Comparison of real-time PCR protocols for differential laboratory diagnosis of amebiasis

Specific identification of Entamoeba spp. in clinical specimens is an important confirmatory diagnostic step in the management of patients who may be infected with Entamoeba histolytica, the species that causes clinical amebiasis. Distinct real-time PCR protocols have recently been published for identification of E. histolytica and differentiation from the morphologically identical nonpathogenic Entamoeba dispar. In this study, we compared three E. histolytica real-time PCR techniques published by December 2004. The limits of detection and efficiency of each real-time PCR assay were determined using DNA extracted from stool samples spiked with serially diluted cultured E. histolytica trophozoites. The ability of each assay to correctly distinguish E. histolytica from E. dispar was evaluated with DNA extracted from patients' stools and liver aspirates submitted for confirmatory diagnosis. Real-time PCR allowed quantitative analysis of the spiked stool samples, but major differences in detection limits and assay performance were observed among the evaluated tests. These results illustrate the usefulness of comparative evaluations of diagnostic assays.     

Nucleotide sequence and phylogenetic analyses of the DNA polymerase gene of Anticarsia gemmatalis nucleopolyhedrovirus

The DNA polymerase from Anticarsia gemmatalis nucleopolyhedrovirus (AgMNPV) was identified and sequenced, and its amino acid sequence was compared with other viral DNA polymerases to identify conserved regions and to reconstruct a phylogenetic tree. The sequence analysis of the AgMNPV DNA polymerase gene revealed the presence of a 2976 nucleotides open reading frame (ORF) encoding a polypeptide of 991 amino acid residues with a predicted molecular mass of 114.7 kDa. Among the baculovirus DNA polymerase genes identified to date, the AgMNPV DNA polymerase gene shared maximum amino acid sequence identity with the DNA polymerase gene of Choristoneura fumiferana nucleopolyhedrovirus defective strain (CfDEFNPV) (94%). The alignment of 140 virus sequences, 23 of them from baculovirus, showed that, of the 10 conserved regions identified, 5 are exclusive to baculoviruses (R1, R5, R9, R6 and R10), only 2 of them (R6 and R10) previously described as such in the literature. Our analysis, based on their positions in the AgMNPV DNA polymerase model, suggests that R9 and R10 could interact with DNA. Phylogenetic analysis of DNA polymerase sequences places the enzyme from AgMNPV within the cluster containing the polymerases of Group I Nucleopolyhedrovirus and suggests that the AgMNPV DNA polymerase is more closely related to that of CfDEFNPV than to those of other baculoviruses.     

Vestibular–Podokinetic interaction without vestibular perception

After prolonged stepping in place relative to space over the center of a rotating turntable, blindfolded subjects cannot step on the stationary platform without unknowingly turning themselves relative to space, a phenomenon termed podokinetic after rotation (PKAR). We asked what role the resulting vestibular stimulation might play in the expression of this form of PKAR. A method of servo-stabilizing the body relative to space during PKAR was devised to compare PKAR expression with and without significant vestibular stimulation. Simulated estimates of average central vestibular response profiles were obtained by passing the averaged unidirectional body angular velocity profiles relative to space through a first-order model of the canal system (τ=15 s). Such simulation results suggested that during normal PKAR performed on a stationary platform, the average central vestibular response would likely rise to peak levels equivalent to that induced by about 9°/s within the frequency range of natural head movement. In the servo-stabilized condition, the simulated response was reduced to insignificant levels. Experimental results demonstrated that in the unstabilized condition the rate of rise of PKAR angular velocity was roughly four times slower than in the stabilized condition. We conclude that the normal expression of PKAR conducted on a stationary platform tends to be substantially slowed by interaction with an unperceived vestibular response.     

