Bimodal distribution of Trypanosoma cruzi antibody levels in blood donors from a highly endemic area of Argentina: what is the significance of low-reactive samples?

BACKGROUND: Low-level seroreactive donor samples that are inconsistently detected by different Trypanosoma cruzi immunoassays are common, but the population distribution has not been reported in an endemic region. The objective was to understand the distribution of low-level reactive samples using highly sensitive immunoassays and the relationship with epidemiologic evidence of exposure to T. cruzi. STUDY DESIGN AND METHODS: Blood donors (BDs) were recruited in two blood banks located in Chaco province, in northeastern Argentina, from June 2006 to March 2007. Donors completed a Chagas exposure questionnaire and provided blood samples. All samples were tested in parallel with five contemporary and commercially available enzyme immunoassays for T. cruzi and a subgroup by a chemiluminescent assay. RESULTS: Of the 1423 enrolled donors, 304 (21.4%) tested positive on all assays while 93 (6.5%) were reactive on at least one assay (inconclusive). Epidemiologic evidence of exposure to T. cruzi was significantly higher among positive and inconclusive donors compared to seronegative BD (p values range from 0.01 to <0.001 depending on the exposure). Histograms of the signal-to-cutoff values from all positive samples showed clear bimodal distributions for the whole parasite lysate assays, but not for the one recombinant antigen-based assay. Low antibody level responses were present in 30% to 40% of the reactives, depending on the assay. CONCLUSION: The population of individuals exposed to T. cruzi in highly endemic regions has a bimodal distribution of antibody response to the parasite. Although the clinical significance of low-level reactivity is not fully established, these results may reflect evolving seroreversions after spontaneously resolved infections.

Molecular cloning, tissue distribution and daily expression of cry1 and cry2 clock genes in European seabass (Dicentrarchus labrax).

Biological rhythms are driven by circadian oscillators, which are ultimately controlled by the cyclic expression of clock genes. Cryptochromes (CRY), blue light photoreceptors, belong to the negative elements of the transcriptional feedback loop into the molecular clock. This paper describes the cloning and characterization of two cryptochromes (cry1 and 2) in European seabass, which is considered an interesting chronobiology model due to its dual (diurnal/nocturnal) behavior. The cloned cDNA fragments encoded for two proteins of 567 and 668 amino acids, which included the FAD-binding and the DNA-photolyase domains. Moreover, both proteins had a high homology with cryptochrome proteins (Cry) of other teleost fish. These cry1 and 2 genes were expressed in several tissues of seabass (brain, liver, heart, retina, muscle, spleen, gill and intestine). In addition, the daily expression of cry1 was rhythmic in brain, heart and liver with the acrophase around ZT 03:15 h (after the onset of lights). Similarly, the cry2 daily expression was rhythmic in liver, peaking at ZT 03:28 h, whereas in brain the acrophase was at ZT 11:08 h (shortly prior to the offset of lights). These findings provide new elements to help understanding the functioning of the molecular clock of seabass.

Circadian regulation of phosphodiesterase 6 genes in zebrafish differs between cones and rods: Implications for photopic and scotopic vision.

A correlation is known to exist between visual sensitivity and oscillations in red opsinand rhodopsin gene expression in zebrafish, both regulated by the clock gene. This indicates that an endogenous circadian clock regulates behavioural visual sensitivity, apart from the regulation exerted by the pineal organ. However, the specific mechanisms for cones (photopic vision) and rods (scotopic vision) are poorly understood. In this work, we performed gene expression, cosinor and immunohistochemical analyses to investigate other key genes involved in light perception, encoding the different subunits of phosphodiesterase pde6 and transducin GαT, in constant lighting conditions and compared to normal light-dark conditions. We found that cones display prominent circadian oscillations in mRNA levels for the inhibitory subunit gene pde6ha that could contribute to the regulation of photopic sensitivity by preventing overstimulation in photopic conditions. In rods, the mRNA levels of the inhibitory subunit gene pde6ga oscillate under normal conditions and dampen down in constant light but continue oscillating in constant darkness. There is an increase in total relative expression for pde6gb in constant conditions. These observations, together with previous data, suggest a complex regulation of the scotopic sensitivity involving endogenous and non-endogenous components, possibly present also in other teleost species. The GαT genes do not display mRNA oscillations and therefore may not be essential for the circadian regulation of photosensitivity. In summary, our results support different regulation for the zebrafish photopic and scotopic sensitivities and suggest circadian regulation of pde6ha as a key factor regulating photopic sensitivity, while the regulatory mechanisms in rods appear to be more complex.

