Exploring views of members of the public and policymakers on the acceptability of population level dietary and active-travel policies: a qualitative study.

BACKGROUND: There is limited evidence on what shapes the acceptability of population level dietary and active-travel policies in England. This information would be useful in the decision-making process about which policies should be implemented and how to increase their effectiveness and sustainability. To fill this gap, we explored public and policymakers' views about factors that influence public acceptability of dietary and active-travel policies and how to increase public acceptability for these policies. METHODS: We conducted online, semi-structured interviews with 20 members of the public and 20 policymakers in England. A purposive sampling frame was used to recruit members of the public via a recruitment agency, based on age, sex, socioeconomic status and ethnicity. Policymakers were recruited from existing contacts within our research collaborations and via snowball sampling. We explored different dietary and active-travel policies that varied in their scope and focus. Interviews were transcribed verbatim and analysed using thematic reflexive analysis with both inductive and deductive coding. RESULTS: We identified four themes that informed public acceptability of dietary and active-travel policies: (1) perceived policy effectiveness, i.e., policies that included believable mechanisms of action, addressed valued co-benefits and barriers to engage in the behaviour; (2) perceived policy fairness, i.e., policies that provided everyone with an opportunity to benefit (mentioned only by the public), equally considered the needs of various population subgroups and rewarded 'healthy' behaviours rather than only penalising 'unhealthy' behaviours; (3) communication of policies, i.e., policies that were visible and had consistent and positive messages from the media (mentioned only by policymakers) and (4) how to improve policy support, with the main suggestion being an integrated strategy addressing multiple aspects of these behaviours, inclusive policies that consider everyone's needs and use of appropriate channels and messages in policy communication. CONCLUSIONS: Our findings highlight that members' of the public and policymakers' support for dietary and active-travel policies can be shaped by the perceived effectiveness, fairness and communication of policies and provide suggestions on how to improve policy support. This information can inform the design of acceptable policies but can also be used to help communicate existing and future policies to maximise their adoption and sustainability.

Applying genomic approaches to delineate conservation strategies using the freshwater mussel Margaritifera margaritifera in the Iberian Peninsula as a model.

Effective conservation actions to counteract the current decline of populations and species require a deep knowledge on their genetic structure. We used Single Nucleotide Polymorphisms (SNPs) to infer the population structure of the highly threatened freshwater pearl mussel Margaritifera margaritifera in the Iberian Peninsula. A total of 130 individuals were collected from 26 locations belonging to 16 basins. We obtained 31,692 SNPs through Genotyping by Sequencing (GBS) and used this dataset to infer population structure. Genetic diversity given as observed heterozygosity was low. Pairwise FST comparisons revealed low levels of genetic differentiation among geographically close populations. Up to 3 major genetic lineages were determined: Atlantic, Cantabrian and Douro. This structure suggests a close co-evolutionary process with brown trout (Salmo trutta), the primordial fish host of this mussel in the studied area. Some sub-basins showed some genetic structuring, whereas in others no intrapopulation differentiation was found. Our results confirm that genetic conservation units do not match individual basins, and that knowledge about the genetic structure is necessary before planning recovery plans that may involve relocation or restocking. The same reasoning should be applied to strictly freshwater species that are sessile or have restricted dispersal abilities and are currently imperiled worldwide.

Phylogeography, historical demography and habitat suitability modelling of freshwater fishes inhabiting seasonally fluctuating Mediterranean river systems: a case study using the Iberian cyprinid Squalius valentinus.

