Search

SUMMARY This study investigated the fatigue failure load of simplified monolithic yttria partially stabilized zirconia polycrystal restorations cemented to a dentin-like substrate using different luting systems. Disc-shaped ceramic (Zenostar T, 10 mm Ø × 0.7 mm thick) and dentin-like substrate (10 mm Ø × 2.8 mm thick) were produced and randomly allocated into eight groups, without or with thermocycling (TC=5–55°C/12,000×): “cement” (RelyX Luting 2 – glass ionomer cement [Ion], [Ion/TC]; RelyX U200 – self-adhesive resin cement [Self], [Self/TC]; Single Bond Universal+RelyX Ultimate – MDP-containing adhesive + resin cement [MDPAD + RC], [MDP-AD + RC/TC]; ED Primer II+Panavia F 2.0 – Primer + MDP-containing resin cement [PR + MDP-RC], [PR + MDP-RC/TC])). Each luting system was used as recommended by the manufacturer. Staircase methodology (20 Hz; 250,000 cycles) was applied for obtaining the fatigue failure loads. Fractographic characteristics were also assessed. At baseline, the Ion group presented the lowest fatigue load, although it was statistically similar to the Self group. The resin-based cement systems presented the highest fatigue performance, with the Ion group being only statistically equal to the Self group. Thermocycling influenced the groups differently. After aging, the MDP-AD + RC presented the highest mean, followed by the PR + MDP-RC and Self groups, while the Ion group had the lowest mean. Fractographic analysis depicted all failures as radial cracks starting at the zirconia intaglio surface. The luting system with MDP-containing adhesive applied prior to the resin cement presented the highest fatigue failure load after aging, presenting the best predictability of stable performance. Despite this, monolithic zirconia presents high load-bearing capability regardless of the luting agent.

Pneumonia caused by opportunistic fungi is a serious complication in immunocompromised patients. Hypercalcemia has been described in renal transplantation associated with Pneumocystis jirovecii (PJP) or Histoplasma capsulatum (HCP) pneumonia.We describe 5 patients who underwent kidney transplant between 2014 and 2019 and developed hypercalcemia before the diagnosis of pulmonary fungal infection: 4 patients with PJP and 1 with HCP. We assessed calcium metabolism and kidney function by total and ionized calcium, phosphorus, intact parathormone (iPTH), 25-OH vitamin D, 1,25(OH)2 vitamin D, and serum creatinine levels.Mean albumin-corrected calcium and ionized calcium were 12.56 mg/dL (range, 10.8-13.8 mg/dL) and 1.57 mmol/L (range, 1.43-1.69 mmol/L). Patients were normocalcemic, at 10.12 mg/dL (range, 9.6-10.5 mg/dL), before diagnosis and resolved hypercalcemia after antifungal treatment, at 8.86 mg/dL (range, 8.0-9.5 mg/dL). All patients had low or normal iPTH values, at 29.1 pg/mL (range, 3-44 pg/mL), with higher PTH levels 3 months before diagnosis and after treatment, at 147.3 pg/mL (range, 28.1-479 pg/mL) and 117.5 pg/mL (range, 18.2-245 pg/mL), respectively. The mean value for 25-OH vitamin D was 30.8 ng/mL (range, 14.6-62.8 ng/mL). This supports a PTH-independent mechanism, and we postulated an extrarenal production of 1,25(OH)2 vitamin D.In kidney transplant patients, hypercalcemia independent of PTH and refractory to treatment should alert for the possibility of opportunistic fungal pneumonia.

The methanol extract of the seed of Sphenocentrum jollyanum Pierre, was subjected to partition chromatography in gradient elutions, using vacuum liquid chromatography (VLC) and HPLC techniques. It led to the isolation of two compounds labelled as DO5d-2 and DO6, whose structures were determined by complete analyses of their spectroscopic data, utilizing modern techniques (LC-MS, 1H- and 13C-NMR, DEPT-NMR, HSQC, HMBC, COSY and NOESY). The Essential oils of the leaf and stem were extracted by hydro-distillation using Clevenger-typed apparatus and analysed by GC-MS technique. It was subsequently subjected to antimicrobial analysis using the agar diffusion method. The major isolated compound – DO6, was further characterized as ‘columbin’, while DO5d-2 was characterized as a new clerodane type furano-diterpene, which has not been previously reported. Essential oils obtained by hydro distillation from leaf and stem of Sphenocentrum jollyanum Pierre, were dominated by aldehydes in 10.93% and 15.14% respectively. Fifty (50) compounds were identified in leaf oil comprising 68.73% of it, with phytol (6.62%) and n-hexanol (6.28%) as most abundant compounds. Twenty-eight (28) compounds identified in the stem oil comprised 44.90% containing pentadecanal (14.72%) and phytol (6.38%) as its prominent compounds. The antimicrobial result showed the oil from the leaf was active against all the bacterial and fungi tested, while that from the stem revealed moderate activity against the tested bacterial and fungi.