Behavioral Characterization of dmrt3a Mutant Zebrafish Reveals Crucial Aspects of Vertebrate Locomotion through Phenotypes Related to Acceleration.

Vertebrate locomotion is orchestrated by spinal interneurons making up a central pattern generator. Proper coordination of activity, both within and between segments, is required to generate the desired locomotor output. This coordination is altered during acceleration to ensure the correct recruitment of muscles for the chosen speed. The transcription factor Dmrt3 has been proposed to shape the patterned output at different gaits in horses and mice. Here, we characterized dmrt3a mutant zebrafish, which showed a strong, transient, locomotor phenotype in developing larvae. During beat-and-glide swimming, mutant larvae showed fewer and shorter movements with decreased velocity and acceleration. Developmental compensation likely occurs as the analyzed behaviors did not differ from wild-type at older larval stages. However, analysis of maximum swim speed in juveniles suggests that some defects persist within the mature locomotor network of dmrt3a mutants. Our results reveal the pivotal role Dmrt3 neurons play in shaping the patterned output during acceleration in vertebrates.

WHO comparative evaluation of serologic assays for Chagas disease.

BACKGROUND: Evaluation of commercially available test kits for Chagas disease for use in blood bank screening is difficult due to a lack of large and well-characterized specimen panels. This study presents a collaborative effort of Latin American blood centers and the World Health Organization (WHO) to establish such a panel. STUDY DESIGN: A total of 437 specimens, from 10 countries were collected and sent to the WHO Collaborating Center in São Paulo and used to evaluate 19 screening assays during 2001 through 2005. Specimens were assigned a positive or negative status based on concordant results in at least three of the four confirmatory assays (indirect immunofluorescence, Western blot, radioimmunoprecipitation assay, and recombinant immunoblot). RESULTS: Of the 437 specimens, 168 (39%) were characterized as positive, 262 (61%) were characterized as negative, and 7 (2%) were judged inconclusive and excluded from the analysis. Sensitivity and specificity varied considerably: 88 to 100 and 60 to 100 percent, respectively. Overall, enzyme immunoassays (EIAs) performed better than the other screening assays. Four EIAs had both parameters higher than 99 percent. Of the four confirmatory assays, only the RIPA gave a 100 percent agreement with the final serologic status of the specimens. CONCLUSION: The sensitivities and specificities of at least four of the commercially available EIAs for Chagas disease are probably high enough to justify their use for single-assay screening of blood donations. Our data suggest that the majority of commercially available indirect hemagglutination assays should not be used for blood donor screening and that the RIPA could be considered a gold standard for evaluating the performance of other assays.

Estimation of sensitivity and specificity of several Trypanosoma cruzi antibody assays in blood donors in Argentina.

