Clinical features and immune mechanisms directly linked to the altered liver function in patients with rheumatoid arthritis.

BACKGROUND: The aim of this study was to explore the impact of arthritis on liver function using different approaches in vivo and in vitro. METHODS: A cross-sectional study was performed on 330 non-obese/non-T2DM subjects: 180 RA patients, 50 NAFLD non-RA patients, and 100 healthy donors (HDs). A longitudinal study was conducted on 50 RA patients treated with methotrexate for six months. Clinical and laboratory parameters and markers of liver disease were collected. Mechanistic studies were carried out in both the CIA mouse model and hepatocytes treated with anti-citrullinated protein antibodies (ACPAs). RESULTS: RA patients have an increased risk of suffering from liver disease independent of obesity or T2DM. This risk was associated with factors such as insulin resistance, autoantibodies, inflammation, and component C3. Methotrexate treatment for six months was associated with liver abnormalities in those newly-diagnosed patients having CV risk factors. ACPAs induced a defective hepatocyte function, promoting IR and inflammation. The induction of arthritis in mice caused the infiltration of immune cells in the liver and increased inflammatory, apoptotic, and fibrotic processes. CONCLUSION: RA patients may experience mild to moderate liver inflammation due to the infiltration of T, B cells, and macrophages, and the action of ACPAs. This is independent of obesity or diabetes and linked to systemic inflammation, and disease activity levels. The negative effects of methotrexate on liver function could be restricted to the concomitant presence of cardiovascular risk factors.

Binding by calmodulin is coupled to transient unfolding of the third FF domain of Prp40A.

Human pre-mRNA processing protein 40 homolog A (hPrp40A) is a splicing factor that interacts with the Huntington's disease protein huntingtin (Htt). Evidence has accumulated that both Htt and hPrp40A are modulated by the intracellular Ca2+ sensor calmodulin (CaM). Here we report characterization of the interaction of human CM with the third FF domain (FF3 ) of hPrp40A using calorimetric, fluorescence and structural approaches. Homology modeling, differential scanning calorimetry and small angle X-ray scattering (SAXS) data show FF3 forms a folded globular domain. CaM was found to bind FF3 in a Ca2+ -dependent manner with a 1:1 stoichiometry and a dissociation constant (Kd ) of 25 ± 3 µM at 25°C. NMR studies showed that both domains of CaM are engaged in binding and SAXS analysis of the FF3 -CaM complex revealed CaM occupies an extended configuration. Analysis of the FF3 sequence showed that the anchors for CaM binding must be buried in its hydrophobic core, suggesting that binding to CaM requires unfolding of FF3 . Trp anchors were proposed based on sequence analysis and confirmed by intrinsic Trp fluorescence of FF3 upon binding of CaM and substantial reductions in affinity for Trp-Ala FF3 mutants. The consensus model of the complex showed that binding to CaM binding occurs to an extended, non-globular state of the FF3 , consistent with coupling to transient unfolding of the domain. The implications of these results are discussed in the context of the complex interplay of Ca2+ signaling and Ca2+ sensor proteins in modulating Prp40A-Htt function.

Small-angle X-ray scattering to quantify the incorporation and analyze the disposition of magnetic nanoparticles inside cells.

Access to detailed information on cells loaded with nanoparticles with nanoscale precision is of a long-standing interest in many areas of nanomedicine. In this context, designing a single experiment able to provide statistical mean data from a large number of living unsectioned cells concerning information on the nanoparticle size and aggregation inside cell endosomes and accurate nanoparticle cell up-take is of paramount importance. Small-angle X-ray scattering (SAXS) is presented here as a tool to achieve such relevant data. Experiments were carried out in cultures of B16F0 murine melanoma and A549 human lung adenocarcinoma cell lines loaded with various iron oxide nanostructures displaying distinctive structural characteristics. Five systems of water-dispersible magnetic nanoparticles (MNP) of different size, polydispersity and morphology were analyzed, namely, nearly monodisperse MNP with 11 and 13 nm mean size coated with meso-2,3-dimercaptosuccinic acid, more polydisperse 6 nm colloids coated with citric acid and two nanoflowers (NF) systems of 24 and 27 nm in size resulting from the aggregation of 8 nm MNP. Up-take was determined for each system using B16F0 cells. Here we show that SAXS pattern provides high resolution information on nanoparticles disposition inside endosomes of the cytoplasm through the structure factor analysis, on nanoparticles size and dispersity after their incorporation by the cell and on up-take quantification from the extrapolation of the intensity in absolute scale to null scattering vector. We also report on the cell culture preparation to reach sensitivity for the observation of MNP inside cell endosomes using high brightness SAXS synchrotron source. Our results show that SAXS can become a valuable tool for analyzing MNP in cells and tissues.