BACKGROUND The impact of renal transplantation (RT) in the elderly with many comorbid conditions is a matter of concern. The aim of our study was to assess the impact of RT on the survival of patients older than 60 years compared with those remaining on the waiting list (WL) according to their comorbidities. METHODS In this multicentric observational retrospective cohort study, we included all patients older than 60 years old admitted on the WL from 01 January 2006 to 31 December 2016. The Charlson comorbidity index (CCI) score was calculated for each patient at inclusion on the WL. Kidney donor risk index was used to assess donor characteristics. RESULTS One thousand and thirty-six patients were included on the WL of which 371 (36%) received an RT during a median follow-up period of 2.5 (1.4-4.1) years. Patient survival was higher after RT compared to patients remaining on the WL, 87%, 80%, and 72% versus 87%, 55%, and 30% at 1, 3, and 5 years, respectively. After RT survival at 5 years was 37% higher for patients with CCI ≥ 3, and 46% higher in those with CCI < 3, compared with patients remaining on the WL. On univariate and multivariate analysis, patient survival was independently associated with a CCI of ≥3 (hazard ratio 1.62; confidence interval 1.09-2.41; P < 0.02) and the use of calcineurin-based therapy maintenance therapy (hazard ratio 0.53; confidence interval 0.34-0.82; P < 0.004). CONCLUSIONS Our study showed that RT improved survival in patients older than 60 years even those with high comorbidities. The survival after transplantation was also affected by comorbidities.

The outcome of patients who return to dialysis after Kidney allograft failure (KAF) remains unclear. Our aim was to compare the outcome of KAF patients vs two different types of transplant naive incident dialysis (TNID) patients, those on the waiting list (WL) and those with a kidney transplant contraindication (KTC).We performed an observational study using data from the Argentinian Dialysis Registry between 2005 and 2016. We compare mortality between KAF, WL, and KTC.We included 75 722 patients of which 2734 were KAF. Survival between the three cohorts (KAF vs WL (n = 14 630) vs KTC (n = 58 358) revealed a significant difference (log-rank test: P .0001) indicating worse survival for KTC patients and best survival for WL. We found that KAF patients had as poor outcome as KTC patients after multivariate adjustment. Cox regression showed that age65 years: HR: 1.845 (1.79-1.89) P .0001, transient catheter: HR: 1.303 (1.26-1.34) P .0001, diabetic: HR: 1.273 (1.22-1.31) P .0001, hepatitis C: HR: 1.156 (1.09-1.22) P .0001, and albumin: HR: 1.247 (1.21-1.28) P .0001 were associated with mortality.Patients who return to dialysis after KAF have higher mortality than WL patients and similar to KTC patients.

SUMMARYObjective:To evaluate the effect of hydrofluoric acid (HF) etching and thermocycling (Tc) on fatigue failure load of feldspathic ceramic restorations cemented with two resin cements.Methods:Disc-shaped feldspathic ceramic (Vitablocs Mark II; Ø=10 mm, 1.0-mm thick) and G10 epoxy resin (Ø=10 mm, 2.5-mm thick) specimens were made and randomly allocated considering three factors: ceramic etching (ie, with vs without 10% HF plus silane application), resin cement (ie, self-adhesive [RelyX U200; U200] or conventional [Multilink Automix; MA]), and Tc (ie, with vs without 5-55°C/12,000 cycles). Adhesive cementation followed each manufacturer's instructions. The fatigue test (n=20) was based on the staircase approach (250,000 cycles; 20 Hz). Contact angle, surface topography, and fractography analysis were also executed. Specific statistical tests were employed for each outcome (α=0.05).Results:The interaction of HF and Tc factors decreased the fatigue resistance for both cements (U200 542.63>U200/HF-Tc 495.00; MA 544.47>MA/HF-Tc 506.84). Comparing the cements associated with HF or Tc, there was statistical superiority for MA (U200-Tc 537.37Conclusion:HF etching plus silane agent increased the ceramic surface free energy and its wettability, but it did not provide better results in terms of fatigue resistance compared with silane agent application only. The association of HF etching and aging significantly reduced the fatigue resistance of the material, regardless of the resin cement used.

