Factors related to inter-observer reproducibility of conventional Pap smear cytology: a multilevel analysis of smear and laboratory characteristics.

OBJECTIVE: To identify factors that influence the inter-observer reproducibility of the routine, conventional Pap smear cytology (Pap smear test) in a network of certificated laboratories in a middle-income Latin American country. METHODS: Twenty-six laboratories provided each an average of 26 negative for malignancy (NILM) and high-grade squamous intraepithelial lesion (HSIL) Pap smears. An external panel reviewed the slides. The kappa index and multilevel logistic regression were used to estimate the reproducibility and odds ratios (OR) of a false result with 95% confidence intervals (95% CI), respectively. Results are presented for laboratories that collect (collector laboratories) and do not collect (non-collector laboratories) samples. RESULTS: The agreements ranged widely (median kappa 0.51, range 0.16-0.70). The overall false-positive (FP) and false-negative (FN) rates were 31% (95% CI 27-35) and 11% (95% CI 7-17). Among collector laboratories (N = 14), a bigger sample collection volume decreased the probability of a FP (OR-adjusted 0.05, 95% CI 0.02-0.1) whereas the number of quality defects (OR-adjusted 1.67, 95% CI 1.25-2.24), high workload (OR-adjusted 5.52, 95% CI 3.85-7.92) and collection by cytotechnologists (OR-adjusted 1.28, 95% CI 1.15-1.42) or health professionals (OR-adjusted 2.26, 95% CI 2.04-2.49) instead of nursing assistants increased it. Among non-collector laboratories (N = 9), the FP rate increased with the number of quality defects (OR-adjusted 1.86, 95% CI 1.06-3.26) but decreased if the samples were collected by health professionals instead of nursing assistants (OR-adjusted 0.37, 95%CI 0.17-0.80). No significant associations were observed for FN. CONCLUSIONS: Staff in charge of cervical sampling significantly determined the reproducibility of the Pap smear test, but this depended on whether the laboratory collects samples or read samples collected elsewhere.

Knowledge of Pap screening and human papillomavirus among women attending clinics in Medellín, Colombia.

This study evaluated Pap screening and human papillomavirus (HPV) knowledge in a population of Colombian women as a possible contributing factor of low cervical cancer screening success. This is a descriptive, cross-sectional analysis of 454 women who were approached in five different hospitals and clinics throughout Medellín, Colombia. Of them, 449 females agreed to participate and answered a standardized face-to-face questionnaire regarding Pap screening and HPV knowledge. Using logistic regression, predictors of both Pap and HPV knowledge were examined. Overall, 76.3% of the participants exhibited a high level of Pap screening knowledge, while only 7.8% showed high level of HPV knowledge. Of the 449 women, 71.5% reported that it had been 1 year or less since their last Pap test, while 7.8% reported never having had a Pap test or not having had a recent test. Factors influencing Pap screening knowledge included education level and insurance; factors influencing HPV knowledge included education level and age. The high level of Pap screening knowledge and use do not explain the high cervical cancer rates in Colombia. The results of this study suggest that educational efforts should be focused on increasing women's knowledge and awareness of HPV in anticipation of the availability of HPV vaccines and HPV tests for screening.

Phylogenetically related, clinically different: human papillomaviruses 6 and 11 variants distribution in genital warts and in laryngeal papillomatosis.

Genital warts (GWs) and laryngeal papillomatosis (LP) are two usually benign pathologies related to infection with human papillomaviruses (HPVs), mainly HPV6 and HPV11. The aim of this work was to describe the genetic diversity of HPV6 and HPV11 isolates found in GWs and LPs, and to analyse the differential involvement of viral variants in either lesion. A total of 231 samples diagnosed as GWs (n = 198) or LP (n = 33) and caused by HPV6 or HPV11 monoinfections were analysed. The phylogenetic relationships of the retrieved viral sequences were explored. We have identified the long control region and the intergenic E2-L2 region as the two most variable regions in both HPV6 and HPV11 genomes. We have generated new HPV6 (n = 166) or HPV11 (n = 65) partial sequences from GWs and LPs lesions spanning both regions and studied them in the context of all available sequences of both types (final n = 412). Our results show a significant (p <0.01) differential presence of HPV6 variants among both pathologies, with HPV6 B variants being preferentially found in GW versus LP samples. No differential involvement of HPV11 variants was observed. Our findings suggest that different HPV6 variants may either show differential tropism or have different potential to induce lesions in different epithelia.

