Assessment of DNA Integrity From Trap-Captured Boll Weevil (Coleoptera: Curculionidae) for Use in a New PCR-Based Diagnostic Tool.

The boll weevil, Anthonomus grandis grandis Boheman (Coleoptera: Curculionidae), is a major pest of commercial cotton (Gossypium hirsutum) in the southern United States and throughout Central and South America. Efforts are underway to develop a PCR-based diagnostic tool that can be used to rapidly and accurately differentiate boll weevils from other weevil species that are commonly captured in pheromone traps. However, the quantity and integrity of weevil DNA must be sufficient for a successful PCR assay. Currently, active eradication programs service traps weekly, but post-eradication programs service traps at 2- or 3-wk intervals. Consequently, captured weevils may be dead, dismembered, and exposed to environmental conditions for prolonged periods which may adversely affect the quantity and quality of weevil DNA. We documented DNA quantity and integrity in boll weevils and weevil body parts aged in traps over a 3-wk period under field conditions. The quantity of DNA extracted from whole weevils, heads, abdomens, and legs generally remained sufficient (> 1 ng/µl) for successful PCR amplification throughout the 21-d period. The integrity (fragment length) of extracted DNA declined over time but generally was sufficient (> 700 bp) for successful amplification. PCR amplification of three marker genes validated that the quality and integrity of DNA extracted from dead weevils and individual weevil body parts aged in traps up to 21 d remained at sufficient levels for the PCR-based assay. However, our data also suggested that rain events may accelerate degradation of weevil DNA.

The possible cost effectiveness of peripheral blood stem cell mobilization with cyclophosphamide and the late addition of G-CSF.

The purpose of this study was to develop a cost-effective protocol for the mobilization of peripheral blood stem cells (PBSC) in patients with malignancy. Thirty consecutive patients were randomized to mobilize PBSC with the late addition of a standard 250 microg dose of G-CSF (Neutrogen) from day 8 or early addition of the same dose of G-CSF from day 2, following cyclophosphamide (CY) 4 g/m2. The median yield of CD34+ cells from evaluated patients was 7.87 x 10(6)/kg (range, 2.06-27.25), collected in a median of four apheresis (range, 2-9). Target CD34 + cell doses > or = 2.0 x 10(6)/kg were achieved in all patients able to be evaluated. There were no statistically significant differences in CD34+ cell yields or toxicities. Overall engraftment occurred with median days to neutrophils > or = 0.5 x 10(9)/L or platelets > 20 x 10(9)/L of 11 and 17 days, respectively. However, the duration of G-CSF administration was markedly shorter in the late use of G-CSF group than in the early use of G-CSF group, with a median of 9 days compared with 15 days (p<0.001). PBSC harvesting after priming with CY plus delayed use of G-CSF made it a safe and cost-effective procedure.

Assessment of biomarkers in paired primary and recurrent colorectal adenocarcinomas.

PURPOSE: Recurrent colorectal cancers respond poorly to anticancer treatment including radiotherapy. To better understand the biological characteristics of the recurrent colorectal tumor, we investigated various biomarkers regulating cell proliferation and cell loss in paired primary and recurrent colorectal tumor specimens within each individual. METHODS AND MATERIALS: From a total of 11 colorectal adenocarcinoma patients, 22 specimens of paired primary and recurrent tumors were obtained for analysis. Apoptosis was evaluated by TUNEL labeling of apoptotic DNA fragmentation. Other biomarkers including proliferating cell nuclear antigen (PCNA), p53, WAF1, p34cdc2, and cyclins B1 and D1 were analyzed by immunohistochemical stains. RESULTS: PCNA index (PCNAI) showed an increase in 6 and a decrease in 5 recurrent tumors compared to primary tumors. Median PCNAI in primary and recurrent tumors were 33.5 and 48.3, respectively (p = 0.16). In contrast, the apoptotic index (AI) decreased in 9 of 11 recurrent tumors compared to primary tumors. Median AI decreased from 4.3 in primary tumors to 1.4 in recurrent tumors (p = 0.04). The p53 expression increased in more than half of recurrent tumors compared to primary tumors. Mean staining score increased from 0.7 in primary tumors to 1.2 in recurrent tumors (p = 0.059). WAF1 and cyclin B1 did not show significant change. In contrast, both cyclin D1 and p34cdc2 increased significantly in recurrent tumors. These two biomarkers showed increased expression in 8 (cyclin D1) and 7 (p34cdc2) recurrent tumors, respectively, compared to their primary counterparts. Mean staining scores of both biomarkers in recurrent tumors increased by more than twofold compared to those in primary tumors and these differences were statistically significant (cyclin D1, p = 0.007; p34cdc2, p = 0.008). CONCLUSION: This study showed significantly decreased apoptosis in recurrent colorectal tumors compared to their primary counterparts. The underlying regulatory mechanisms included increased expression of p53 and altered cell cycle regulators such as increased cyclin D1 and p34cdc2. With further study, it may be used for developing a new therapeutic strategy for the treatment of recurrent colorectal cancer.