BACKGROUND: The absence of a gold standard test for Trypanosoma cruzi antibodies represents a problem not only for the evaluation of screening tests, but also for appropriate blood donor counseling. The aim of this study was to estimate the sensitivity and specificity of multiple blood donor screening tests for T. cruzi antibodies in Argentina. STUDY DESIGN AND METHODS: From June 2006 to March 2007 a sample of 1455 blood donors was recruited from two blood banks in Chaco province, an area of Argentina with highly endemic T. cruzi infection. Samples were tested by three epimastigote lysate enzyme immunoassays (EIAs), one recombinant antigen EIA, two indirect hemagglutination assay (IHA) tests, a particle agglutination assay (PA), and a research trans-sialidase inhibition assay (TIA). Sensitivity and specificity were estimated using latent class analysis (LCA). RESULTS: LCA estimated the consensus prevalence of T. cruzi infection at 24.5%. Interassay correlation was higher among the four EIA tests and TIA compared to IHA tests. Assay sensitivities varied from 96 to 99.7 for different EIAs, 91% for TIA, 84% for PA, and 66 to 74% for IHA tests. Relative to the LCA, assay specificities were from 96% to almost 100%. CONCLUSION: Based on the comparison of several tests in a large population from an endemic area for T. cruzi infection, our data showed an adequate sensitivity for EIA tests in contrast to PA and IHA assays. The latter tests should no longer be used for blood donor screening.

Management of hepatocellular carcinoma.

Hepatocellular carcinoma is the fifth most common cancer in the world and is the third cause of cancer-related death with varying prevalence according to endemic risk factors. Despite therapeutic advances, there has not been significant improvement in the overall survival of patients who have hepatocellular cancer in the last 2 decades. Treatment selection should be based on tumor characteristics and the underlying liver disease.

A gas-filled abdominal cyst in an elderly woman: A giant colonic diverticulum case report.

INTRODUCTION: Giant colonic diverticulum (GCD), a rare complication of the diverticular disease, can present with a wide range of nonspecific symptoms as abdominal pain and bowel obstruction. Its diagnosis represents a challenge that mainly depends on imaging findings. PRESENTATION OF CASE: We report the case of a 79 year-old female patient that came to our emergency department complaining of 5-day history of hypogastric pain and constipation. Physical examination reveled a 15cm hypogastric round, tender and tympanic mass. Enhanced abdominal CT scan showed a large air-filled cyst adjacent to a diverticular sigmoid colon without evidence of intra-abdominal free air or fluid. Based on the radiological features, GCD was suspected and surgical treatment performed. The mass and the sigmoid colon were resected. The postoperative course was uneventful. Histopathology confirmed the preoperative diagnosis. DISCUSSION: GCD, defined as a diverticulum larger than 4cm, represents a rare complication of the diverticular disease. Usually abdominal X-ray and computed tomography (CT) scan show a gas-filled structure, sometimes communicating with the adjacent colon. GCD resection and segmental colectomy are strongly recommended even in asymptomatic cases due to the high incidence and severity of complications. CONCLUSION: Because of its rarity and variable and non-specific clinical presentation, the diagnosis of GCD depends mainly on imaging findings. The gold standard treatment is surgical resection of the GCD and the compromised colon with primary anastomosis when possible.

Antibodies to hepatitis C and other viral markersin multi-transfused patients from Argentina.

BACKGROUND: The knowledge of transfusion-transmitted viral infections in Argentina is scarce. A regional study organized by the Pan American Health Organization let us asses the current status. OBJECTIVES: To estimate the prevalence of HCV, HBV and HIV infection in a population of multi-transfused Argentinean patients. STUDY DESIGN: Multi-center, cross sectional study of 504 patients from national referral institutions in Buenos Aires, who had received more than ten units of blood products in more than two occasions. Demographic and clinical data were collected using a standardized questionnaire. Blood samples were analyzed for a-HCV, a-HIV, HBsAg and a-HBcore. RESULTS: Patients belonged to five diagnostic categories: onco-hematology (309; 61.3%); hemophilia (96; 19%); acute blood loss (54; 10.7% ); hemoglobinopathies (35; 6.9%); and hemodialysis (5; 1% ); five patients (1%) had two of the previous conditions. Overall prevalence rates of viral markers were: a-HCV 9.3% (CI(95%): 6.7-12.0); a-HBcore 4.8% (CI(95): 2.8-6.7); a-HIV 1.2% (CI(95%): 0.14-2.2) and HBsAg 0.20%(CI(95%): 0.2-0.59). The highest prevalence rates were found among patients living with hemophilia (PLH). There was a significant statistical association (p < 10(-5), OR =78.8 [29.7-209.7]) between a-HCV infection and having been transfused before 1993, when screening blood donors for a-HCV became mandatory in our country. The subpopulation of patients exposed to transfusion before 1993 was conformed mostly by PLH (70.9%) and hemoglobinopathies (18.6%). In this subpopulation, we found a significant association (p < 10(Dot;), OR -40 [5.68-281.66]) between years of exposure to transfusion and a-HCV among the patients under the median age (21.95 years old); however, there was no association for those above the median age (p=0.111). CONCLUSION: a-HCV was found to be the most prevalent infection in the multi-transfused patient population under study. Most infected individuals were PLH, transfused before 1993. This study will provide support for further research aimed at improving blood safety in Argentina.

