Chemical and biological profile of the fractionated extract of the species Campylopus savannarum (Müll. Hal.) Mitt.

The indiscriminate use of allopathic drugs has selected resistant bacterial and fungal populations which represents a severe public health problem worldwide. On the other hand, plants are in a prominent position due to the capability to synthesize structurally complex bioactive metabolites that can be an alternative to resistant microorganisms' control. In this work, we evaluated the chemical composition and the antimicrobial, antioxidant, and cytotoxic potential of the fractionated extract of C. savannarum in ethyl acetate. The extract of C. savannarum was divided into 12 fractions that were submitted to phytochemical screening, minimum inhibitory concentration (MIC), reduction of 1,1-diphenyl-2-picrylhydrazine (DPPH), and hemolytic activity of sheep erythrocytes assays. During the investigation, all extract fractions presented alkaloids, triterpenoids, steroids, and phenolic compounds in qualitative analyses, while in the quantitative evaluation, we observed the presence of both phenols and flavonoids in these fractions. Among the fraction, the highest phenolic content was observed in the Cs23-24 fraction (2.480 mg EAG/g), while the Cs31-34 fractions presented the highest amount of flavonoid (182.25 µg EQ/100 mg). Nine of the 12 fractions of the moss species' extract showed antimicrobial action Against Gram-positive bacteria: Bacillus cereus, Staphylococcus aureus, Micrococcus luteus, Bacillus subtilis; Gram-negative bacteria: Pseudomonas aeruginosa; and also antifungal activity against Candida albicans and Candida glabata. The cytotoxic assay demonstrated that the tested fractions did not induce hemolysis at concentrations 10 and 100(µG/ML). In the antioxidant evaluation, the Cs55-69 fractions were the ones that presented the highest scavenging activity (57, 0%) followed by the Cs45-54 fraction (42.7%). Overall, the evaluation of the biological potential of the fractionated extracts of Campylopus savannarum showed promising data, in the search for natural antimicrobial compounds.

Administration of a single dose of a PGF2α analogue (dinoprost) before sexual tests did not improve ram's sexual behaviour.

The aim of this study was to determine if a single administration of dinoprost tromethamine before each test enhances sexual behaviour in inexperienced rams, and if the possible effect is maintained after the rams gain sexual experience. The study was conducted in northeaster Mexico during autumn (breeding season). The study was performed with 26 Saint Croix rams (16.1 ± 1.1 months old). Rams were allocated to two treatments, which were subjected to 15 tests with 2 restrained non-oestrous ewes. While rams from one group (group DIN) received a single dose of dinoprost (10 mg im, Lutalyse, Zoetis) immediately before the test, the other rams remained untreated (group CON). Sexual behaviours were recorded during 5 min. The data were analysed with a mixed model that included the treatment, number of test and their interaction as main effects, and the individual as a random effect. DIN rams displayed less ano-genital sniffings (6.6 ± 0.8 vs 9.7 ± 0.8, P = 0.007), matings (0.11 ± 0.05 vs 0.29 ± 0.05, P = 0.03), and had a lower mating/total mounts ratio (0.05 ± 0.03 vs 0.16 ± 0.03, P = 0.006) than CON rams. There were no other treatment effects, but there were significant effects of time and interactions between treatment and time in all the behaviours. Overall, administration of a single dose of dinoprost before testing did not have positive effects in the development of rams' sexual behaviour, and even, might have negative effects. However, it should be considered that in this study only one prostaglandin analogue, one dose of this analogue, administered immediately before the tests was used, so more studies involving other analogues, doses, and/or regimes of administration should be done.

Functionalized diamond nanothreads from benzene derivatives.

Diamond nanothreads (DNTs) are fully sp3-bonded one-dimensional carbon nanostructures, synthesized recently through compression of crystalline benzene. They possess outstanding mechanical strength, suitable for the development of novel nanostructured reinforced materials. In this article, we use density functional theory calculations to investigate the feasibility and physical properties of functionalized DNTs. We show that the stacking and covalent bonding of benzene derivative molecules (toluene, aniline, phenol and fluorobenzene) may lead to stable configurations analogous to benzene-derived DNTs, with functional groups (-CH3, -NH2, -OH, -F) covalently attached to the surface. The same principle was also applied to pyridine, an aromatic heterocyclic compound, resulting in DNTs containing N heteroatoms within the sp3 C-C chain. We show that the mechanical properties remain practically unaltered compared to the original material, and that the electronic properties can be tuned upon functionalization. The presence of polar functional groups on DNT surfaces are expected to affect their compatibility with other materials and solvents, enabling the development of novel processes and technological applications using DNTs.