Daylight photodynamic therapy for prevention of new actinic keratosis and keratinocyte carcinomas in organ transplants. A cryotherapy-controlled randomized clinical trial.

BACKGROUND: Organ transplant recipients (OTR) have a higher risk of actinic keratosis (AK) and keratinocyte carcinomas (KC). There are no clinical trials assessing the effectiveness of daylight photodynamic therapy (DPDT) to prevent new AK and KC in OTR. OBJECTIVES: To determine whether repeated treatments of field cancerization with DPDT are effective in preventing new AK and KC in OTR. METHODS: A randomized, intra-subject controlled, evaluator-blind, split-face and/or scalp trial, from April 2016 to October 2018. Participants were OTR older than 18 years, 1-year posttransplant, with at least 5 AK on each hemi-face/hemi-scalp. One side received six field treatments with DPDT: two sessions 15 days apart at baseline, two at 3 months and two at 9 months after baseline. Control side received lesion-directed treatment with cryotherapy (double freeze-thaw) at baseline, 3 and 9 months. Total number of lesions (AK and KC) at 21 months, number of new AK and KC at 3, 9, 15 and 21 months and treatment preferences were analysed. RESULTS: Of 24 men included, 23 were analysed at 3 months; and 21, at 9, 15 and 21 months. Mean (SD) age was 69.8 years (9.2). The total number of lesions at 21 months was 4.7 (4.3) for DPDT and 5.8 (5.0) for control side; P = 0.09. DPDT showed significantly lower means [SD] of new lesions compared to control side at 3 months (4.2 [3.4] vs. 6.8 [4.8]; P < 0.001), 9 months (3.0 [3.3] vs. 4.3 [3.4]; P = 0.04) and 15 months (3.0 [4.6] vs. 4.8 [5.0]; P = 0.02), and non-significant at 21 months (3.7 [3.5] vs. 5.0 [4.5]; P = 0.06). Most participants preferred DPDT. CONCLUSION: DPDT showed potential effectiveness in preventing new AK and KC in OTR by consecutive treatments of field cancerization. The preference for DPDT could facilitate adherence to the long-term treatment necessary in these patients.

Fat Quality Index and Risk of Cardiovascular Disease in the Sun Project.

OBJECTIVES: To examine the association between a dietary fat quality index (FQI), and the risk of incident cardiovascular events or deaths in the Seguimiento Universidad de Navarra (SUN) cohort. DESIGN: Longitudinal analysis during 10.1 years of median follow-up. Cox models were used to estimate adjusted hazard ratios (HR) of incident cardiovascular diseases (CVD) according to tertiles of FQI and of different fat subtypes. SETTING: University of Navarra, Spain. PARTICIPANTS: 19,341 middle-aged adults. MEASUREMENTS: Fat intake was measured with a validated food-frequency questionnaire. The FQI was calculated according to the ratio: (monounsaturated+polyunsaturated) / (saturated+trans fatty acids). RESULTS: We observed 140 incident cases of CVD. No association was found for FQI (HR=0.94, 95 %CI 0.61-1.47 for the highest vs the lowest tertile, p for trend=0.884). No significant associations were found for different dietary fat subtypes on CVD risk. The results suggest no clear association between a higher FQI and a higher amount of energy from fat and incidence of CVD (p for interaction: 0.259 and p for trend only among participants with a percentage of energy from fat ≥35% of total energy: 0.272). CONCLUSION: In this Mediterranean cohort, the FQI was not associated with cardiovascular events. A "heart-healthy diet" should focus its attention on dietary fat sources and should use an overall dietary pattern approach, rather than limiting the focus on fat subtypes. More research is needed to validate dietary advice on specific fatty acids intake or saturated fatty acids replacements for reducing CVD risk.

[Influence of initial protocolized treatment with steroids in length of stay and costs of community acquired pneumonia]. / Influencia de la protocolización del tratamiento inicial con corticoides en la estancia y costo de las neumonías adquiridas en la comunidad. Estudio prospectivo 2015.