The Mediterranean freshwater fish fauna has evolved under constraints imposed by the seasonal weather/hydrological patterns that define the Mediterranean climate. These conditions have influenced the genetic and demographic structure of aquatic communities since their origins in the Mid-Pliocene. Freshwater species in Mediterranean-type climates will likely constitute genetically well-differentiated populations, to varying extents depending on basin size, as a consequence of fragmentation resulting from drought/flood cycles. We developed an integrative framework to study the spatial patterns in genetic diversity, demographic trends, habitat suitability modelling and landscape genetics, to evaluate the evolutionary response of Mediterranean-type freshwater fish to seasonal fluctuations in weather. To test this evolutionary response, the model species used was Squalius valentinus, an endemic cyprinid of the Spanish Levantine area, where seasonal weather fluctuations are extreme, although our findings may be extrapolated to other Mediterranean-type species. Our results underscore the significant role of the Mediterranean climate, along with Pleistocene glaciations, in diversification of S. valentinus. We found higher nuclear diversity in larger drainage basins, but higher mitochondrial diversity correlated to habitat suitability rather than basin size. We also found strong correlation between genetic structure and climatic factors associated with Mediterranean seasonality. Demographic and migration analyses suggested population expansion during glacial periods that also contributed to the current genetic structure of S. valentinus populations. The inferred models support the significant contribution of precipitation and temperature to S. valentinus habitat suitability and allow recognizing areas of habitat stability. We highlight the importance of stable habitat conditions, fostered by typical karstic springs found on the Mediterranean littoral coasts, for the preservation of freshwater species inhabiting seasonally fluctuating river systems.

Spatial heterogeneity in the Mediterranean Biodiversity Hotspot affects barcoding accuracy of its freshwater fishes.

Incomplete knowledge of biodiversity remains a stumbling block for conservation planning and even occurs within globally important Biodiversity Hotspots (BH). Although technical advances have boosted the power of molecular biodiversity assessments, the link between DNA sequences and species and the analytics to discriminate entities remain crucial. Here, we present an analysis of the first DNA barcode library for the freshwater fish fauna of the Mediterranean BH (526 spp.), with virtually complete species coverage (498 spp., 98% extant species). In order to build an identification system supporting conservation, we compared species determination by taxonomists to multiple clustering analyses of DNA barcodes for 3165 specimens. The congruence of barcode clusters with morphological determination was strongly dependent on the method of cluster delineation, but was highest with the general mixed Yule-coalescent (GMYC) model-based approach (83% of all species recovered as GMYC entity). Overall, genetic morphological discontinuities suggest the existence of up to 64 previously unrecognized candidate species. We found reduced identification accuracy when using the entire DNA-barcode database, compared with analyses on databases for individual river catchments. This scale effect has important implications for barcoding assessments and suggests that fairly simple identification pipelines provide sufficient resolution in local applications. We calculated Evolutionarily Distinct and Globally Endangered scores in order to identify candidate species for conservation priority and argue that the evolutionary content of barcode data can be used to detect priority species for future IUCN assessments. We show that large-scale barcoding inventories of complex biotas are feasible and contribute directly to the evaluation of conservation priorities.

Tiramina en la orina de los parkinsonianos idiopáticos / Urine excretion of tyramine in idiopathic parkinson disease

Hemos tabulado los datos, recuperados por pubmed.gov, de la excreción urinaria de la tiramina pues sus niveles anormales, previos a la aparición de signos clínicos, permitirían la identificación preclínica precoz de los parkinsonianos idiopáticos. El tamizado para excreción de tiramina en la orina debería realizarse, especialmente en enfermos con Parkinson, antes de cualquier tratamiento farmacológico, y en grupos de alto riesgo,. Se deben llevar a cabo estudios cuantitativos para completar las cifras normales de tiramina en la orina según el sexo, la edad, el ritmo circadiano, la dieta y otros factores que puedan afectar dicha excreción.

Heterogeneous mechanisms of azole resistance in Candida albicans clinical isolates from an HIV-infected patient on continuous fluconazole therapy for oropharyngeal candidosis.