Sensitivity and specificity of serological tests, histopathology and immunohistochemistry for detection of Toxoplasma gondii infection in domestic chickens.

Since free-range chickens are important for the epidemiology of toxoplasmosis, this study evaluated the sensitivity and specificity of different laboratory techniques for the diagnosis of Toxoplasma gondii in these animals. Serum samples from 135 adult domestic chickens were tested for anti-T. gondii antibodies by the indirect fluorescent antibody test (IFAT), enzyme-linked immunosorbent assay (ELISA), modified agglutination test (MAT), and indirect hemagglutination test (IHAT). Tissue samples from all animals were analyzed by histopathology, immunohistochemistry and mouse bioassay (gold standard). Fifty-four chickens were positive for T. gondii in the bioassay. The sensitivity and specificity of the different tests were, respectively, 85% and 56% for ELISA; 80% and 52% for IFAT; 76% and 68% for MAT; 61% and 80% for IHAT; 7% and 98% for immunohistochemistry, and 6% and 98% for histopathology. The MAT was the most effective method for the diagnosis of T. gondii infection in chickens, followed by ELISA. Histopathology and immunohistochemistry are useful tools for the diagnosis of T. gondii infection in chickens due to their specificity.

Long-term evaluation of gait initiation in six Parkinson's disease patients with bilateral subthalamic stimulation.

Defined as the transient state between standing and walking, gait initiation is negatively affected in Parkinson's disease (PD), which often results in significant disability. Although deep brain stimulation (DBS) is the most common surgical procedure for PD, the long-term effects of DBS on gait initiation are not well studied. The present study evaluated the long-term effects of subthalamic nucleus (STN) DBS on the preparation phase of gait initiation using principal component (PC) analysis. Six patients with PD who had undergone STN DBS and 24 healthy control subjects were evaluated. PD subjects were assessed 11.3±10.3 (P1) and 78.9±10.6 (P2) months after surgery. PD subjects were tested with STN DBS in two conditions: without medication and with medication. PC analysis was applied separately for the vertical, anterior-posterior and medial-lateral components of ground reaction force (GRF) recorded during gait initiation. Three PC scores were chosen by the scree test for each GRF component and all these PC scores were used for calculating a standard distance between healthy controls and PD subjects. The Friedman test showed a significant difference in standard distance among conditions (P=0.004), with the post-hoc test recognizing differences among P1 conditions and P2 medication-on condition. The eigenvector loading factors pointed to major differences between PD conditions surrounding the maximum amplitude of vertical and anterior-posterior GRF. For the studied sample, all distances increased in the follow-up evaluation (P2) with and without medications, indicating a worsening in gait initiation after seven years.

Effect of TNF-α production inhibitors on the production of pro-inflammatory cytokines by peripheral blood mononuclear cells from HTLV-1-infected individuals

Human T lymphotropic virus type 1 (HTLV-1) is the causal agent of myelopathy/tropical spastic paraparesis (HAM/TSP), a disease mediated by the immune response. HTLV-1 induces a spontaneous proliferation and production of pro-inflammatory cytokines by T cells, and increasing interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) levels are potentially involved in tissue damage in diseases related to HTLV-1. This exaggerated immune response is also due to an inability of the natural regulatory mechanisms to down-modulate the immune response in this group of patients. TNF-α inhibitors reduce inflammation and have been shown to improve chronic inflammatory diseases in clinical trials. The aim of this study was to evaluate the ability of pentoxifylline, forskolin, rolipram, and thalidomide to decrease in vitro production of TNF-α and IFN-γ in cells of HTLV-1-infected subjects. Participants of the study included 19 patients with HAM/TSP (mean age, 53 ± 11; male:female ratio, 1:1) and 18 HTLV-1 carriers (mean age, 47 ± 11; male:female ratio, 1:2.6). Cytokines were determined by ELISA in supernatants of mononuclear cell cultures. Pentoxifylline inhibited TNF-α and IFN-γ synthesis with the minimum dose used (50 µM). The results with forskolin were similar to those observed with pentoxifylline. The doses of rolipram used were 0.01-1 µM and the best inhibition of TNF-α production was achieved with 1 µM and for IFN-γ production it was 0.01 µM. The minimum dose of thalidomide used (1 µM) inhibited TNF-α production but thalidomide did not inhibit IFN-γ production even when the maximum dose (50 µM) was used. All drugs had an in vitro inhibitory effect on TNF-α production and, with the exception of thalidomide, all of them also decreased IFN-γ production.

