contrast, women with p53 over-expression had significantly shorter OS than those without p53 over-expression (11.5 versus 14.4 months; P = 0.002). In addition, triple negative tumors were more frequent in African-American than in Caucasian patients, and were associated with a significant reduction in OS (8.7 versus 12.9 months; P = 0.008). CONCLUSION: None of the biomarkers was predictive of treatment response in women with metastatic breast cancer; however, survival differed according to hormone receptor and p53 status. Triple negative tumors were more frequent in African-American patients and were associated with a shorter survival.

Black or African American↗