Comparative analysis of a liquid-based Pap test and concurrent HPV DNA assay of residual samples. A study of 608 cases

OBJECTIVE: To retrospectively study the HPV DNA assay of residual samples from the ThinPrep Pap Test (Cytyc Corporation, Boxborough, Massachusetts, U.S.A.) PreserveCyt (Cytyc) vial as a quality improvement (QI) indicator for management of patients with abnormal cervical cytology. STUDY DESIGN: Six hundred eight residual sample vials of liquid-based Pap-Test specimens were selected for the study based on Pap-test results from October 1998 to March 2001. The specimen vials were forwarded to the reference laboratory (American Medical Laboratories, Chantilly, Virginia, U.S.A.) for HPV DNA assay using the Hybrid Capture System method (Digene Corporation, Gaithersburg, Maryland, U.S.A.). At the time of HPV DNA assay, the residual samples were between 8 days to 10 months old, and each vial contained 4 mL. Of the 608 study cases, 76 were WNL, 115 contained BCC, 172 contained ASCUS, 179 were LSIL and 66 were HSIL. In this study, the 191 WNL and BCC cases were designated as the disease-free control group. The HPV DNA typing results were reported as low-risk, high/intermediate-risk or HPV DNA "not detected" HPV types. The HPV DNA testing results were compared to the Pap-Test diagnoses and statistical analysis performed. RESULTS: The following information reflects the percentage of HPV DNA-positive cases based on the Pap-Test diagnoses: 16.2% in WNL and BCC, 51.1% in ASCUS, 94.4% in LSIL and 98.4% in HSIL. Sensitivity (95.5%), specificity (83.7%), false negative value (4.4%), false positive value (16.2%) and predictive value of a positive (88.3%) and negative (93.5%) Pap-Test were calculated on the basis of HPV DNA testing results for 436 cases that were diagnosed as either SIL or negative (WNL and BCC). ASCUS (172) Pap-Test cases were considered borderline--disease positive and excluded from statistical analysis. CONCLUSION: The HPV DNA assay of residual samples from ThinPrep Pap-Test liquid-based specimens is an objective adjunct to the gynecologic cytology QI protocol and is the gold standard reference test for triaging women with equivocal cytologic diagnoses. The great value of HPV DNA testing is its high sensitivity (95.5%), specificity (83.7%) and negative predictive value (93.5%). HPV DNA testing results can be used as a tool to better determine the need for referrals for colposcopic biopsy, especially for patients with an ASCUS diagnosis. The residual Pap-Test specimens are stable and reproducible for HPV DNA typing. A working flow chart for our gynecologic cytology QI program was produced from the Pap-Test and HPV DNA assay results. This offer presents the added benefit of minimizing the problem of sample variation. The prevalence of HPV infection was 16.2% in this study.     

Stenurus globicephalae Baylis et Daubney, 1925 (Nematoda: Pseudaliidae) from a false killer whale,Pseudorca crassidens (Cetacea: Delphinidae), stranded on the coast of Uruguay

Stenurus globicephalae Baylis et Daubney, 1925 (Nematoda: Pseudaliidae) was found in the cranial air sinuses of a false killer whale, Pseudorca crassidens (Owen), stranded on the coast of Uruguay in 1999. Although this species has been reported once in P. crassidens from the North Atlantic, this is the first record for South America. A total of 920 specimens were obtained, of which 663 were females (body length: 4.34 +/- 0.45 cm) and 257 were males (2.99 +/- 0.18 cm). Morphometric details are presented for S. globicephalae in this host, which do not show significant differences from those parasitizing Globicephala melas (Traill), but are distinct from those parasitizing Peponocephala electra (Gray). The host's skull revealed loss of osseous mass with the disappearance of the left zygomatic arch, and the left jaw had three osseous fenestrations in the region related to the organ of acoustic reception. These lesions support the hypothesis that this infection, known as stenurosis, was related to the stranding.     

Effect of some biocides on non-tuberculous mycobacteria

The incidence of disease and nosocomial infections produced by non tuberculosis mycobacteria (NTM) has increased in immunocompetent patients, but also and more frequently, in immunosuppressed patients. Several studies have disclosed that mycobacteria are more resistant to biocides than non-sporulating bacteria; in addition, some species are particularly resistant. The biocide action of sodium hypochloride and glutaraldehyde on Mycobacterium aviumintracellulare, Mycobacterium gordonae, Mycobacterium fortuitum and Mycobacterium chelonae was studied, using a modified Kelsey Maurer test. For the different species, both the minimal inhibitory concentration (MIC) and the minimal action time were determined. Effectiveness of sodium hypochloride and glutaraldehyde against the different mycobacterial species varied. The same was true for different isolates of the same species. Sodium hypochloride effective MIC and exposure time (killing of 99.99% of all NTM) were 0.2% and 5 minutes, respectively. In order to achieve 100% killing, 0.5% MIC and 15 minute exposure were needed. In the case of glutaraldehyde, 99.99% of the bacteria were killed with 1% MIC and a 15 minute exposure. An effectiveness of 100% was achieved with a 2% MIC of glutaraldehyde and a 15 minute exposure. Sodium hypochloride and glutaraldehyde are effective biocides for mycobacteria. The first biocide is cheap and effective at low concentrations, but its corrosive and oxidant nature makes it impossible for use in hospitals or with laboratory equipment. Glutaraldehyde (neither corrosive nor oxidant) is a safe alternative for disinfection of this type of equipment. However, it is important to bear in mind that these pathogens may develop resistance to biocides.     