Molecular mechanisms of azole resistance in Candida albicans include alterations in the target enzyme and increased efflux of drug, but the impact of specific treatment regimens on resistance has not been established. A patient with advanced AIDS was enrolled in a longitudinal study to receive continuous oral fluconazole (FLU) 200 mg/day for the treatment of oropharyngeal candidosis (OPC). Oral cultures were obtained at time of enrollment, during episodes of OPC and quarterly for surveillance. The patient had five symptomatic relapses on continuous FLU during 43 months. All OPC episodes were successfully treated with increasing doses of FLU although increased FLU MICs were detected for C. albicans isolates with progression of time. DNA-typing techniques demonstrated that resistance developed in a persistent strain of C. albicans. Both FLU-resistant and isogenic isolates with reduced susceptibility were detected in the same clinical samples through multiple episodes. Analysis of molecular mechanisms of resistance revealed overexpression of MDR and CDR genes encoding efflux pumps (but not ERG11) in isolates with decreased FLU susceptibility. In addition, the presence of the G464S amino acid substitution in their lanosterol demethylase, affecting its affinity for FLU, was also detected. However, other isogenic, but FLU-susceptible isolates recovered from the same samples did not harbour the mutation, indicating microevolution of yeast populations within the oral cavity. In this patient, the continuous antifungal pressure exerted by FLU resulted in development of resistance of multifactorial nature. Despite their clonal origin, different subpopulations of C. albicans demonstrated distinct resistance mechanisms, including concomitant presence and absence of functional point mutations in ERG11 genes.

Prevalence of molecular mechanisms of resistance to azole antifungal agents in Candida albicans strains displaying high-level fluconazole resistance isolated from human immunodeficiency virus-infected patients.

Molecular mechanisms of azole resistance in Candida albicans, including alterations in the target enzyme and increased efflux of drug, have been described, but the epidemiology of the resistance mechanisms has not been established. We have investigated the molecular mechanisms of resistance to azoles in C. albicans strains displaying high-level fluconazole resistance (MICs, > or =64 microg/ml) isolated from human immunodeficiency virus (HIV)-infected patients with oropharyngeal candidiasis. The levels of expression of genes encoding lanosterol 14alpha-demethylase (ERG11) and efflux transporters (MDR1 and CDR) implicated in azole resistance were monitored in matched sets of susceptible and resistant isolates. In addition, ERG11 genes were amplified by PCR, and their nucleotide sequences were determined in order to detect point mutations with a possible effect in the affinity for azoles. The analysis confirmed the multifactorial nature of azole resistance and the prevalence of these mechanisms of resistance in C. albicans clinical isolates exhibiting frank fluconazole resistance, with a predominance of overexpression of genes encoding efflux pumps, detected in 85% of all resistant isolates, being found. Alterations in the target enzyme, including functional amino acid substitutions and overexpression of the gene that encodes the enzyme, were detected in 65 and 35% of the isolates, respectively. Overall, multiple mechanisms of resistance were combined in 75% of the isolates displaying high-level fluconazole resistance. These results may help in the development of new strategies to overcome the problem of resistance as well as new treatments for this condition.

Identification of continuous B-cell epitopes on the protein moiety of the 58-kiloDalton cell wall mannoprotein of Candida albicans belonging to a family of immunodominant fungal antigens.