Kidney transplant patients (KTPs), and particularly those with advanced chronic kidney rejection, may be affected by opportunistic infections, metabolic alterations and vascular and oncologic diseases that promote clinical conditions that require a variety of treatments, the combinations of which may predispose them to hyponatremia. Salt and water imbalance can induce abnormalities in volemia and/or serum sodium depending on the nature of this alteration (increase or decrease), its absolute magnitude (mild or severe) and its relative magnitude (body sodium:water ratio). Hyponatremia appears when the body sodium:water ratio is reduced due to an increase in body water or a reduction in body sodium. Additionally, hyponatremia is classified as normotonic, hypertonic and hypotonic and while hypotonic hyponatremia is classified in hyponatremia with normal, high or low extracellular fluid. The main causes of hyponatremia in KTPs are hypotonic hyponatremia secondary to water and salt contraction with oral hydration (gastroenteritis, sepsis), free water retention (severe renal failure, syndrome of inappropriate antidiuretic hormone release, hypothyroidism), chronic hypokalemia (rapamycin, malnutrition), sodium loss (tubular dysfunction secondary to nephrocalcinosis, acute tubular necrosis, tubulitis/rejection, interstitial nephritis, adrenal insufficiency, aldosterone resistance, pancreatic drainage, kidney–pancreas transplant) and hyponatremia induced by medication (opioids, cyclophosphamide, psychoactive, potent diuretics and calcineurinic inhibitors). In conclusion, KTPs are predisposed to develop hyponatremia since they are exposed to immunologic, infectious, pharmacologic and oncologic disorders, the combinations of which alter their salt and water homeostatic capacity.

Introduction Living kidney donor (LKD) transplantation is increasing due to organ shortage. Clinical studies have shown that the risk of developing end-stage renal disease (ESRD) in donors is similar to that in the general population. Our goal was to evaluate postdonation renal outcomes assessed by glomerular filtration rate (GFR), proteinuria, and blood pressure. Methods A total of 210 LKD transplants were performed at Hospital Italiano de Buenos Aires between 2000 and 2014. Postdonation outcomes were analyzed in 109 donors. GFR was assessed by 24-hour creatinine clearance (as 24-hour ClCr) and estimated using the Modification of Diet in Renal Disease (MDRD) and Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) equations. Additionally, we correlated the predonation renal functional reserve (RFR) with postdonation GFR. Donor results were compared to the expected GFR (adjusted to age and single kidney). Other renal outcome indicators measured were albuminuria and blood pressure, and they were compared (predonation and postdonation) using univariate analysis. Results A total of 109 patients were followed up for 47 ± 34 months (range, 12–168): 70% were female, age at donation was 48.58 years (range, 25–70), and predonation serum creatinine was 0.85 ± 0.17 mg/dL. Postnephrectomy GFR (24-hour ClCr) was significantly lower compared to predonation GFR (105.38 ± 21.78 mL/min/1.73 m2 vs 90.14 ± 17.78 mL/min/1.73 m2). However, postdonation GFR was not significantly different compared to the expected GFR. No differences were found for blood pressure or albuminuria. Age >50 and an RFR ( Conclusions In this population of LKD, renal outcome (24-hour CrCl, albuminuria, and blood pressure) was within the expected outcome for healthy individuals after uninephrectomy.

Physical therapy modalities are commonly prescribed to relieve pain and to restore full function of older adults suffering from musculoskeletal diseases. The choice of a specific physical modality depends on patient’s specific disease, his health conditions, and comorbidities.