Slight influence of the estrous cycle stage on the mucosal and systemic specific antibody response induced after vaginal and intraperitoneal immunization with protoxin Cry1Ac from Bacillus thuringiensis in mice

Since the immune response appears to be variable according to the hormonal stage of the mammalian female, the aim of this study was to determine whether estrous cycle stage modifies the mucosal and systemic immune responses induced by intraperitoneal and vaginal immunization of mice with protoxin Cry1Ac. We tested the influence of three immunizations on the specific antibody response elicited at estrus and diestrus, that were the same estrous cycle stages at which the antigen was applied. Both intraperitoneal and vaginal immunization of mice with Cry1Ac either at estrus or diestrus induces specific antibody responses at serum, vagina and large intestine. The stage of the estrous cycle have little or non influence in the magnitude of the response induced, since only at serum the IgM was slightly higher at estrus than at diestrus by both routes. At the large intestine only the IgA response elicited via the intraperitoneal route changed, being higher at diestrus, whereas at the vagina IgA response induced did not change significantly due to the cycle stage. Present results suggest that Cry1Ac may be used as an antigen carrier as it can elicit antibody responses at systemic level and at several mucosal sites including the vagina that are not modified significantly throughout the reproductive cycle.     

Halocidin: a new antimicrobial peptide from hemocytes of the solitary tunicate,Halocynthia aurantium

From hemocytes of the tunicate Halocynthia aurantium we purified a new antimicrobial peptide named halocidin. The native peptide had a mass of 3443 Da and comprised two different subunits containing 18 amino acid residues (WLNALLHHGLNCAKGVLA) and 15 residues (ALLHHGLNCAKGVLA), which were linked covalently by a single cystine disulfide bond. Two different monomers were separately synthesized and used to make three additional isoforms (15 residue homodimer, 18 residue homodimer, heterodimer). In antimicrobial assays performed with synthetic peptides of halocidin, it was confirmed that congeners of the 18 residue monomer were more active than those of the 15 residue monomer against methicillin-resistant Staphylococcus aureus and multidrug-resistant Pseudomonas aeruginosa.     

Phosphorylation of cucumber mosaic virus RNA polymerase 2a protein inhibits formation of replicase complex

The 2a (polymerase) protein of cucumber mosaic virus (CMV) was shown to be phosphorylated both in vivo and in vitro. In vitro assays using 2a protein mutants and tobacco protein kinases showed that the 2a protein has at least three phosphorylation sites, one of which is located within the N-terminal 126 amino acid region. This region is essential and sufficient for interaction with the CMV 1a protein. When phosphorylated in vitro, the 2a protein N-terminal region failed to interact with the 1a protein. Since the 1a-2a interaction is essential for the replication of CMV, this suggests that phosphorylation of the N-terminal region of the 2a protein negatively modulates the interaction in vivo, and may have a regulatory role acting directly in viral infection.     

Green fluorescent protein-labeled recombinant fluobody for detecting the picloram herbicide

A green fluorescent protein-labeled fluobody was designed to develop a simple immunoassay method for detecting picloram herbicide in an environmental sample. The gfp gene was successfully inserted into the pSJF2 vector harboring the picloram-specific antibody fragment to yield pSJF2GFP. Picloram spiking in an environmental river sample could be indirectly detected by observing the fluorescence intensity value of the gfp-fluobody, exhibiting specific sensitivity to free picloram with an IC50 value of 50 ppb. Using the gfp-fluobody immunoassay avoids the enzyme-substrate reaction for calorimetric detection that is required in an enzyme-linked immunosorbent assay (ELISA).     

Selective action of acetogenin mitochondrial complex I inhibitors

Five annonaceous acetogenins, rolliniastatin-1 [structure: see text], rolliniastatin-2 [structure: see text], laherradurin [structure: see text], squamocin [structure: see text], annonacin [structure: see text], and rotenone as a reference, differing in their NADH oxidase inhibition activity, have been evaluated for antifeedant, insecticidal, trypanocidal and cytotoxic effects on insect, mammalian and tumor cells. All the test compounds were toxic to Leptinotarsa decemlineata, demonstrated selective cytotoxicity to insect Sf9 cells and a panel of tumor cell lines with the multidrug-resistant SW480 (P-glycoprotein+, Pgp+) being the most sensitive one. Compounds [structure: see text] and rotenone had post-ingestive effects against Spodoptera littoralis larvae while [structure: see text] and rotenone were active against Trypanosoma cruzi. Based on their biochemical properties (inhibition of the mitochondrial NADH oxidase activity), the in vivo effects of these compounds on S. littoralis and their cytotoxic effects on Sf9 and tumor cells were more predictable than their effect on T. cruzi and mammalian cells.     

J-binding protein increases the level and retention of the unusual base J in trypanosome DNA

The nuclear DNA of Trypanosoma brucei and other kinetoplastid flagellates contains the unusual base beta-d-glucosyl-hydroxymethyluracil, called J, replacing part of the thymine in repetitive sequences. We have described a 100 kDa protein that specifically binds to J in duplex DNA. We have now disrupted the genes for this J-binding protein (JBP) in T. brucei. The disruption does not affect growth, gene expression or the stability of some repetitive DNA sequences. Unexpectedly, however, the JBP KO trypanosomes contain only about 5% of the wild-type level of J in their DNA. Excess J, randomly introduced into T. brucei DNA by growing the cells in the presence of the J precursor 5-hydroxymethyldeoxyuridine, is lost by simple dilution as the KO trypanosomes multiply, showing that JBP does not protect J against removal. In contrast, cells containing JBP lose excess J only sluggishly. We conclude that JBP is able to activate the thymine modification enzymes to introduce additional J in regions of DNA already containing a basal level of J. We propose that JBP is a novel DNA modification maintenance protein.