The 58-kiloDalton mannoprotein (mp58) on the surface of Candida albicans is highly immunogenic, is expressed by all C. albicans isolates tested, and elicits strong antibody responses during candidiasis. It belongs to a family of immunodominant fungal antigens with representatives also in different species of Aspergillus. The amino acid sequence of the protein portion of mp58 as deduced from the DNA sequence of its encoding gene (FBP1/PRA1) was used to synthesize a complete set of overlapping dodecapeptides (overlap, 7; offset, 5) covalently attached to the surface of derivatized polyethylene pins. The pin-coupled peptides were used in a modified enzyme-linked immunosorbent assay (ELISA) to identify continuous epitopes recognized by a number of antiserum preparations containing anti-mp58 antibodies. This comprehensive epitope-scanning study revealed the presence of multiple immunoreactive continuous B-cell epitopes within the protein sequence. Regions of increased reactivity included both the amino and carboxy termini of the mature protein (encompassing amino acid residues 16 to 50 and 286 to 299, respectively) and four internal regions spanning amino acids at positions 66 to 92, 121 to 142, 148 to 192, and 211 to 232. Further delineation of epitopic regions and identification of the boundaries of the antigenic sites was performed upon ELISA testing with a second Pepset consisting of completely overlapping 8-mer peptides spanning these reactive regions in the protein moiety of mp58. The highly reactive epitopic region at the C terminus of the protein was further evaluated using both window net and replacement net analyses. A synthetic peptide corresponding to the last 10 amino acid residues at the C terminus of the protein was immunogenic when injected into mice after being coupled to a carrier protein. Moreover, antibodies in the resulting sera specifically recognized the homologous mp58 in ELISAs and immunoblot assays. Delineation of the antibody responses to mp58 could provide the basis for the development of novel immunity-based prophylactic, therapeutic, and diagnostic techniques for the management of candidiasis.

IFNalpha 2b induces apoptosis and proteasome-mediated degradation of p27Kip1 in a human lung cancer cell line.

IFNs are a family of cytokines involved in antiviral defense, cell growth regulation and immune activation. IFNs either inhibit cell proliferation or control apoptosis depending on factors such as cell type and state of cell differentiation. It is important to determine how IFN-induced gene products interact with other cellular proteins to produce these responses. We have investigated the effect of IFNalpha 2b on a human small cell lung carcinoma (SCLC) cell line H82. We have found that IFNalpha efficiently induces apoptosis in H82 cells. The induction of apoptosis by IFNalpha 2b is accompanied by decreased levels of c-myc and Cdk2. We have also observed that in H82 cells IFNalpha induces downregulation of p27 and this is in contrast to the upregulation of p27 observed in other cell types where IFNs induce cell cycle arrest. IFNalpha-induced downregulation of p27 is due to protein destabilization and can be prevented by the proteasome inhibitor LLnL. The data suggest that in H82 cells, IFNalpha 2b induces degradation of p27Kip1 independently of CDK2 kinase activity and through a ubiquitin or ubiquitin-related pathway and that the degradation of p27Kip1 could be a molecular event of importance for IFN-induced apoptosis in cancer cells.

Comparison of in vitro activities of voriconazole and five established antifungal agents against different species of dermatophytes using a broth macrodilution method.

The in vitro activities of voriconazole against 19 different species of dermatophytes were compared with those of terbinafine, itraconazole, ketoconazole, griseofulvin, and fluconazole. MICs were determined according to a National Committee for Clinical Laboratory Standards broth macrodilution method. Voriconazole appeared more active than ketoconazole, griseofulvin, and fluconazole and less active than itraconazole and terbinafine. Based on these results, voriconazole merits further investigation as a potentially useful agent for the treatment of dermatophytosis.

[Azole resistance in Candida albicans]. / Resistencia de Candida albicans a los azoles.

The molecular mechanisms of azole resistance in Candida albicans include alterations in the target enzyme (lanosterol 14-demethylase) and overexpression of efflux transporters that decrease the intracellular concentration of the drug. Although the rate of azole resistance in systemic isolates of C. albicans remains very low, resistance to fluconazole appears as an important issue in the management of oropharyngeal candidiasis (OPC) in patients with AIDS. In order to establish the prevalence of resistance to azole antifungal agents in this setting, we investigated the molecular mechanisms of resistance to azoles in highly resistant C. albicans isolates (fluconazole MIC 64 mg/l) from HIV-infected patients with OPC. Antifungal susceptibility testing of serial C. albicans isolates was performed by NCCLS methodology. Strain identity was investigated by DNA-typing techniques. Overexpression of genes encoding lanosterol 14-demethylase (erg11) and efflux transporters (mdr1 and cdr) implicated in the development of resistance was monitored in matched sets of susceptible and resistant isolates. In addition, erg11 genes were PCR-amplified and their nucleotide sequences determined in order to detect point mutations. A combination of different mechanisms of resistance contributed to the development of resistance to fluconazole. The multifactorial character of the azole resistance in C. albicans makes necessary the development of approaches to overcome the problem. Accordingly, new triazoles have been developed; new classes of antifungals are being investigated; and combinations with inhibitors of efflux transporters are being studied.