Early phase of spatial mismatch negativity is localized to a posterior “where” auditory pathway

The auditory mismatch negativity (MMN) is an event-related potential that reflects early processing of changes in acoustic stimulus features. Although the MMN has been well characterized by previous work, the number, roles, and anatomical locations of its cortical generators remain unresolved. Here, we report that the MMN elicited by occasional deviations in sound location is comprised of two temporally and anatomically distinct phases: an early phase with a generator posterior to auditory cortex and contralateral to the deviant stimulus, and a later phase with generators that are more frontal and bilaterally symmetric. The posterior location of the early-phase generator suggests the engagement of neurons within a putative “where” pathway for processing spatial auditory information.     

Novel L2HGDH mutations in 21 patients with L-2-hydroxyglutaric aciduria of Portuguese origin

We studied 21 patients, from 18 families, with L-2-hydroxyglutaric aciduria (L-2-HGA), a rare neurometabolic disorder with a homogeneous presentation: progressive neurodegeneration with extrapyramidal and cerebellar signs, seizures, and subcortical leukoencephalopathy. Increased levels of L-2-hydroxyglutaric acid in body fluids proved the diagnosis of L-2-HGA in all 21 patients. We analyzed the L-2-HGA gene (L2HGDH), recently found to be mutated in consanguineous families with L-2-HGA, and identified seven novel mutations in 15 families. Three mutations appeared to be particularly prevalent in this Portuguese panel: a frameshift mutation (c.529delC) was detected in 12 out of 30 mutant alleles (40%), a nonsense mutation (c.208C>T; p.Arg70X) in 7/30 alleles (23%), and a missense mutation (c.293A>G; p.His98Arg) in four out of 30 mutant alleles (13%), suggesting that common origin may exist. Furthermore, two novel missense (c.169G>A; p.Gly57Arg, c.1301A>C; p.His434Pro) and two splice error (c.257-2A>G, c.907-2A>G) mutations were found. All the mutations presumably lead to loss-of-function with no relationship between clinical signs, progression of the disease, levels of L-2-HGA and site of the mutation. In the three remaining families, no pathogenic mutations in the L-2-HGA were found, which suggests either alterations in regulatory regions of the gene or of its intervening sequences, compound heterozygosity for large genomic deletion and, or further genetic heterogeneity.     

Kallmann syndrome: 14 novel mutations in KAL1 and FGFR1 (KAL2)

Kallmann syndrome (KAL) combines hypogonadotropic hypogonadism and anosmia. Hypogonadism is due to Gonadotropin Releasing Hormone (GnRH) deficiency and anosmia is related to hypoplasia of the olfactory bulbs. Occasional symptoms include renal agenesis, bimanual synkinesia, cleft lip palate, dental agenesis. KAL is genetically heterogeneous and two genes have so far been identified, namely KAL1 (Xp22.3) and FGFR1/KAL2 (8p12), which underlie the X chromosome‐linked form and an autosomal dominant form of the disease, respectively. We studied a cohort of 98 unrelated Caucasian KAL patients. We identified KAL1 mutations in 14 patients, of which 7 (c.3G>A (p.M1?), g.IVS1+1G>T, c.570_571insA (p.R191fsX14), c.784G>C (p.R262P), c.958G>T (p.E320X), c.1651_1654delinsAGCT (p.P551_E552delinsSX), c.1711T>A (p.W571R)) have not been previously reported. In addition, we found FGFR1 mutations in 7 patients, namely c.303G>A (p.V102I), C.385A>C (p.D129A), c.810G>A (p.V273M), c.1093_1094delAG (p.R365fsX41), c.1561G>A (p.A520T), c.1836_1837insT (p.Y613fsX42), c.2190C>G (p.Y730X), all of which were novel mutations. In this study, unilateral renal agenesis and bimanual synkinesia were exclusively found associated with KAL1mutations, cleft palate and dental agenesia with FGFR1mutations. © 2004 Wiley‐Liss, Inc.     

Factors associated with unintended pregnancy in Brazil: cross-sectional results from the Birth in Brazil National Survey, 2011/2012

Background

Unintended pregnancy, a pregnancy that have been either unwanted or mistimed, is a serious public health issue in Brazil. It is reported for more than half of women who gave birth in the country, but the characteristics of women who conceive unintentionally are rarely documented. The aim of this study is to analyse the prevalence and the association between unintended pregnancy and a set of sociodemographic characteristics, individual-level variables and history of obstetric outcomes.