Background Proteinuria (P) is a early sign of inflammation and renal damage. It has an important role in the detection, diagnosis, and monitoring of renal disease in transplanted patients. The aim of this study was to examine the correlation between random urinary proteinuria/creatininuria index (P/CI) and 24-hour total protein excretion among stable renal transplant patients. Materials and Methods We obtained 1511 samples of 24-hour protein excretion (24-hr P) with corresponding P/CI were obtained from 197 adult patients beyond 6 months post-transplantation between 2009 and 2011. The population was divided into 2 groups: One to obtain a population of justification (755) and another, of validation (755). A scatter graft yielded was obtained by Pearson's coefficient of correlation. A “receiver operater characteristic curve” analysis was carried out to evaluate the sensitivity and specifity of PCI and 24hr-P, showing a cutoff of 0.15 for PCI. Results The PCI and 24 hr P Pearson's correlation was significant (r = 0.89; P = .0001). The sensitivities of the P/CI for the justification and the validation samples were 97% and 94%, respectively; the a cutoff was 0.15. Their negative predictive values for P/CI were 92% and 84% respectively (cutoff, 0.15). The specificity was below 50% in both groups. Conclusions We observed a significant correlation between P/CI and 24 hr P. The sensitivity was slightly higher than the specificity (50%) but the negative predictive value was >92%. The use of P/CI seemed to be adequate for screening of protein excretion during renal transplant recipient follow-up.

The Modification of Diet in Renal Disease (MDRD) equation is widely used to estimate glomerular filtration rate (eGFR) in kidney transplant (KT) patients. The novel Chronic Kidney Disease-Epidemiology Collaboration equation (CKD-EPI) could improve accuracy of GFR estimation. Our aim was to compare both equations for staging of CKD in KT patients.In a cohort of KT patients, correlation of eGFR according to MDRD and CKD-EPI with 24-hour creatinine clearance (24h-CrCl) was evaluated. Concordance between both equations for CKD staging was performed. MDRD was used for initial CKD staging. The mean difference of GFR between 24h-CrCl and each equation was calculated and Bland-Altman analysis applied.A cohort of 463 KT patients were studied: 67% female, overall average age 46 ± 14 years, 41% living donor, mean time of transplantation 71 months (3-95), and mean serum creatinine 1.68 ± 1.03 mg/dL. For the whole cohort (all CKD stages), eGFR by CKD-EPI was 5.33 mL/min/1.73 m(2) higher than by MDRD (P .01). For CKD stages 1, 2, and 3A, the mean eGFR differences (CKD-EPI - MDRD) were 13.98 ± 3.27, 8.2 ± 1.98, and 5.34 ± 1.32 mL/min/1,73 m(2), respectively. The percentage of patients with eGFR 60 mL/min/1.73 m(2) decreased from 63.8% according to MDRD to 53.9% with the use of CKD-EPI. In women and patients ≤65 years old, eGFR by CKD-EPI was 5.98 and 5.81 mL/min/1.73 m(2) higher, respectively, than by MDRD (P .01).The novel CKD-EPI reduces the number of patients with eGFR 60 mL/min/1.73 m(2) and consequently assigns lower CKD stages to our KT population.

There are several immunological and non-immunological factors related to renal graft deterioration, and histological lesions such as interstitial fibrosis and tubular atrophy overlap with those observed in aging kidneys. Consequently, it has been proposed that kidney transplant senescence could contribute to graft loss. The process of cell senescence displays characteristics such as an increased expression of specific aging suppressor genes, shortened telomeres, mitochondrial changes, increased expression of negative regulators of the cell cycle, and immunological senescence. Additionally, tubular frailty characterizes the aged kidney, making it more susceptible to ischemia, reperfusion, toxic injury, and consequently, to inflammation. Moreover, renal tissue injury predisposes the older graft not only to progressive deterioration due to glomerular hyperfiltration, but also triggers acute rejection due to increased immunogenicity. In conclusion, renal graft senescence is a complex process, and its better understanding will help the nephrologist in its management in order to achieve a longer graft survival.

The tacrolimus is metabolized primarily by CYP3A5, a member of the single nucleotide polymorphism family. It shows cytochrome P450 (SNP) in intron 3, which consists of a change of base, G for A, producing a stop codon. The result is a nonfunctional protein (allele *3). Allele *1 is the wild type. The patients that show the allelic variant *3 in homozygosis (G/G) are slow metabolizers of the immunosuppressant, increasing its concentration in blood. In contrast, heterozygote A/G alleles *1/*3 are intermediate metabolizers, whereas those of allele *1 in homozygosis (A/A) are normal metabolizers. The aim of this study was to determine CYP 3A5 polymorphism among adult renal transplant recipients and the general Argentinean population. We analyzed 21 recipients and 36 healthy controls. All subjects gave written informed consent approved by the local committee. To determine the polymorphism, we extracted DNA from peripheral blood and used polymerase chain reaction (PCR) to amplify intron 3 of the CYP 3A5. The presence of variant was confirmed by direct sequencing. Among the controls the CYP3A5 genotype *3/*3 (G/G) was detected in 32 individuals, 4 showed *1/*3 (A/G), and none had *1/*1 (A/A); among the recipients, the results were as follows: 18, 2, and 1, respectively. The frequencies of polymorphism in both groups were similar, although they differred from those published for other populations. These results are the basis for the development of a pharmacogenomic program applied to organ transplantation. The genetic polymorphisms can determine responses to drugs. The molecular diagnosis must be transferred to clinical practice so as to guide selection of medicine and drug doses to be optimal for each individual.