Impaired angiogenic balance and suppression of tumorigenicity in HeLa cells chronically exposed to interferon-alpha.

We have previously reported that IFNalpha-chronic treatment for 41 days induced a partial phenotype reversion on HeLa cells along with a down-regulation of HPV18 mRNA levels. However, tumorigenicity of these cells in nude mice was unchanged. Interestingly, after 1 year of IFNalpha-chronic exposition, HeLa cells failed to induce s.c. tumors when injected into nude mice. In such experimental conditions both HPV18 DNA integration pattern and viral DNA copy number present in HeLa cells remained intact in the nontumorigenic phenotype cells. As result of the treatment with IFNalpha, HeLa cells rendered more resistant to lysis mediated by activated natural killer cells in vitro. Furthermore, IFNalpha-chronic treatment was able to induce VEGF and decrease bFGF mRNA expression, suggesting a potential effect on the angiogenic behavior of these tumoral cells. Thus, long-term treatment of HeLa cells with IFNalpha can accomplish a reversion of the malignant phenotype by a sequential multistep mechanism, in which the antiangiogenic effect of IFNalpha could be one of the contributing events.

Prevalence and risk factors of tinea unguium and tinea pedis in the general population in Spain.

This study prospectively evaluated the prevalence and risk factors of tinea unguium and tinea pedis in the general adult population in Madrid, Spain. One thousand subjects were clinically examined, and samples of nails and scales from the interdigital spaces of the feet were taken from those patients presenting with signs or symptoms of onychomycosis and/or tinea pedis, respectively. In addition, a sample from the fourth interdigital space of both feet was collected from all individuals with a piece of sterilized wool carpet. Tinea unguium was defined as a positive direct examination with potassium hydroxide and culture of the etiological agent from subjects with clinically abnormal nails. Patients with positive dermatophyte cultures of foot specimens were considered to have tinea pedis. The prevalence of tinea unguium was 2.8% (4.0% for men and 1.7% for women), and the prevalence of tinea pedis was 2.9% (4.2% for men and 1.7% for women). The etiological agents of tinea unguium were identified as Trichopyton rubrum (82.1%), followed by Trichopyton mentagrophytes var. interdigitale (14.3%) and Trichopyton tonsurans (3.5%). Trichophyton rubrum (44.8%) and Trichophyton mentagrophytes (44.8%), followed by Epidermophyton floccosum (7%) and T. tonsurans (3.4%), were the organisms isolated from patients with tinea pedis. The percentage of subjects who suffered simultaneously from both diseases was 1.1% (1.7% for men and 0.6% for women). In a multivariate logistic regression analysis, age (relative risk [RR], 1.03) and gender (RR, 2.50) were independent risk factors for tinea unguium, while only gender (RR, 2.65) was predictive for the occurrence of tinea pedis. In both analyses, the presence of one of the two conditions was associated with a higher risk for the appearance of the other disease (RR, >25).