Relationship between the Arg72Pro polymorphism of p53 and outcome for patients with traumatic brain injury.

OBJECTIVE: To determine the relationship between the Arg72Pro polymorphism of p53 and the outcome after traumatic brain injury (TBI) in humans. DESIGN AND SETTING: A prospective study was carried out in a ten-bed surgical intensive care unit (SICU) of a university hospital. PATIENTS: The study included 90 caucasian patients who had experienced a severe TBI within the previous 24 h. The exclusion criterion was a previous deficit in the central nervous system. MEASUREMENTS AND RESULTS: The main recorded outcomes were values on the Glasgow Outcome Scale (GOS) at discharge from the SICU (GOS-0) and 6 months later (GOS-6). A blood sample was taken from each patient and all samples were analyzed by an allele-specific polymerase chain reaction for detection of Arg72Pro polymorphism of p53. These polymorphisms were tested for their association with values of GOS-0 and GOS-6. A two-tailed value of p<0.05 was considered statistically significant. The frequency of the argine/argine (Arg/Arg) genotype was greater among the patients who had a bad outcome at GOS-0 (69 vs 31% in the bad-outcome group; p=0.029). A multiple logistic regression analysis showed that patients with the Arg/Arg variation had a 2.9-fold greater risk of having a bad outcome at discharge from the SICU (95% confidence interval, 1.05-8.31; p=0.039 ). There was no similar relationship with respect to GOS-6. CONCLUSION: The Arg/Arg genotype of the Arg72Pro polymorphism in p53 is associated with increased likelihood of a bad outcome at discharge from the SICU.

[Epidermoid carcinoma of gallbladder: analysis of our casuistic]. / Carcinoma epidermoide de vesícula: análisis de nuestra casuística.

BACKGROUND: The most common type of gallbladder cancer is the adenocarcinoma. The squamous cell carcinoma represents only a 0-12% of all gallbladder tumors. METHODS: 124 cases of malignant neoplasias of the gallbladder were diagnosed during the last 33 years in the Department of Surgery of our hospital. From these cases, 5 were squamous cell carcinomas, representing 2.41% of our series. CASE REPORT: The ratio female: male was 1.5:1, and the mean age was 50.2 years. Liver involvement was observed in 4 of 5 patients at the moment of diagnosis. Four patients underwent surgery and one received palliative treatment with percutaneous bile dreinage. The mean survival was 14.5 months. CONCLUSION: The tumor extention at the time of diagnosis is generally advanced and the outcome is not promising in this kind of gallbladder cancer.

[Beta thalassemia major in Argentina]. / Beta talasemia mayor en la Argentina.