Effect of TNF-α production inhibitors on the production of pro-inflammatory cytokines by peripheral blood mononuclear cells from HTLV-1-infected individuals.

Human T lymphotropic virus type 1 (HTLV-1) is the causal agent of myelopathy/tropical spastic paraparesis (HAM/TSP), a disease mediated by the immune response. HTLV-1 induces a spontaneous proliferation and production of pro-inflammatory cytokines by T cells, and increasing interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) levels are potentially involved in tissue damage in diseases related to HTLV-1. This exaggerated immune response is also due to an inability of the natural regulatory mechanisms to down-modulate the immune response in this group of patients. TNF-α inhibitors reduce inflammation and have been shown to improve chronic inflammatory diseases in clinical trials. The aim of this study was to evaluate the ability of pentoxifylline, forskolin, rolipram, and thalidomide to decrease in vitro production of TNF-α and IFN-γ in cells of HTLV-1-infected subjects. Participants of the study included 19 patients with HAM/TSP (mean age, 53 ± 11; male:female ratio, 1:1) and 18 HTLV-1 carriers (mean age, 47 ± 11; male:female ratio, 1:2.6). Cytokines were determined by ELISA in supernatants of mononuclear cell cultures. Pentoxifylline inhibited TNF-α and IFN-γ synthesis with the minimum dose used (50 µM). The results with forskolin were similar to those observed with pentoxifylline. The doses of rolipram used were 0.01-1 µM and the best inhibition of TNF-α production was achieved with 1 µM and for IFN-γ production it was 0.01 µM. The minimum dose of thalidomide used (1 µM) inhibited TNF-α production but thalidomide did not inhibit IFN-γ production even when the maximum dose (50 µM) was used. All drugs had an in vitro inhibitory effect on TNF-α production and, with the exception of thalidomide, all of them also decreased IFN-γ production.

Gait initiation evaluation after deep brain stimulation for Parkinson's disease: A 7-year follow-up.

This study evaluated the long-term effects of deep brain stimulation of the subthalamic nucleus (DBS-STN) on gait initiation. Six Parkinson's disease (PD) patients who had undergone DBS-STN and 31 control subjects were evaluated. PD subjects were assessed at two different time periods: 11.3 ± 10.3 (P1) and 78.9 ± 10.6 (P2) months after surgery. Subjects under stimulation were tested in two conditions: without medication and with medication. Principal components (PC) analysis was separately applied on vertical, anterior-posterior and medial-lateral ground reaction force (GRF) from gait initiation, during the anticipatory postural adjustment (APA) phase. Three PC scores were chosen by the scree test for each GRF component. The higher loading factors pointed to major differences between controls and PD patients on maximum APA amplitude for vertical and anterior-posterior GRF. Friedman test showed a significant difference in standard distance among conditions (P = 0.006), with the post-hoc test recognizing differences only between P1 and P2 in the medication-on condition. All distances increased in the follow-up evaluation (P2), when considering the same medication condition, indicating a worsening in gait initiation after 7 years of follow-up.

Quantitative evaluation of the effects of subthalamic stimulation on gait in Parkinson's disease patients using principal component analysis.

BACKGROUND: Principal component analysis (PCA) was applied to the ground reaction force (GRF) for evaluating the deep brain stimulation of the subthalamic nucleus (DBS-STN) effects in Parkinson's disease (PD) subjects with and without medication. METHODS: Ten subjects who underwent DBS-STN were evaluated under the following four conditions: without treatment (mof-sof), with stimulation (mof-son), with medication (mon-sof), and with both treatments (mon-son). A control group of 30 subjects was also evaluated. PCA was applied separately on each GRF component. Broken stick criterion selected eight principal components (PC) from vertical GRF and one from each horizontal GRF. A standard distance was calculated using these 10 PCs and the gait speed to measure how far the PD group's gait was from the normal pattern. RESULTS: The standard distance allowed classifying normal and PD subjects in the mof-sof condition with 100% accuracy, sensitivity, and specificity. The same distance was calculated for mon-sof, mof-son, and mon-son conditions. The smallest mean standard distance was found in the mon-son condition, which was significantly different from mof-sof (Friedman test with Dunn post-hoc, p < .05). CONCLUSION: PCA allowed the quantitative evaluation of treatment effects, indicating that DBS-STN improves the GRF pattern in PD subjects, primarily in the medication on state.