OBJECTIVE: The aim of the study was to analyze the impact of steroid treatment in patients with community acquired pneumonia (CAP), both in length of stay and economical cost of admission at a clinical university hospital. METHODS: Prospective study of admitted patients with the diagnosis of CAP, both in Internal Medicine and Infectious diseases department. The study was conducted from January to march 2015; patients receiving steroids from diagnosis to end of antibiotic treatment were classified as group I; otherwise, they were considered in group II. Administration of steroids was done according to the criteria of the responsible. Cost was stablished according to CAP Diagnostic Related Group (DRG). RESULTS: Prevalence of patients younger than 65 year-old was higher in group I (p<0.05). In bivariate analyses, mean admission time was lower in group I (5.37 vs 8.88 days) (p<0.0005) and also economical cost (2,361 euros vs 3,907 euros) (p<0.0005). In multivariate analysis, factors independently associated to higher cost (>3,520 euros) were COPD (OR=2.602; 95% CI 1.074-6.305) and group II (patients with no steroids) (OR=6.2; p=0,007). CONCLUSIONS: No administration of steroids in patients with CAP was associated, together with COPD, with higher economical cost (evaluated by DRG/length of stay).

Loading dose of Dexdor® and optimal sedation during oral and maxillofacial ambulatory surgery procedures: An observational study. / Dosis de carga de Dexdor® y nivel óptimo de sedación durante procedimientos de cirugía oral y maxilofacial en régimen ambulatorio: un estudio observacional.

INTRODUCTION: Dexdor® do not include the possibility of loading dose, which could increase time to achieve adequate sedation for ambulatory procedures. The objective of this study was to evaluate the effect of several loading dose of dexmedetomidine in the time to achieve and maintain an optimal level of sedation and its clinical hemodynamic repercussion. MATERIAL AND METHODS: The IRB approved this observational study for patients that underwent oral and maxillofacial ambulatory surgery under dexmedetomidine at the University of Navarra Clinic from February 2013 to November 2014. According to the loading dose the patients were grouped into 3 categories:<0.5, 0.5, and>0.5µg/kg. Optimal level of sedation was defined as bispectral index<85. Data were analyzed using survival analysis techniques. Vasoactive drugs requirements was evaluated using exact logistic regression. RESULTS: Eighty-one patients were evaluated. Hazard ratios for patients in 0.5 and >0.5µg/kg loading dose categories for achieving a bispectral index<85 were 1.5 (95% CI 0.9, 2.6) and 1.8 (95% CI 0.8, 3.9), respectively, compared with the lowest category. Five patients (6.2%) required atropine for bradycardia. Patients in the group>0.5µg/kg showed greater risk of requiring atropine compared with the group<0.5µg/kg (odds ratio 2.2; 95% CI 0.03, 183). CONCLUSION: Loading dose of dexmedetomidine>0.5µg/kg appears minimize the time to achieve and maintain an optimal level of sedation during the first 60min of procedure. Further investigation to elucidate the association between loading dose of dexmedetomidine and subsequent atropine requirements may be warranted.

In Vitro Activities of Novel Antimicrobial Combinations against Extensively Drug-Resistant Acinetobacter baumannii.

Extensively drug-resistant (XDR) Acinetobacter spp. have emerged as a cause of nosocomial infections, especially under conditions of intensive care. Unfortunately, resistance to colistin is increasing and there is a need for new therapeutic options. We aimed to study the effect of some novel combinations against XDR Acinetobacter baumannii in an in vitro pharmacokinetics-pharmacodynamics (PK/PD) model. Three nonrelated clinical strains of XDR A. baumannii were investigated. Antibiotic-simulated regimens were colistin at 3 MU every 8 h (q8h) (first dose, 6 MU), daptomycin at 10 mg/kg of body weight q24h, imipenem at 1 g q8h, and ertapenem at 1 g q24h. Combination regimens included colistin plus daptomycin, colistin plus imipenem, and imipenem plus ertapenem. Samples were obtained at 0, 1, 2, 4, 8, and 24 h. Among the single-agent regimens, only the colistin regimen resulted in significant reductions in log10 CFU per milliliter compared to the control for all the strains tested. Although colistin achieved bactericidal activity at 4 h, it was not able to reach the limit of detection (1 log10 CFU/ml). One strain had significant regrowth at 24 h without the emergence of resistance. Daptomycin-colistin combinations led to a significant reduction in levels of log10 CFU per milliliter that were better than those achieved with colistin as a single-agent regimen, reaching the limit of detection at 24 h against all the strains. The combination of imipenem plus ertapenem outperformed the colistin regimen, although the results did not reach the limit of detection, with significant regrowth at 24 h. Similarly, colistin-plus-imipenem combinations reduced the levels of log10 CFU per milliliter at 8 h, with significant regrowth at 24 h but with development of resistance to colistin. We have shown some potentially useful alternatives for the treatment of extensively drug-resistant A. baumannii. Among them, the daptomycin-colistin combination was the most effective and should be investigated in future studies.