An analysis of beta thalassemia major patients seen at Hospital Juan P. Garrahan was carried out in order to determine the characteristics and outcome of the population. From August 1987 to July 2000, 45 patients were admitted (27 males-18 females). The most common beta globin gene defects were C-39 (30.7%); IVS-I nt 110 (20%); IVS-I nt 6 (13.3%); IVS-I nt 1(4%). alpha globin genes were normal in 42 patients, 1 patient had triplicate and cuadriplicate alpha globin genes and 2 patients were not analyzed. Six patients of 5 families were heterozygous for -158G gamma mutation. Allogeneic stem cell transplantation was performed in 7 patients, with an identical sibling. Transfusion-related infections and alloantibodies were detected in 6.7% patients. Growth assessment showed no significant difference in the stature of girls compared to the reference population, but 5 boys had short stature. There is a tendency to short trunk. Growth velocity was normal at prepubertal age. No X-ray lesions related to desferrioxamine were observed. Delayed puberty and hypogonadotropic hypogonadism were found in 35.7% and abnormalities in GH/IGF-I axis in 12.5% of the patients. Impaired glucose tolerance was found in 2 patients. No patient developed diabetes mellitus, thyroid or adrenal insufficiency. One patient had cardiac complications. Forty-two patients are alive and 3 died (cardiac failure 1, central nervous system bleeding 1, sepsis 1). We conclude that beta thalassemia major, originated mainly from Italian immigrants, has a cumbersome treatment and is severely hindered by the lack of adequate economic resources in our patients.

Neuroprotective effects of human umbilical cord mesenchymal stromal cells in an immunocompetent animal model of Parkinson's disease.

Microglial activation in the substantia nigra (SN) is a ubiquitous feature in PD which could mediate toxic effects. Human mesenchymal stromal cells (hMSCs) possess immunomodulatory properties. We evaluated whether the transplantation of hMSCs obtained from umbilical cord had a neuroprotective effect in a not-immunosuppressed rat Parkinson's disease (PD) model. Rats receiving hMSCs in the SN displayed significant preservation in the number of dopaminergic neurons in the SN at 21 days after lesion and an improved performance in behavioral tests compared to control rats. However, no differences in any inflammatory parameter tested were found. These results suggest that grafted hMSCs exert neuroprotection but not neuromodulatory effects on degenerating dopaminergic neurons.

Daily rhythms of clock gene expression, glycaemia and digestive physiology in diurnal/nocturnal European seabass.

Seabass is a fish species with dual (diurnal/nocturnal) feeding behavior, although little is known about changes in its molecular clock, physiology and metabolism linked to this dual behavior. In the research described here possible differences in clock gene expression in central (brain) and peripheral (liver) oscillators, and in physiology (blood glucose and amylase activity in mid-intestine) were studied in seabass with diurnal or nocturnal self-feeding patterns under LD 12:12h (light:dark) (lights on=Zeitgeber Time (ZT) 00:00h). The results revealed that per1 expression in brain shows daily rhythmicity with the acrophase (Φ) around the lights offset (ZT 12:00h, Cosinor, p<0.01) in both diurnal and nocturnal seabass. In liver, per 1 daily levels of expression were higher in diurnal fish (univariate GML, p<0.02). Daily blood glucose variations were observed in both groups (ANOVA I, p<0.01), with higher glucose levels occurring at night in nocturnal as well as in diurnal fish, although only diurnal seabass displayed a significant daily rhythm (Φ=ZT 16:52h, Cosinor, p<0.02). The highest values of amylase activity coincided with the feeding-phase of fish; that is, in nocturnal seabass the maximum was reached at ZT 18:00h (ANOVA I, p<0.01), whereas in diurnal seabass the Φ was ZT 03:39h (Cosinor, p<0.02). In short, our findings indicated that the feeding rhythm (diurnal vs. nocturnal) strongly influenced the daily patterns of digestive function and clock gene expression in the liver (feeding-entrained clock), but not in the brain (light-entrained clock).

Circadian rhythms of self-feeding and locomotor activity in zebrafish (Danio Rerio).