Ao se apropriar das Tecnologias da Informação e Comunicação (TICs), a Geografia materializa os seus conteúdos didáticos, via objetos de aprendizagem, como instrumento auxiliar do processo ensino-aprendizagem, privilegiando a interatividade entre o educando e os saberes didáticos. Nesse sentido, esta pesquisa teve como foco central elaborar um objeto de aprendizagem sobre alfabetização cartográfica e validá-lo com os sujeitos do ensino e aprendizagem de Geografia. Como objetivos intrínsecos, tiveramse: (a) investigar práticas pedagógicas para o ensino e aprendizagem de Geografia, por meio de objeto de aprendizagem; (b) desenvolver e efetivar o objeto de aprendizagem em Geografia; (c) analisar a aplicação prática do objeto de aprendizagem na quinta série do Ensino Fundamental. Ressalta-se que o objeto de aprendizagem, desenvolvido em uma perspectiva interdisciplinar e interativa, de caráter construtivista, vem a contribuir como alternativa prática e viável, atraindo e motivando, com melhores resultados, à aprendizagem dos alunos. Os aspectos positivos desse recurso didático podem ser comprovados através da validação do objeto de aprendizagem junto aos alunos de quinta série do Ensino Fundamental.

The aim of this study was to evaluate growth hormone (GH) and ghrelin levels in response to physical exercise in athletes.Two different exercise workloads were administered in two different groups of athletes. Group A athletes (19 males, 18 females; mean age +/- standard deviation: 25+/-6.7 years), performing a 60-90 min training session at approximately 80% of VO2max, were sampled for GH and ghrelin determinations before and immediately at the end of a training session on-the-field. Group B athletes (4 males; mean age: 28.2+/-7.2 years) performed two consecutive 30-min cycling sessions at 80% of individual VO2max at different time intervals between bouts (2 and 6 h) in two different days. GH and ghrelin concentrations were determined in blood samples collected at 15-min intervals during exercise and following 1 h of recovery.In group A athletes, GH levels increased after the training session (P0.0001), with no differences between males and females. In male athletes, ghrelin levels significantly decreased after the training session (from 1 506.4+/-859 to 1 254.8+/-661.7 pg/mL, P0.05), while no significant changes were found in females. No correlations were observed between GH and ghrelin levels at rest and after training. In group B athletes, GH levels significantly increased after the first exercise bouts (peak: 26.8+/-11.2 and 17.3+/-3.5 ng/mL, respectively), while the pattern of GH response was different after the second bout of exercise performed at 2-h or 6-h interval. In fact, peak GH concentration in response to the second bout (4.3+/-1.6 ng/mL) was lower (P0.01) than that of the first bout when the interval elapsed was only 2 h, while a recovery of GH responsiveness was evident after the 6-h interval between the two exercise bouts (11.9+/-3.3 ng/mL). As far as ghrelin levels are concerned, no significant changes were observed during and after the two exercise bouts performed at the different time intervals.GH responses to prolonged exercise bouts (60-90 min) are associated with changes in ghrelin levels only in male athletes, while repeated exercise bouts of lower duration (30 min), capable to determine marked GH responses, are divorced from changes in ghrelin concentrations.

O artigo descreve o processo de desenvolvimento dos objetos de aprendizagem de Geografia, que compõem a série denominada Capitão Tormenta e Paco. A série é composta por quatro objetos que contemplam os conteúdos de Fusos Horários, Movimentos da Terra, Redes Geográficas e Estações do Ano. O artigo descreve as fases de desenvolvimento dos objetos, referindo-se à metodologia RIVED, à linha pedagógica adotada e às fases de desenvolvimento ao mesmo tempo em que vai apresentando as interfaces dos objetos. O artigo é finalizado apontando os ganhos da equipe com o desenvolvimento e o atendimento às recomendações para efetivar a qualidade e usabilidades dos objetos de aprendizagem desenvolvidos.