Plasmodium yoelii: cloning and characterization of the gene encoding for the mitochondrial heat shock protein 60.

Heat shock proteins are a highly conserved group of proteins required for the correct folding, transport, and degradation of other proteins in vivo. The Hsp70, Hsp90, and Hsp60 families are among the most widely studied families. Hsp60 is found in eubacteria, mitochondria, and chloroplasts, where, in cooperation with Hsp10, it participates in protein folding and translocation of proteins to the organelles. We have cloned and characterized the Hsp60 gene of Plasmodium yoelii (PyHsp60). PyHsp60 is a single-copy gene, located on chromosome 9, 10, or 11. The PyHsp60 cDNA sequence showed an open reading frame of 1737 nucleotides that codes for a polypeptide of 579 amino acids, with 93% amino acid identity to Plasmodium-falciparum Hsp60 (PfHsp60). Cloning and sequencing of a genomic PCR clone showed the presence of a 201-bp intron, located 141 bp downstream of the ATG codon. A single, heat-inducible, 2.3-kb transcript was detected in Northern blots of RNA isolated from blood stage parasites. Mouse antisera raised against a DNA vaccine vector that expresses PyHsp60 recognized sporozoites and liver- and blood-stage parasites by indirect fluorescent antibody test (IFAT). By Western blot, these antisera reacted with the mycobacterial Hsp65 and recognized a protein of approximately 65 kDa in P. yoelii sporozoites and P. falciparum blood stages. These results show that PyHsp60 and PfHsp60 genes are homologous and that of the PyHsp60 gene encodes a heat-inducible, intracellular protein that is expressed in several of the developmental stages of P. yoelii.

Immunogenicity and protective efficacy of a Plasmodium yoelii Hsp60 DNA vaccine in BALB/c mice.

The gene encoding the 60-kDa heat shock protein of Plasmodium yoelii (PyHsp60) was cloned into the VR1012 and VR1020 mammalian expression vectors. Groups of 10 BALB/c mice were immunized intramuscularly at 0, 3, and 9 weeks with 100 microg of PyHsp60 DNA vaccine alone or in combination with 30 microg of pmurGMCSF. Sera from immunized mice but not from vector control groups recognized P. yoelii sporozoites, liver stages, and infected erythrocytes in an indirect fluorescent antibody test. Two weeks after the last immunization, mice were challenged with 50 P. yoelii sporozoites. In one experiment the vaccine pPyHsp60-VR1012 used in combination with pmurGMCSF gave 40% protection (Fisher's exact test; P = 0.03, vaccinated versus control groups). In a second experiment this vaccine did not protect any of the immunized mice but induced a delay in the onset of parasitemia. In neither experiment was there any evidence of a protective effect against the asexual erythrocytic stage of the life cycle. In a third experiment mice were primed with PyHsp60 DNA, were boosted 2 weeks later with 2 x 10(3) irradiated P. yoelii sporozoites, and were challenged several weeks later. The presence of PyHsp60 in the immunization regimen did not lead to reduced blood-stage infection or development of parasites in hepatocytes. PyHsp60 DNA vaccines were immunogenic in BALB/c mice but did not consistently, completely protect against sporozoite challenge. The observation that in some of the PyHsp60 DNA vaccine-immunized mice there was protection against infection or a delay in the onset of parasitemia after sporozoite challenge deserves further evaluation.