To investigate daily feeding rhythms in zebrafish, the authors have developed a new self-feeding system with an infrared photocell acting as a food-demand sensor, which lets small-size fish such as zebrafish trigger a self-feeder. In this paper, the authors used eight groups of 20 fish. Locomotor activity rhythms were also investigated by means of infrared sensors. Under a 12 h:12 h light (L)-dark (D) cycle, zebrafish showed a clear nocturnal feeding pattern (88.0% of the total daily food-demands occurring in the dark phase), concentrated during the last 4 h of the dark phase. In contrast, locomotor activity was mostly diurnal (88.2% of total daily activity occurring in the light phase). Moreover, both feeding and locomotor rhythms were endogenously driven, as they persisted under free-running conditions. The average period length (τ) of the locomotor and feeding rhythms was shorter (τ = 22.9 h) and longer (τ = 24.6 h) than 24 h, respectively. During the time that food availability was restricted, fish could only feed during ZT0-ZT12 or ZT12-ZT16. This resulted in feeding activity being significantly modified according to feeding time, whereas the locomotor activity pattern remained synchronized to the LD cycle and did not change during this trial. These findings revealed an independent phasing between locomotor and feeding activities (which were mostly nocturnal or diurnal, respectively), thus supporting the concept of multioscillatory control of circadian rhythmicity in zebrafish.

Clarithromycin-nifedipine interaction as possible cause of vasodilatory shock.

OBJECTIVE: To report a case of vasodilatory shock possibly resulting from a clarithromycin-nifedipine interaction. CASE SUMMARY: A 77-year-old male with uncontrollable hypertension developed shock, heart block, and multiorgan failure 2 days after clarithromycin was added to his antihypertensive treatment (nifedipine, captopril, doxazosin). Invasive monitoring revealed hyperdynamic shock with decreased systemic vascular resistances. DISCUSSION: Nifedipine is metabolized by isoenzyme CYP3A4. This metabolic pathway is inhibited by clarithromycin, thus potentially increasing the plasma nifedipine concentration, which may lead to excessive calcium-channel blocker effects. Clinical manifestations of excessive calcium-channel blockade comprise hypotension or vasodilatory shock and heart block, as in our case. An objective causality assessment revealed that this drug interaction was the possible cause of these adverse effects. Because of an initial diagnosis of septic shock, suspicion of this entity was delayed and specific diagnostic and treatment were not possible. CONCLUSIONS: This life-threatening clinical picture, including shock and heart block, was possibly the result of a pharmacokinetic interaction between clarithromycin and nifedipine.

[External performance evaluation of screening in blood banks in Argentina: results and strategies for improvement]. / Evaluación externa del desempeño en el tamizaje de bancos de sangre en Argentina: resultados y estrategias para mejorarlo.

OBJECTIVES: Because there was no program for the external evaluation of the quality of the screening for transfusion-transmitted infections (TTIs) in blood banks in Argentina, in 1999 the Hemotherapy Service of Garrahan Hospital, in Buenos Aires, launched an ongoing external evaluation program, with the support of the Pan American Health Organization and the Argentine Hemotherapy and Immunohematology Association. METHODS: A panel of 12 samples that were reactive to all the markers screened for in Argentine blood banks was distributed. The panel was delivered to 52 laboratories in 1999, 102 laboratories in 2000, and 118 laboratories in 2001. The participating laboratories were classified into one of four categories according to their performance: A: 0 or 1 false positive (FP) results; B: 2 or 3 FPs; C: 4 or more FPs; and D: at least 1 false negative result. Workshops were held with the participants in order to jointly analyze the results. RESULTS: Out of the laboratories that received the panel of samples, the percentage of laboratories that sent in their results was 92.3% in 1999, 92.2% in 2000, and 83.9% in 2001. These response levels demonstrate the interest in evaluation activities. However, the annual average percentage of the laboratories that received a "D" classification was very high (30%), which indicates that there are problems in the performance of TTI screening. This poor showing could be related to the difficulty in taking corrective measures throughout the system, given the large number of laboratories involved. CONCLUSIONS: Evaluations of this type should be included in Argentina's National Blood Program as a tool for the ongoing assessment of the blood banking situation, for decision-making, and for monitoring the quality of screening.