Do Skin Prick Test and In Vitro Techniques Diagnose Sensitization to Peach Lipid Transfer Protein and Profilin Equally Well in Allergy to Plant Food and Pollen?

OBJECTIVE: To compare the skin prick test (SPT) with in vitro techniques (single and multiplex fluorescence enzyme-immunoassay [FEIA]) for detecting sensitization to profilin and lipid transfer protein (LTP). METHODS: We retrospectively studied 181 patients with pollen and/or plant food allergy and 61 controls. SPT was performed with date palm profilin (Pho d 2) and peach LTP (Pru p 3), and specific IgE (sIgE) to Phl p 12 and Pru p 3 was analyzed using single FEIA and microarray. RESULTS: Fifteen of 201 patients with negative results for LTP in the SPT were sensitized to this allergen in the in vitro tests, and 18 of 41 patients with positive results for LTP in the SPT were not sensitized according to the in vitro tests. Seventeen of 186 patients with negative results for profilin in the SPT were sensitized to Phl p 12 by serum sIgE, and 30 out of 56 patients with positive results for profilin in SPT were not sensitized to Phl p 12 according to the other tests. Moderate agreement was observed between the 3 techniques studied. CONCLUSIONS: SPT is a sensitive technique for detecting sensitization to LTP and profilin. Its results are similar to those of in vitro techniques, especially in patients with negative SPT results for peach LTP and palm tree profilin.

Early skin testing is effective for diagnosis of hypersensitivity reactions occurring during anesthesia.

Allergic skin tests have to be performed 4-6 weeks after an allergic anesthetic reaction. Patients with allergic reactions during anesthesia were prospectively included (n = 44). Skin tests were performed in two stages: (i) Stage 1 (S1), 0-4 days after the reaction; and (ii) Stage 2 (S2), 4-8 weeks after. Five (11.5%) surgical procedures were suspended due to the reaction. Positive skin tests were obtained in 25/44 patients (57%). Allergic diagnosis was carried out at S1 in 15/25 (60%) and at S2 in 10/25 (40%). Three patients resulted positive only in S1. Overall agreement among S1 and S2 skin tests was 70.45%. The kappa statistic was 0.41 (P-value = 0.002). Odds ratio of obtaining a false negative in S1 (compared with S2) was 3.33. Early allergological study is useful, could minimize false negatives, but should be considered as a complement to late skin tests.

Silica SBA-15 template assisted synthesis of ultrasmall and homogeneously sized copper nanoparticles.

The synthesis of spherical copper nanoparticles with extremely narrow size distribution by electroless copper deposition on mesoporous silica support is described. The materials were characterized by nitrogen sorption, transmission electron microscopy, x-ray diffractometry and Fourier transform infrared spectroscopy. The copper nanoparticles have a cubic crystalline structure and an average particle size of 5.5 +/- 0.8 nm. The copper nanoparticles are stable, without detectable oxidation or further agglomeration under ambient conditions even after months. These results demonstrate that electroless copper reduction can be conducted and constrained within the mesoporous silica framework, which pave the way for engineered mesoreactors.

Biexponential analysis of diffusion-tensor imaging of the brain in patients with cirrhosis before and after liver transplantation.

BACKGROUND AND PURPOSE: DTI has shown increased MD of water molecules in the brain of patients with cirrhosis, consistent with low-grade edema. This study further characterizes this edema by using biexponential analysis of DTI data, a technique that may differentiate cytotoxic and vasogenic edema. MATERIALS AND METHODS: A total of 41 patients with cirrhosis awaiting liver transplantation and 16 healthy controls were studied by DTI by using a single-shot echo-planar technique with 11 b-values (range, 0-7500 s/mm(2)) and 6 noncollinear directions. Measurements were fitted to biexponential function to determine MD and FA for the fast and slow diffusion components. Regions of interest were selected in the parietal white matter and corticospinal tract. The assessment was repeated 1 year after liver transplantation in 24 of these patients. RESULTS: In parietal white matter, patients with cirrhosis showed an increase in fast MD and a decrease in fast FA that normalized after liver transplantation. In the corticospinal tract, there was an increase in fast and slow MD that normalized after transplantation, and a decrease in FA that persisted posttransplantation. There was no association of DTI parameters with minimal HE (n =12). CONCLUSIONS: Biexponential analysis of DTI supports the presence of edema in the brain of patients with cirrhosis that reverts after transplantation. In parietal white matter, the increase in brain water was mainly located in the interstitial compartment, while the corticospinal tract showed a mixed pattern (intra- and extracellular). In addition, the findings on posttransplantation were consistent with microstructural damage along the corticospinal tract.

