Diagnóstico de la infección por Treponema pallidum en pacientes con sífilis temprana y neurosífilis mediante reacción de la polimerasa en cadena / Laboratory diagnosis of Treponema pallidum infection in patients with early syphilis and neurosyphilis through a PCR-based test

Syphilis is a sexually transmitted disease caused by Treponema pallidum. The diagnosis is based mainly in clinical presentation and non-specific assays. PCR-based diagnosis has been suggested as an attractive alternative method. The aim of this study was the validation of a PCR-based test for the diagnosis of early syphilis (ES) and neurosyphilis (NS). Clinical samples of mucocutaneous lesions and cerebrospinal fluid (CSF) specimens from patients previously diagnosed for ES and NS respectively using an enlarged gold standard, were tested by PCR. The reaction was done using primers targeting the tpN47gene. Twenty out of 21 mucocutaneous samples from patients diagnosed with ES were positive by PCR, with a clinical sensitivity of 95 percent. Four out of 8 CSF samples from patients previously diagnosed with NS were positive by PCR, with a clinical sensitivity of 50 percent. The clinical specificity for both ES and NS was 100 percent. The PCR sensitivity and specificity for mucocutaneous samples allowed us to implement this assay in our laboratory for routine diagnosis. Although the sensitivity of the PCR in CSF was low, it may be useful to support clinical diagnosis.

La sífilis es una enfermedad de transmisión sexual producida por Treponema pallidum, cuyo diagnóstico se realiza presuntivamente basándose en aspectos clínicos y análisis de especificidad limitada. La reacción de la polimerasa en cadena (RPC) ha sido planteada como una alternativa diagnóstica de mayor sensibilidad y especificidad. El objetivo de este trabajo fue validar una RPC para el diagnóstico de sífilis temprana (ST) y neurosífilis (NS). Se utilizaron muestras de lesiones muco-cutáneas y de LCR de pacientes con sospecha de cursar ST y NS respectivamente, previamente diagnosticados, utilizando un estándar de oro ampliado. La RPC fue realizada con partidores dirigidos al gen tpN47. De las 21 muestras de pacientes con ST, la RPC resultó positiva en 20, lo que resulta en una sensibilidad clínica de 95 por ciento. De las 8 muestras de pacientes con NS, la RPC resultó positiva en 4, obteniéndose una sensibilidad clínica de 50 por ciento. La especificidad clínica para ST y NS fue de 100 por ciento. La excelente sensibilidad y especificidad de la RPC para muestras muco-cutáneas permitió la exitosa implementación de este análisis en nuestro laboratorio para el diagnóstico de rutina. Si bien la sensibilidad de la RPC en LCR es baja, es muy útil para apoyar el diagnóstico clínico.

Medición de 17-OH progesterona sanguínea en recién nacidos chilenos: antecedentes para implementar un programa de detección neonatal de hiperplasia suprarrenal congénita / 17 OH progesterone measurement in blood of chilean newborns: as a background to start a congenital adrenal hyperplasia screening program

Background: The early diagnosis and therapy of congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency can prevent adrenal crises and erroneous gender assignment in affected newborns. To achieve this goal neonatal mass-screening programs have been developed, measuring blood 17 alpha-hydroxyprogesterone (17OHP). In Chile there is no experience with this type of screening. Aim: To develop a method for measuring 17OHP in filter paper blood specimens. To obtain reference ranges and determine neonatal 17OHP threshold levels according to gestational age and birth weight. To analyze factors affecting the cost-efficiency ratio and suggest recommendations for the organization of a neonatal screening program for CAH in Chile. Material and methods: Nine hundred twenty two newborns were studied. 17OHP was measured using double antibody radioimmunoassay in filter paper blood samples obtained 48 h after birth. Reference ranges were determined according to gestational age and birth weight and a cutoff point of 25 ng/ml was established. Results: Seventeen newborns had 17OHP over the cutoff value. They were assessed by a pediatric endocrinologist and in none of them, CAH was confirmed. Therefore the false positive rate of the determination was 1.8 percent. Among these newborns with elevated 17OHP, 66 percent had a birth weight below 1.5 kg and 5.8 percent, a birth weight between 1.5 and 2.5 kg. The cost per reported result was US $ l. Timing of the recall was between the 3 and 10 days of life. No newborn missed the follow-up. Discussion: To increase the cost-efficiency ratio of an eventual neonatal screening program, newborns with birth weights below 1.5 kg should be excluded and cutoff points should be defined according to birth weight

Exceso de mineralocorticoides en hipertensos esenciales: enfoque clínico diagnóstico / Evidences for mineralocorticoid excess in essential hypertension: clinic and diagnosis viewpoint