PCR with random primers (RAPD analysis) performed on the DNA of embryogenic and non-embryogenic suspension cultured rice and of transformed rice plants allows the evaluation of the extent of DNA changes in the different biological materials. This is thus suggested as a convenient approach, in combination with restriction analysis and Southerns blotting, to evaluate the integrity of the foreign gene, the stability of the insertion site and the stability of the whole genome.

Sequence analysis of twelve DNA fragments, which had previously been found to be extensively amplified in suspension-cultured rice cells, revealed that two of them, isolated on plasmids designated pE10 and pE11, have sequences identical to distinct regions of chloroplast DNA (ct-DNA). Both sequences are part of an extrachromosomal circular DNA molecule (ECD). The molecular structure of the ECD was investigated by a combination of restriction analysis, standard and pulsed-field gel electrophoresis, hybridization with ct-DNA probes and amplification by the polymerase chain reaction in the presence of oligonucleotide primers homologous to selected regions of rice ct-DNA. The results showed that a continuous and unrearranged stretch of ct-DNA from the long single-copy region, of at least 28 kbp in length, is present in the ECD. It was estimated that the number of copies of the ECD in cultured cells was almost equivalent to that of ct-DNA molecules in rice leaves, while the ratio of ECD to ct-DNA molecules in the cultured cells was approximately 200:1.

Sequence analysis of twelve DNA fragments, which had previously been found to be extensively amplified in suspension-cultured rice cells, revealed that two of them, isolated on plasmids designated pE10 and pE11, have sequences identical to distinct regions of chloroplast DNA (ct-DNA). Both sequences are part of an extrachromosomal circular DNA molecule (ECD). The molecular structure of the ECD was investigated by a combination of restriction analysis, standard and pulsed-field gel electrophoresis, hybridization with ct-DNA probes and amplification by the polymerase chain reaction in the presence of oligonucleotide primers homologous to selected regions of rice ct-DNA. The results showed that a continuous and unrearranged stretch of ct-DNA from the long single-copy region, of at least 28 kbp in length, is present in the ECD. It was estimated that the number of copies of the ECD in cultured cells was almost equivalent to that of ct-DNA molecules in rice leaves, while the ratio of ECD to ct-DNA molecules in the cultured cells was approximately 200:1.

International audience; Protoplasts of Nicotiana tabacum were transformed with the APH(3')II gene, which confers kanamycin resistance. Plants resistant to kanamycin were differentiated and 3 of them were chosen at random. These were used to study the stability of the foreign gene after a cycle of dedifferentiation, to produce calli, and differentiation, to produce new plants. The effect of the selective pressure was analyzed by performing dedifferentiation and differentiation in the presence or absence of kanamycin. Inbred plants were also produced from the original transformed plants and used as control. Southern blot analysis of DNA extracted from 66 regenerated plants showed in all cases that no detectable alteration occurred both in gene structure and insertion site. Furthermore the specific activity of the APH(3')II enzyme was shown to be at high level in all regenerated plants regardless of the fact that they were regenerated in the presence or absence of kanamycin. The results described here are experimental evidence that a hybrid forcing gene is rather stable in a heterologous genome even after dedifferentiation of the transformed plants and differentiation in vitro, i.e. in those conditions known to be correlated with extensive somaclonal variation.

The plasmid pE10 is a pBR322-derived plasmid carrying a 4.5 kb rice (Oryza sativa L.) repeated DNA sequence. The cloned sequence has been shown to be amplified in cultured rice cells. The analysis of practically intact chromosomal rice DNA molecules by pulsed field gel electrophoresis has now shown that the amplification is associated with the appearance of extrachromosomal molecules. In fact, pE10 hybridizes exclusively with unfractionated DNA from leaf protoplasts, while it recognizes predominantly an extrachromosomal DNA molecule (ECD) of about 45 kb and its multiples in the case of protoplasts from cultured cells. Insensitivity to the action of the exonuclease Bal31 suggests that the molecule is circular. Analysis of restriction endonuclease products with both standard horizontal and pulsed field gel electrophoresis suggest that the extrachromosomal DNA, and its chromosomal counterpart, is composed of tandemly repeated units of about 7 kb. Thus, the smaller extrachromosomal circle should contain 6–7 repeats, while the sequence cloned in pE10 is a subset of this repeat. The extrachromosomal DNA represents about 1 % of total rice DNA and its level of amplification is not affected by the different phases of growth in culture.