Comparison among probabilistic neural network, support vector machine and logistic regression for evaluating the effect of subthalamic stimulation in Parkinson disease on ground reaction force during gait.

Deep brain stimulation of the subthalamic nucleus (DBS-STN) is an approved treatment for advanced Parkinson disease (PD) patients; however, there is a need to further evaluate its effect on gait. This study compares logistic regression (LR), probabilistic neural network (PNN) and support vector machine (SVM) classifiers for discriminating between normal and PD subjects in assessing the effects of DBS-STN on ground reaction force (GRF) with and without medication. Gait analysis of 45 subjects (30 normal and 15 PD subjects who underwent bilateral DBS-STN) was performed. PD subjects were assessed under four test conditions: without treatment (mof-sof), with stimulation alone (mof-son), with medication alone (mon-sof), and with medication and stimulation (mon-son). Principal component (PC) analysis was applied to the three components of GRF separately, where six PC scores from vertical, one from anterior-posterior and one from medial-lateral were chosen by the broken stick test. Stepwise LR analysis employed the first two and fifth vertical PC scores as input variables. Using the bootstrap approach to compare model performances for classifying GRF patterns from normal and untreated PD subjects, the first three and the fifth vertical PCs were attained as SVM input variables, while the same ones plus the first anterior-posterior were selected as PNN input variables. PNN performed better than LR and SVM according to area under the receiver operating characteristic curve and the negative likelihood ratio. When evaluating treatment effects, the classifiers indicated that DBS-STN alone was more effective than medication alone, but the greatest improvements occurred with both treatments together.

Assessment of the effects of subthalamic stimulation in Parkinson disease patients by artificial neural network.

This study aims at using a probabilistic neural network (PNN) for discriminating between normal and Parkinson disease (PD) subjects using as input the principal components (PCs) derived from vertical component of the ground reaction force (vGRF). The trained PNN was further used for evaluating the effects of deep brain stimulation of the subthalamic nucleus (STN DBS) on PD, with and without medication. A sample of 45 subjects (30 normal and 15 PD subjects who underwent STN DBS) was evaluated by gait analysis. PD subjects were assessed under four test conditions: without treatment (mof-sof), only with stimulation (mof-son) or medication (mon-sof), and with combined treatments (mon-son). PC analysis was applied on vGRF, where six PC scores were chosen by the broken stick test. Using a bootstrap approach for the PNN model, and the area under the receiver operating characteristic curve (AUC) as performance measurement, the first three and fifth PCs were selected as input variables. The PNN presented AUC = 0.995 for classifying controls and PD subjects in the mof-sof condition. When applied to classify the PD subjects under treatment, the PNN indicated that STN DBS alone is more effective than medication, and further vGRF enhancement is obtained with combined therapies.

Application of principal component analysis in vertical ground reaction force to discriminate normal and abnormal gait.

Discrete parameters from ground reaction force (GRF) are been considered in gait analysis studies. However, principal component analysis (PCA) may provide additional insight into gait analysis for considering the complete pattern of GRF. This study aimed at testing the application of PCA to discriminate the vertical GRF pattern between control group (CG) and patients with lower limb fractures (FG), as well as proposing a score to quantify the abnormality of gait. Thirty-eight healthy subjects participated of CG and 13 subjects in FG, five subjects from FG were also evaluated after physiotherapeutic treatment (FGA). The GRF was measured by an instrumented treadmill. Principal component coefficients (PCCs) were obtained by singular value decomposition using GRF of complete stride. Two, four and six PCCs were used to obtain the standard distance (D). The classification between groups was mainly given by the first PC, which indicated higher loading factors during push off of affected side and heel strike of unaffected side. The classification performance achieved 92.2% accuracy with two PCCs, 94.1% with four PCCs and 96.1% with six PCCs. Four subjects reached normal boundary after treatment, with all FGA subjects presenting decreased D. This study demonstrates that PCA is an adequate method for discriminating normal and abnormal gait and D allows an objective evaluation of the progress and effectiveness of rehabilitation treatment.

[Salivary and lacrymal gland disorders and HTLV-1 infection]. / Atteinte des glandes salivaires et lacrymales et infection par le HTLV-1.