Risk factors of A/H1N1 etiology in pneumonia and its impact on mortality.

OBJECTIVE: Pandemic flu has changed the epidemiology of pneumonia, thus challenging the prediction of etiology and outcome. We analyze the risk factors to predict influenza A/H1N1 infection in patients with pneumonia, and the impact of this etiology on mortality during a pandemic period. Differences between pneumonia with or without A/H1N1 coinfection are described. METHODS: Retrospective observational study in 364 consecutive patients hospitalized with pneumonia during the A/H1N1 pandemic flu, April-December 2009. RESULTS: 294 patients (80.5%) had A/H1N1(-) pneumonia, 47 (13.2%) A/H1N1(+) pneumonia, and 23 (6.3%) coinfection. Mortality during hospitalization was 24/294 (8.2%), 8/47 (16.7%), 2/23 (8.7%) respectively. A regression logistic analysis (Area under curve, AUC 0.81) to predict A/H1N1(+) pneumonia identified four independent variables: age < 60 years (Odds ratio, OR 5.9), multilobar infiltrates (OR 7.7), C-reactive protein (CRP) < 10 mg/dL (OR 2.8), and leukopenia < 5000/mm(3) (OR 3.4). Risk factors for in-hospital mortality in the whole group were A/H1N1 (+) etiology and LDH > 600 IU/L (OR 4.1) when adjusting for PSI, and hypoxemia (OR 4.2) when adjusting for CURB 65 (AUC 0.81). Heart disease (OR 27.4) and LDH > 600 IU/L (OR 10.5) were risk factors for in-hospital mortality in A/H1N1(+) patients (AUC 0.81) CONCLUSION: Leukopenia, multilobar infiltrates, CRP<10 mg/dl and age < 60 years were independently associated with A/H1N1(+) etiology. Pandemic A/H1N1(+) increased mortality pneumonia. Heart disease and LDH > 600 were independently associated with mortality in A/H1N1(+) pneumonia.

Review article: the design of clinical trials in hepatic encephalopathy--an International Society for Hepatic Encephalopathy and Nitrogen Metabolism (ISHEN) consensus statement.

BACKGROUND: The clinical classification of hepatic encephalopathy is largely subjective, which has led to difficulties in designing trials in this field. AIMS: To review the current classification of hepatic encephalopathy and to develop consensus guidelines on the design and conduct of future clinical trials. METHODS: A round table was convened at the 14th International Society for Hepatic Encephalopathy and Nitrogen Metabolism (ISHEN) meeting. Key discussion points were the nomenclature of hepatic encephalopathy and the selection of patients, standards of care and end-points for assessing the treatment and secondary prevention of hepatic encephalopathy. RESULTS: It was generally agreed that severity assessment of hepatic encephalopathy in patients with cirrhosis, whether made clinically or more objectively, should be continuous rather than categorical, and a system for assessing the SONIC (Spectrum of Neuro-cognitive Impairment in Cirrhosis) was proposed. Within this system, patients currently classified as having minimal hepatic encephalopathy and Grade I hepatic encephalopathy would be classified as having Covert hepatic encephalopathy, whereas those with apparent clinical abnormalities would continue to be classified as overt hepatic encephalopathy. Some aspects of the terminology require further debate. Consensus was also reached on the patient populations, standards of care and endpoints to assess clinical trial outcomes. However, some compromises had to be made as there is considerable inter- and intravariability in the availability of some of the more objective surrogate performance markers. CONCLUSIONS: The objectives of the round table were met. Robust, defendable guidelines for the conduct of future studies into hepatic encephalopathy have been provided. Outstanding issues are few and will continue to be discussed.

Temporomandibular joint synovial chondromatosis with a traumatic etiology.