The emergence capability of maize seeds in a kaolinitic red tropical soil from Zimbabwe, subjected to rainfall simulation, was investigated under three kinds of treatment: untreated, 2.5 T·ha−1 and 5 T·ha−1 of phosphogypsum. Samples treated with gypsum exhibited a strong increase of crust strength and a decrease of the percentage of the cracked areas; they also showed a lower bulk density and a higher residual moisture content in air-dry condition. Gypsum translocation was limited to the first millimeters of the soil. A possible explanation advanced is the loss of brittle behaviour in treated samples, due to greater retention of clay in the surface soil layer; non-brittle behaviour by the treated soil would hamper soil cracking by emerging seeds and their emergence through cracks.

The occurrence of genomic modifications in transgenic rice plants recovered from protoplasts and their transmission to the self-pollination progeny has been verified with the random amplified polymorphic DNA (RAPD) approach. The plant was the Indica-type rice (Oryza sativa L.) cultivar Chinsurah Boro II. The analysed material was: (1) microspore-derived embryogenic rice cells grown in suspension culture, (2) transgenic plants recovered from protoplasts produced from the cultured cells and (3) the self-pollination progeny (two successive generations) of the transgenic plants. DNA purified from samples of these materials was PCR-amplified with different random oligonucleotide primers and the amplification products were analysed by agarose gel electrophoresis. Band polymorphism was scored and used in band-sharing analyses to produce a similarity matrix. Relationships among the analysed genomes were expressed in a dendrogram. The extensive DNA changes evidenced in cultured cells demonstrate the occurrence of somaclonal variation in the material used to produce protoplasts for gene transfer. Quantitatively reduced DNA changes were also found in the resulting transgenic plants and in their self-pollination progenies. While confirming the stability of the foreign gene in transgenic plants, this work gives molecular evidence for the occurrence of stable genomic changes in transgenic plants and points to in vitro cell culture as the causative agent. RAPDs are shown to be a convenient tool to detect and estimate the phenomenon at the molecular level. The methodology is also proposed as a fast tool to select those transgenic individuals that retain the most balanced genomic structure and to control the result of back-crosses planned to restore the original genome.

Lactobacilli play a crucial role in maintaining the ecological equilibrium of the vaginal niche, preventing the colonization of exogenous microorganisms. Although many studies have discussed the mechanisms displayed by lactobacilli in counteracting several urogenital pathogens, a few data are available on the interaction between lactobacilli and Chlamydia trachomatis. This study aimed to elucidate the molecular bases of the interaction among vaginal lactobacilli, the sexually transmitted pathogen C. trachomatis and the epithelial cervical cells. We evaluated the in vitro activity of 15 Lactobacillus strains, belonging to different species (i.e., L. crispatus, L. gasseri, L. vaginalis), against C. trachomatis. In particular, we evaluated the capability of lactobacilli cells to interfere with C. trachomatis infection in HeLa cells, by exclusion assays. Lactobacilli significantly reduced C. trachomatis infectivity, being L. crispatus the most active species. Although a dose-dependent effect was noticed, a significant antagonistic activity was maintained even at lower doses. As other Gram-positive bacteria (i.e., Streptococcus agalactiae, Enterococcus faecalis, and Bacillus subtilis) failed to interfere with C. trachomatis infectivity, Lactobacillus activity proved to be specific. The potential mechanism of protection was investigated in Lactobacillus crispatus BC5, chosen as the model strain. The incubation of HeLa cell line with BC5 cells induced important modifications in the epithelial plasma membrane, by altering lipid composition and α5 integrin subunit exposure. When α5 integrin subunits were masked by a specific blocking antibody or ITGA5 gene expression was silenced, Chlamydia infection was significantly reduced. It follows that α5 integrin subunit is crucial for the pathogen infection process, and the anti-Chlamydia activity can be directly linked to membrane properties modifications in cervical cells. The three Gram-positive bacteria used as controls failed to modify the expression of α5β1 integrin. In conclusion, we identified a potential molecular mechanism at the basis of the protection exerted by L. crispatus BC5 against C. trachomatis, getting insights into the role of the cervico-vaginal microbiota for the woman’s health.