INTRODUCTION: The frequency and severity of salivary and lacrymal gland human T-cell lymphotropic virus type 1 (HTLV-1) infection were assessed in HTLV-1 plus patients, presenting with neurological deficit (tropical spastic paraparesis/HTLV-1 associated myelopathy [TSP/HAM]) or not. The mechanism of this deficit was investigated. MATERIAL AND METHODS: A case-control study was made from April 2002 to December 2005, in an area strongly endemic for HTLV-1. The patients were classified in three groups: group 1 with 16 patients presenting with TSP/HAM; group 2 with 67 HTLV-1 carriers and group 3 with 29 healthy volunteers. The dry syndrome was investigated by history taking and by oral and ophthalmological clinical examination. Immunological and biological screening for rhumatoid factors, antinuclear antibodies, and antibodies against soluble nuclear antigens (SSA, SSB). Peripheral blood was separated by density gradient and mononuclear cells were recovered to dose interferon-gamma and tumor necrosis factor-alpha. Patients in the three groups were assessed for salivary flow by stimulated weighing using Saxon's test. A Chi-2 test, a variance analysis (Anova), and the Spearman rank correlation test were used for the statistical analysis. RESULTS: The dry syndrome was mild and more common in group 1 patients (75%). In group 2, 22% of the patients presented with functional signs of buccal mucosa dryness comparable to those observed in group 1. No correlation was found between salivary flow and screened pro-inflammatory cytokines. DISCUSSION: Our results show that hyposialia is an important part of the disease induced by HTLV-1, even in virus carriers without neurological deficit. Its mechanism seems different than that of the Gougerot-Sjögren syndrome.

Levels of serum chemokines discriminate clinical myelopathy associated with human T lymphotropic virus type 1 (HTLV-1)/tropical spastic paraparesis (HAM/TSP) disease from HTLV-1 carrier state.

Approximately 5% of people infected with human T lymphotropic virus type 1 (HTLV-1) develop clinical myelopathy or tropical spastic paraparesis (HAM/TSP) that is associated with high-levels of Th1 cytokines, interferon (IFN)-gamma and tumour necrosis factor (TNF)-alpha. Chemokines are known to induce cytokine secretion and direct the trafficking of immune cells to sites of disease. The present study measured serum chemokines correlated with autonomously released IFN-gamma in cell cultures. HTLV-1 infection was defined by enzyme-linked immunosorbent assay (ELISA) and confirmed by Western blot. Subjects included HTLV-1 carriers (n = 56), patients with HAM/TSP (n = 31) and healthy HTLV-1 seronegative volunteer controls (n = 20). Serum chemokines and IFN-gamma autonomously released by mononuclear cells in culture were quantified by ELISA. Compared to HTLV-1 carriers, serum chemokines in HAM/TSP patients showed significantly increased levels of CXCL9 and CXCL10, significantly diminished levels of CCL2 and similar amounts of CCL11 and CCL24. In contrast, CCL11 and CCL24 were significantly lower in serum of HAM/TSP patients than either control. IFN-gamma was positively correlated with CXCL9 and CXCL10 when HAM/TSP and HTLV-1 carriers were used as a combined group. However, despite a large proportion of HTLV-1 carriers having high IFN-gamma levels, these chemokines were not increased in carriers. This study showed that high levels of CXCL9 and CXCL10 in the systemic circulation and low serum CCL2 levels are features of HAM/TSP. HTLV-1 infection and Tax and/or additional viral encoded factor-mediated pathological processes triggering T cell activation with autogenous IFN-gamma release are probably involved in regulating chemokine release.

Erectile dysfunction and HTLV-I infection: a silent problem.

The human T-lymphotropic virus type I (HTLV-I) is a retrovirus associated with a chronic myelopathy known as HTLV-I-Associated Myelopathy or Tropical Spastic Paraparesis (HAM/TSP). The main objective was to assess the frequency of erectile dysfunction (ED) in HTLV-I-infected individuals from Salvador and other cities from Bahia, Brazil, as well as to verify if sexual dysfunction correlates with urinary symptoms and overall neurological impairment. From January 2001 to April 2004, 218 HTLV-I carriers (111 male and 107 female subjects) had complete clinical, neurological, and urological evaluation. They were assessed using standardized questionnaires to determine urinary complaints (Urinary Distress Inventory) and ED (Brief Male Sexual Function Inventory). Neurological impairment was established by Expanded Disability Status Scale (EDSS) from 0 to 10. HAM/TSP was considered as EDSS> or =2. A total of 17 males had clinically defined HAM/TSP (group 1). From the 94 HTLV-I-infected males, 62 were selected (group 2) and paired by age with patients in group 1. A total of 79 individuals were selected for this study. The age ranged from 35 to 81 y (mean=47.9+/-9.65). The percentage of ED in the studied population was 40.5%. In the HAM/TSP group, ED frequency was 88.2%. The associations among sexual dissatisfaction, erectile dysfunction, urinary symptoms (frequency, nocturia, and urgency) and EDSS> or =2 were statistically significant. In HAM/TSP, there is a slow and progressive degeneration of the lateral funiculus of the spinal cord. HTLV-I-infected individuals present a high frequency of ED and it is closely associated to urinary symptoms and the overall neurological picture. The HTLV-I carriers already had prominent compromise of the sexual activity.