Background: Primary hyperaldosteronism is more frequent among subjects with essential hypertension than previously thought. The prevalence, according to local and international evidence could fluctuate between 9 and 10 percent. Aim: To investigate if subjects with essential hypertension have different aldosterone and renin plasma levels than normotensive subjects. Patients and methods: One hundred twenty five subjects with essential hypertension, not receiving medications for at least two weeks prior to the study and 168 age and sex matched normotensive controls were studied. Blood was drawn between 9 and 10 AM during a sodium free diet to determine plasma aldosterone, plasma renin activity and potassium. Results: Plasma aldosterone was higher in hypertensive subjects than controls (11.6 ñ 7.6 and 9.9 ñ 5.1 ng/dl respectively; p=0.04). Plasma renin activity was lower in hypertensives than controls (1.42 ñ 1.28 and 1.88 ñ 1.39 ng/ml/h respectively; p<0.001). Thus, plasma aldosterone/plasma renin activity ratio was higher in hypertensives (13.8 ñ 13.5 and 8.3 ñ 7.8; p<0.001). A pathological ratio was defined as over 25, corresponding to the mean plus two standard deviations of the control group. Primary hyperaldosteronism was found in 5/125 hypertensives (4 percent) and 1/168 normotensive subject (0.6 percent). None had hypokalemia. Conclusions: Subjects with essential hypertension have higher plasma aldosterone and lower plasma renin activity than normal controls. A plasma aldosterone/plasma renin activity over 25 was defined as abnormal

Alta prevalencia de hiperaldosteronismo primario no diagnosticado en hipertensos catalogados como esenciales / High prevalence of undiagnosed primary hyperaldosteronism among patients with essential hypertension

Background: Classically, primary hyperaldosteronism was diagnosed in no more than 1 percent of patients with hypertension, when hypokalemia was used as the screening test. However, numerous patients with primary hyperaldosteronism do not have hypokalemia and the disease remains undiagnosed. Aim: To assess the prevalence of normokalemic primary hyperaldosteronism among patients classified as having essential hypertension. Patients and methods: One hundred hypertensive patients with a blood pressure over 145/95 were studied. Plasma aldosterone and plasma renin activity were measured in all. A primary hyperaldosteronism was diagnosed when high aldosterone levels (over 16 ng/dl) and low plasma renin activity (below 0.5 ng/ml/h) coexisted in two blood tests or the aldosterone/plasma renin activity ratio was over 50. A probable primary hyperaldosteronism was diagnosed when the ratio was between 25 and 50 and these patients were subjected to a Fludrocortisone test to confirm the diagnosis. A dexametasone suppression test was done to discard glucocorticoid remediable aldosteronism. An adrenal TAC scan was done to all patients with primary hyperaldosteronism. Results: A diagnosis of primary hyperaldosteronism was reached in ten patients. Seven had elevated aldosterone and low plasma renin activity. In three the diagnosis was confirmed with the fludrocortisone test. All ten patients had normal serum potassium levels. Dexametasone suppression test was positive in three patients, that normalized their blood pressure levels. Adrenal TAC scans showed an adenoma in one patient and hyperplasia in another. Conclusions: Primary hyperaldosteronism is more frequent than previously thought, it is overlooked when hypokalemia is used as the screening test and it can only be diagnosed measuring plasma aldosterone and renin activity

Medición de la actividad de renina plasmática a niveles bajos: optimización del método / Measurement of low levels of plasma renin activity: a methodological improvement

Background: The present method to measure plasma renin activity is cumbersome and imprecise, factors that limit its clinical application. Aim: To assess the importance of blood sampling conditions and the usefulness of increasing incubation time to measure plasma renin activity at low levels. Patients and methods: Twenty hypertensive patients, 14 female, aged 14 to 76 years old, were studied. Two blood samples were obtained after a 10 min rest in the sitting position and after a 30 min rest in supine position. One blood sample, of each condition was sent lo the laboratory at room temperature and the other sample was sent refrigerated. Angiotensin I concentration was determined after 3 h of enzymatic incubation at 37°C and, in subjects with an activity of less than 1 ng/ml/h, after 18 h of incubation. Results: No significant differences in plasma renin activity were observed between the samples obtained with different rest times or different transportation methods. In people with low plasma renin activity, the 18 h enzymatic incubation reduced the lower detection from 0.3 to 0.014 ng/ml/h and the coefficient of variation from 14.4 to 3.2 percent. Conclusions: A simplified blood sampling method does not change plasma renin activity values, and tbe longer enzymatic incubation in people with low plasma renin activity improves both the sensitivitv and accuracy of the determination

Diagnóstico molecular de hiperplasia suprarrenal congénita por déficit de 21-hidroxilasa, variedad perdedora de sal, en población chilena / Molecular diagnosis of salt wasting congenital adrenal hyperplasia, caused by deficit of 21-hydroxylase, in the Chilean population

Twenty five patients with salt wasting congenital adrenal hyperplasia, that had 17-hydroxyprogesterone levels above 30 ng/ml, were studied. In all patients, a polymerase chain reaction (PCR) with selective primers was done with extracted genomic DNA, to amplify the active gene and specific primers for normal or mutated alleles of 50 chromosomes of the 25 patients. The higher frequency affected the ASIn2 in 26 percent of cases, followed by mutations Arg357Trp in 22 percent of cases and Gln319Stop in 12 percent and deletion in 12 percent. The frequent genotypes were homozygosity for ASIn2 (16 percent), homozygosity for Arg357Trp (12 percent) and the homozygote deletion of the gene in 12 percent. The most frequent mechanisms of genetic deficiency of 21-hydroxylase were the mutations ASIn2 Arg357Trp. This type of studies allows prenatal diagnosis and genetics counseling