Resistencia de Candida albicans a los azoles / Azole resistance in Candida albicans

Los mecanismos de resistencia a fluconazol en Candida albicans incluyen alteraciones en la enzima diana (lanosterol 14-a-desmetilasa) y sobreexpresión de transportadores activos de membrana que disminuyen la concentración intracelular de los azoles. Aunque el porcentaje de resistencia al fluconazol en aislamientos de C. albicans procedentes de infecciones sistémicas se mantiene pequeño, en la candidiasis orofaríngea en pacientes con sida constituye un grave problema. Para poder establecer la prevalencia de la resistencia al fluconazol en este caso, se han investigado los mecanismos moleculares de resistencia a los azoles en aislamientos de C. albicans resistentes al fluconazol (CMI ³64 mg/l) procedentes de pacientes VIH positivos con candidiasis orofaríngea. Las pruebas de sensibilidad se llevaron a cabo según las normas del NCCLS. La identificación de los aislamientos se realizó empleando técnicas de tipificación de DNA. La sobreexpresión de los genes que codifican para la enzima lanosterol 14-a-desmetilasa (erg11) y para los transportadores de membrana (mdr1 y cdr) se monitorizó por comparación con aislamientos isogénicos sensibles. Asimismo, los genes erg11 fueron amplificados por PCR y secuenciados para la detección de mutaciones concretas. La resistencia al fluconazol en estos aislamientos se debía a la combinación de diferentes mecanismos moleculares de resistencia. El carácter multifactorial de la resistencia de C. albicans a los azoles existentes hace necesario adoptar nuevas estrategias para combatirla, como son el desarrollo de moléculas más potentes, la investigación de nuevos antifúngicos con nuevas dianas y mecanismos de acción, así como combinaciones con inhibidores de los transportadores activos de membrana (AU)

Rifampin does not improve the efficacy of quinolone antibacterial prophylaxis in neutropenic cancer patients: results of a randomized clinical trial.

PURPOSE: To determine whether the addition of rifampin to a quinolone-based antibacterial prophylactic regimen in patients undergoing high-dose chemotherapy (HDC) with peripheral-blood stem-cell transplantation (PBSCT) decreases the incidence of neutropenia and fever, Gram-positive bacteremia, and infection-related morbidity. PATIENTS AND METHODS: Patients with solid tumors undergoing HDC with PBSCT were randomized to receive prophylactic antibiotics with either ciprofloxacin 500 mg orally every 8 hours or the same ciprofloxacin regimen with rifampin 300 mg orally every 12 hours. Prophylaxis was started 48 hours before stem-cell reinfusion. Patients were monitored to document the occurrence of neutropenia and fever, incidence and cause of bacterial infection, time to onset and duration of fever, requirement for intravenous antimicrobials, and length of hospital admission. RESULTS: Sixty-five patients were randomized to receive ciprofloxacin and 65 to receive ciprofloxacin plus rifampin, and from these groups, 62 and 61 were assessable, respectively. The proportion of patients who developed neutropenia and fever was 87% in the group treated with ciprofloxacin and 78% in the group treated with ciprofloxacin and rifampin (P =.25). Although there was a trend toward a reduction in the overall incidence of bacteremia (12 v 4 patients), and Gram-positive bacteremia (8 v 2 patients) with the addition of rifampin, none of these comparisons was statistically significant (P =.05 and P =.09, respectively). CONCLUSION: The results of this study, which demonstrate that rifampin does not improve ciprofloxacin antibacterial prophylaxis in cancer patients undergoing HDC with PBSCT support but that it does increase the occurrence of undesirable side effects, do not support the routine use of rifampin in this setting.

Human papillomavirus type 16 E7 impairs the activation of the interferon regulatory factor-1.