Synovial chondromatosis (SC) is a metaplastic disorder characterized by the formation of cartilaginous nodules inside the articular space. SC is uncommon in the temporomandibular joint (TMJ). A few reports suggest a correlation between a traumatic episode and the development of SC. The authors describe the diagnosis, treatment and follow-up of a patient with unilateral SC of the left TMJ in conjunction with bony resorption on the mandibular condyle and a clear traumatic etiology. They review and comment on previous reports in the literature.

Serum total tryptase levels are increased in patients with active chronic urticaria.

BACKGROUND: We have demonstrated previously mast cell histamine release upon incubation with chronic urticaria (CU) sera, presumably by degranulation. OBJECTIVE: To explore total and mature tryptase in order to assess whether any increase in total tryptase levels is due in part to mast cell degranulation or to mast cell burden. We also wanted to explore differences between the autoimmune groups called idiopathic (serum unable to activate basophils), and to correlate total and mature tryptase levels with different urticaria features. METHODS: We measured total and mature tryptase serum levels in 81 CU patients, 16 atopic donors and 21 healthy control sera. We assessed autoimmunity by measuring the CD63 expression in normal basophil donors upon incubation with CU sera. RESULTS: We found significantly higher levels of total tryptase in the sera of CU patients (6.6 ±4.1 µg/L) than in sera from healthy non-atopic subjects (4.4 ±2.8 µg/L) and from atopic subjects (4.5 ±1.7 µg/L). Mature tryptase levels were undetectable (<1 ng/mL). Total tryptase levels in the autoimmune urticaria group were significantly higher (9.8 ±5.4 µg/L) than the idiopathic urticaria group (4.4 ±2.2 µg/L). A significant difference in total tryptase was found between symptomatic patients (7.3 ±4.1 µg/L) compared with asymptomatic ones (5.7 ±4.1 µg/L) at the time of venesection. No difference was found in mature tryptase levels either. CONCLUSION: Total elevated tryptase levels are not accompanied by an elevated mature tryptase levels, as might be expected if the serum levels reflected mast cell degranulation.

[Paratesticular adenomatoid tumor: a report of nine cases]. / Tumor adenomatoide paratesticular: una serie de nueve casos.

INTRODUCTION: Paratesticular tumors are rare. Most of them are benign, and adenomatoid tumors are most common. These tumors sometimes infiltrate the testicular parenchyma and require differential diagnosis with malignant tumors. In such cases, intraoperative biopsy allows for performing conservative surgery. MATERIALS AND METHODS: A retrospective study of nine patients with paratesticular adenomatoid tumors seen during a nine-year period (2000-2008) is reported. RESULTS AND CONCLUSIONS: Patient age (mean, 49.6 years) and most common initial signs (tender nodule) are reported. The tumor most commonly occurred as a small, usually oval, nodule in the tail of epididymis. Our series included a case each of intraparenchymal tumor of the testis and tumor of the tunica vaginalis. Diagnosis was initially made based on a ultrasound scan and subsequently confirmed by histology. Differential diagnosis and surgical treatment, performed in all patients, are reported, and also the pathological features of surgical specimens.

Differentiation of yeasts growing on dry-cured Iberian ham by mitochondrial DNA restriction analysis, RAPD-PCR and their volatile compounds production.

The efficiency of mitochondrial DNA (mtDNA) restriction analysis, RAPD-PCR and volatile compounds analysis to differentiate yeast biotypes involved in flavour development of dry-cured Iberian ham throughout the ripening process is evaluated. For this purpose, 86 yeasts isolated from Iberian hams in the main ripening stages at different industries of the four Protected Designations of Origin of this product, were used. The combination of mtDNA restriction analysis and RAPD-PCR using the primer (GACA)4 showed a higher variability in the yeast species detected than obtained using only mtDNA restriction analysis. Only two species, Debaryomyces hansenii and Candida zeylanoides, were identified throughout the whole ripening process and a wide diversity of biotypes was found in these two species, with those of D. hansenii predominating. Clear differences between biotypes were detected in the generation of volatile compounds, with the biotype C2-2 of D. hansenii showing the highest concentrations of volatiles. The combined use of mtDNA restriction analysis and RAPD-PCR distinguishes yeast biotypes with different production of volatile compounds. In addition, analysis of the production profile of volatile compounds is needed to differentiate yeast strains of the same biotype recovered at different stages of ripening. Thus, the combination of these three methods could be very useful to select or monitor yeasts as starter cultures in dry-cured meat products.