Fluorometric study of rabbit sperm head membrane phospholipid asymmetry during capacitation and acrosome reaction using Annexin-V FITC.

This study was conducted to evaluate phosphatidylserine translocation in head plasma membrane of Percoll-gradient purified of rabbit cauda epididymal sperm during capacitation and acrosome reaction (AR) using Annexin-V. Propidium iodide was used as control to reject dead or dying cells. The presence and distribution of Annexin-V binding sites were analyzed using flow fluorocytometry and confocal microscopy. After 6 h of incubation of sperm in capacitation medium, the number of cells positively stained with Annexin-V showed a small but significant increment. The Annexin-V binding sites produced during capacitation were found mainly in the post-acrosomal region of the sperm head plasma membrane. After AR induction with progesterone, the localization of phosphatidylserine was changed and the Annexin-V binding sites were found almost only in the acrosomal region, but with higher number of binding sites in the equatorial area. On the contrary, after AR induction with A23187, phosphatidylserine translocation, although predominant over the acrosomal region, was also observed in the post-acrosomal region. Plasma membrane destabilization during capacitation and AR may be important for sperm-oocyte fusion.

Absence seizures effects on reading revealed by video-electroencephalography.

OBJECTIVE: The effects of absence seizures on reading have not been studied. The purpose of this paper is to show some alterations in reading during absence seizures in patients with video-electroencephalographic recordings. METHODS: Patients were selected if absences seizures were the only type of seizures. They were studied at rest with eyes closed and while reading aloud a text by simultaneous bipolar electroencephalographic and video recordings. RESULTS: Eight patients were studied with 34 events. Absences ranged from 1-12 events in twenty-five minutes recording, seizures duration ranged from 1.5-16 seconds. Brief seizures results on non-stop reading, but reading arrest with latency of 1-3 seconds after seizure onset were present in the longer events. Post-absence amnesia ranged from 1-4.5 seconds. Re-start reading occurs mainly in other reading place in the text, but two patients in one event re-started reading in the right place. Classic symptoms during absence were present and reading alterations found were: reading arrest, lost of reading point, bradylexia, dysprosodic reading, grapheme substitution, reading loudness reduction, repeating of phrases. Events occurred without patient seizure awareness, except in one patient by means of the time lost between onset to the end of the event. CONCLUSIONS: This study showed that acute reading alterations during absence seizures are frequent in longer seizures, producing many neuropsychological symptoms that alter reading without seizure awareness.

Beta-thalassaemia in Cubans: novel allele increases the genetic diversity at the HBB locus in the Caribbean.

In order to establish the molecular basis of beta-thalassaemia in Cubans, a total of 75 unrelated individuals, with beta-thalassaemia major (7), Hb S-beta-thalassaemia (28), Hb C-beta-thalassaemia (1), and beta-thalassaemia trait (39) yielding 82 beta-thalassaemia alleles, were analyzed. Seventeen different point mutations were identified accounting for 93% of the beta-thalassaemia alleles studied, revealing a high genetic heterogeneity at the HBB locus in this population. The more prevalent mutations, namely, CD 39 (C --> T) (30.5%), -29 (A --> G) (13.4%), IVS-I-110 (G --> A) (8.5%), and IVS-II-1 (G --> A) (8.5%), reflect the Mediterranean and African predominant ancestry of the extant Cuban population. We also report the identification of a novel allele, IVS-I-108 (T --> C), that possibly activates a cryptic branch site, in a beta-thalassaemia carrier with no other molecular defect within the beta-globin gene and its proximal promoter. This study shows that prenatal diagnosis of beta-thalassaemia should be feasible in about 60% of at-risk pregnancies by direct detection of selected point mutations. However, due to the wide spectrum of mutations, and in order to offer fully informative prenatal diagnosis to more than 87% of at-risk couples, the screening for beta-thalassaemia mutations in Cubans should be performed by using a general point mutation detection method, such as DGGE (denaturing gradient gel electrophoresis).