Methods

Birth in Brazil is a cross-sectional study with countrywide representation that interviewed 23,894 women after birth. The information about intendedness of pregnancy was obtained after birth at the hospital and classified into three categories: intended, mistimed or unwanted. Multinomial regression analysis was used to estimate the associations between intendedness of a pregnancy, and sociodemographic and obstetric variables, calculating odds ratios and 95 % confidence intervals. All significant variables in the bivariate analysis were included in the multinomial multivariate model and the final model retaining variables that remained significant at the 5 % level.

Results

Unintended pregnancy was reported by 55.4 % of postpartum women. The following variables maintained positive and significant statistical associations with mistimed pregnancy: maternal age?<?20 years (OR?=?1.89, 95 % CI: 1.68–2.14); brown (OR?=?1.15, 95 % CI: 1.04–1.27) or yellow skin color (OR?=?1.56, 95 % CI: 1.05–2.32); having no partner (OR?=?2.32, 95 % CI: 1.99–2.71); having no paid job (OR?=?1.15, 95 % CI: 1.04–1.27); alcohol abuse with risk of alcoholism (OR?=?1.25, 95 % CI: 1.04–1.50) and having had three or more births (OR?=?2.01, 95 % CI: 1.63–2.47). The same factors were associated with unwanted pregnancy, though the strength of the associations was generally stronger. Women with three or more births were 14 times more likely to have an unwanted pregnancy, and complication in the previous pregnancies and preterm birth were 40 % and 19 % higher, respectively. Previous neonatal death was a protective factor for both mistimed (OR?=?0.61, 95 % CI: 0.44–0.85) and unwanted pregnancy (OR?=?0.44, 95 % CI: 0.34–0.57).

Conclusions

This study confirms findings from previous research about the influence of socioeconomic and individual risk factors on unintended pregnancy. It takes a new approach to the problem by showing the importance of previous neonatal death, preterm birth and complication during pregnancy as risk factors for unintended pregnancy.

     

Labor and birth care by nurse with midwifery skills in Brazil

Background

The participation of nurses and midwives in vaginal birth care is limited in Brazil, and there are no national data regarding their involvement. The goal was to describe the participation of nurses and nurse-midwives in childbirth care in Brazil in the years 2011 and 2012, and to analyze the association between hospitals with nurses and nurse-midwives in labor and birth care and the use of good practices, and their influence in the reduction of unnecessary interventions, including cesarean sections.

Methods

Birth in Brazil is a national, population-based study consisting of 23,894 postpartum women, carried out in the period between February 2011 and October 2012, in 266 healthcare settings. The study included all vaginal births involving physicians or nurses/nurse-midwives. A logistic regression model was used to examine the association between the implementation of good practices and suitable interventions during labor and birth, and whether care was a physician or a nurse/nurse-midwife led care. We developed another model to assess the association between the use of obstetric interventions during labor and birth to the personnel responsible for the care of the patient, comparing hospitals with decisions revolving exclusively around a physician to those that also included nurses/nurse-midwives as responsible for vaginal births.

Results

16.2 % of vaginal births were assisted by a nurse/nurse-midwife. Good practices were significantly more frequent in those births assisted by nurses/nurse-midwives (ad lib. diet, mobility during labor, non-pharmacological means of pain relief, and use of a partograph), while some interventions were less frequently used (anesthesia, lithotomy position, uterine fundal pressure and episiotomy). In maternity wards that included a nurse/nurse-midwife in labour and birth care, the incidence of cesarean section was lower.

Conclusions

The results of this study illustrate the potential benefit of collaborative work between physicians and nurses/nurse-midwives in labor and birth care. The adoption of good practices in managing labor and birth could be the first step toward more effective obstetric and midwifery care in Brazil. It may be easier to introduce new approaches rather than to eliminate old ones, which may explain why the reduction of unnecessary interventions during labor and birth was less pronounced than the adoption of new practices.