METHODOLOGY: One hundred and twenty clinical isolates of S. pneumoniae were obtained from patients of University Malaya Medical Centre (UMMC). The strains were screened using a multiplex real-time PCR method for the presence of alterations in the genes encoding the penicillin binding proteins: pbp2b, macrolide resistance determinant ermB and the pneumolysin gene, ply. Dual-labelled Taqman probes were used in the real-time detection method comprising three different genes labeled with individual fluorophores at different wavelengths. One hundred and twenty isolates from bacterial cultures and isolates directly from blood cultures samples were analyzed using this assay.
RESULTS: A multiplex PCR comprising the antibiotic resistance genes, ermB and and pneumolysin gene (ply), a S. pneumoniae species specific gene, was developed to characterize strains of S. pneumoniae. Out of the 120 pneumococcal isolates, 58 strains were categorized as Penicillin Sensitive Streptococcus pneumoniae (PSSP), 36 as Penicillin Intermediate Streptococcus pneumoniae (PISP) and 26 as Penicillin Resistant Streptococcus pneumoniae (PRSP). All the 58 PSSP strains harboured the pbp2b gene while the 36 PISP and 26 PRSP strains did not harbour this gene, thus suggesting reduced susceptibility to penicillin. Resistance to erythromycin was observed in 47 of the pneumococcal strains while 15 and 58 were intermediate and sensitive to this drug respectively. Susceptibility testing to other beta-lactams (CTX and CRO) also showed reduced susceptibility among the strains within the PISP and PRSP groups but most PSSP strains were sensitive to other antibiotics.
CONCLUSION: The characterization of pneumococcal isolates for penicillin and erythromycin resistance genes could be useful to predict the susceptibility of these isolates to other antibiotics, especially beta-lactams drugs. We have developed an assay with a shorter turnaround time to determine the species and resistance profile of Streptococcus pneumoniae with respect to penicillin and macrolides using the Real Time PCR format with fluorescent labeled Taqman probes, hence facilitating earlier and more definitive antimicrobial therapy which may lead to better patient management.
STUDY DESIGN: Eighty-six postnasal biopsy samples and 71 fine-needle aspirate samples of neck masses were obtained from patients who were clinically suspect for NPC. Genomic DNA was extracted from the samples, and EBNA1, EBNA2, and LMP genes of EBV were detected by PCR. PCR results were compared with NPC histopathology findings.
RESULTS: The sensitivity of PCR to detect EBNA1 (97.14%), EBNA2 (88.57%), and LMP (91.43%) genes of EBV in nasopharyngeal biopsy samples were higher than those in fine-needle aspirate samples.
CONCLUSION: Detection of EBV by PCR in tissue obtained from nasopharyngeal biopsy and fine-needle aspirate samples of neck masses is a relatively inexpensive, reliable, and accurate method of diagnosing NPC. Detection of EBV genes is on par with histopathological examination (HPE) and superior to fine-needle aspirate cytology.
SIGNIFICANCE: PCR is an ideal tool for suggesting NPC and guiding the diagnostic workup in occult primary tumors, facilitating earlier diagnosis and reducing morbidity and mortality.
MATERIALS AND METHOD: Eighty-seven paraffin-embedded ameloblastoma cases (20 unicystic, 47 solid/multicystic, 3 desmoplastic and 17 recurrent) were subjected to immunohistochemistry for expression of cortactin, N-WASP, WIP, Src kinase and F-actin, and findings correlated with clinicopathological parameters.
RESULTS: Invadopodia proteins (except Src kinase) and F-actin were widely detected in ameloblastoma (cortactin: n = 73/87, 83.9%; N-WASP: n = 59/87; 67.8%; WIP: n = 77/87; 88.5%; and F-actin: n = 87/87, 100%). Protein localization was mainly cytoplasmic and/or membranous, and occasionally nuclear for F-actin. Cortactin, which functions as an actin-scaffolding protein, demonstrated significantly higher expression levels within ameloblastoma tumoral epithelium than in stroma (P < 0.05). N-WASP, which coordinates actin polymerization and invadopodia-mediated extracellular matrix degradation, was overexpressed in the solid/multicystic subtype (P < 0.05). WIP, an upstream regulator of N-WASP, and F-actin were significantly upregulated along the tumour invasive front compared to tumour centres (P < 0.05). Except for males with cortactin overexpression, other clinical parameters (age, ethnicity and anatomical site) showed no significant correlations.
CONCLUSIONS: Present results suggest that local invasiveness of ameloblastoma is dependent upon the migratory potential of its tumour cells as defined by their distribution of cortactin, N-WASP and WIP in correlation with F-actin cytoskeletal dynamics.
MATERIALS AND METHODS: A total of 17 patients were selected fulfilling one of the Bethesda criteria: colorectal cancer diagnosed in a patient aged less than 50 years old, having synchronous and metachronous colorectal cancer or with a strong family history. Immunohistochemical staining was performed on paraffin embedded tumour tissue samples using four antibodies: MLH1, MSH2, MSH6 and PMS2.
RESULTS: Twelve out of 17 patients (70.6%) were noted to have a family history. A total of 41% (n=7) of the patients had abnormal immunohistochemical staining with one or more of the four antibodies. Loss of expression were noted in 13 tumour tissues with a negative staining score <4. Of 13 tumour tissues, four showed loss expression of MLH1. For PMS2, loss of expression were noted in five cases. Both MSH2 and MSH6 showed loss of expression in two tumour tissues respectively.
CONCLUSIONS: Revised Bethesda criteria and immunohistochemical analysis constituted a convenient approach and is recommended to be a first-line screening for Lynch syndrome in Malay cohorts.