We had previously observed that HPV-16 E7 disturbs the Guanylate Binding Protein (GBP)-ISRE reporter activation by IFN-gamma thus suggesting an alteration of the IRF-1 function. In this study we examined the mechanism by which E7 affects the IFN-gamma signals driving the activation of gene transcription. Using 14/2 BRK cells containing dexamethasone-inducible HPV-16 E7 gene, we observed a large inhibition of the IRF-1 DNA binding activity upon E7 induction. Concomitantly, there was no significant change in the levels of IRF-1, indicating that this was not due to reduced levels of IRF-1 expression. Likewise, in vitro translated E7 did not affect the IRF-1 DNA binding activity in nuclear extracts derived from IFN-induced cells, thus indicating that the effects of E7 are upstream of IRF-1's binding to its DNA recognition site. Finally, NFkappaB DNA binding activity was also inhibited under conditional expression of E7. These data indicate that HPV-16 E7 inhibits the IRF-1 and NFkappaB function and this could lead to the impairment of the IFN response in HPV-infected cells. Furthermore, the findings suggest that different events of the IFN inducible signal cascade seem to be target for HPV-16 E7.

Comparison of high-performance liquid chromatographic and microbiological methods for determination of voriconazole levels in plasma.

A new selective high-performance liquid chromatography (HPLC) method with UV detection for the determination of the investigational triazole voriconazole in human plasma by using acetonitrile precipitation followed by reverse-phase HPLC on a C(18) column was compared with a simple agar well diffusion bioassay method with Candida kefyr ATCC 46764 as the assay organism. Pooled plasma was used to prepare standard and control samples for both methods. The results of analyses with spiked serum samples (run as unknowns) were concordant by the bioassay and HPLC methods, with expected values being obtained. HPLC demonstrated an improved precision (3.47 versus 12.12%) and accuracy (0.81 versus 1.28%) compared to those of the bioassay method. The range of linearity obtained by both methods (from 0.2 to 10 microg/ml for HPLC and from 0.25 to 20 microg/ml for the bioassay) includes the range of concentrations of voriconazole (from 1.2 to 4.7 microg/ml) which are considered clinically relevant. Although either methodology could be used for the monitoring of patient therapy, the smaller variability observed with HPLC compared to that observed with the bioassay favors the use of HPLC for pharmacokinetic studies.

Activation of the human p27(Kip1) promoter by IFNalpha 2b.

p27(Kip1) is one of the key regulatory proteins in cell cycle through inhibition of pRB phosphorylation by suppression of the activity of several cyclin/Cdk complexes. The expression of p27(Kip1) has been shown to be controlled by a posttranslational mechanism, although vitamin D(3) and neuronal differentiation can also induce its mRNA. Recently, the p27(Kip1) promoter was isolated and sequenced from a human leukocyte genomic library. In this report, we demonstrate that IFNalpha 2b, activates the human p27(Kip1) promoter-driven luciferase reporter gene in transient expression assays in H82 cells. This induction might involve two IRF 1-like binding sites present in the p27(Kip1) promoter. To our knowledge this is the first report on the direct activation of the human p27(Kip1) promoter by IFNalpha 2b.

Comparison of DNA-based typing methods to assess genetic diversity and relatedness among Candida albicans clinical isolates.

Three serial isolates of Candida albicans were obtained from each of five HIV infected patients with recurrent oropharyngeal candidiasis from the same geographical area. Isolates from one patient remained susceptible to fluconazole whereas serial isolates from the other four patients showed decreasing susceptibilities to the drug. Strain identity was investigated by pulse-field gel electrophoretic (PFGE) separation of chromosomes, restriction fragment length polymorphism (RFLP) of chromosomal DNA, Southern blot analysis with the moderately repetitive probe Ca3 of the materials present in the RFLP gels after transfer to nylon membranes, and random amplification of polymorphic DNA (RAPD). All techniques were able to group isolates obtained from the same patient. Techniques resulting in more complex banding profiles exhibited increased discriminatory power allowing detection of strain variants. Methods resulting in less complex banding patterns, especially Southern hybridization of SfiI digested chromosomal DNA with the moderately repetitive probe Ca3, were more helpful to determine isogenicity among isolates obtained from the same patient. The combination of results from methods with high discriminatory power (to maximize detection of strain variants) and methods resulting in less complex banding patterns (to allow determination of isogenic isolates) should facilitate the delineation of the epidemiology of C. albicans infection.