Initial Data on the Molecular Epidemiology of Cryptosporidiosis in Lebanon

Cryptosporidium spp. represent a major public health problem worldwide and infect the gastrointestinal tract of both immunocompetent and immunocompromised persons. The prevalence of these parasites varies by geographic region, and no data are currently available in Lebanon. To promote an understanding of the epidemiology of cryptosporidiosisin this country, the main aim of this study was to determine the prevalence Cryptosporidium in symptomatic hospitalized patients, and to analyze the genetic diversity of the corresponding isolates. Fecal specimens were collected in four hospitals in North Lebanon from 163 patients (77 males and 86 females, ranging in age from 1 to 88 years, with a mean age of 22 years) presenting gastrointestinal disorders during the period July to December 2013. The overall prevalence of Cryptosporidium spp. infection obtained by modified Ziehl-Neelsen staining and/or nested PCR was 11%, and children <5 years old showed a higher rate of Cryptosporidium spp. The PCR products of the 15 positive samples were successfully sequenced. Among them, 10 isolates (66.7%) were identified as C. hominis, while the remaining 5 (33.3%) were identified as C. parvum. After analysis of the gp60 locus, C. hominis IdA19, a rare subtype, was found to be predominant. Two C. parvum subtypes were found: IIaA15G1R1 and IIaA15G2R1. The molecular characterization of Cryptosporidium isolates is an important step in improving our understanding of the epidemiology and transmission of the infection.     

Conantokins derived from the Asprella clade impart conRl-B, an N-methyl d-aspartate receptor antagonist with a unique selectivity profile for NR2B subunits

Using molecular phylogeny has accelerated the discovery of peptidic ligands targeted to ion channels and receptors. One clade of venomous cone snails, Asprella, appears to be significantly enriched in conantokins, antagonists of N-methyl d-aspartate receptors (NMDARs). Here, we describe the characterization of two novel conantokins from Conus rolani, including conantokin conRl-B that has shown an unprecedented selectivity for blocking NMDARs that contain NR2B subunits. ConRl-B shares only some sequence similarity with the most studied NR2B selective conantokin, conG. The divergence between conRl-B and conG in the second inter-Gla loop was used to design analogues for structure-activity studies; the presence of Pro10 was found to be key to the high potency of conRl-B for NR2B, whereas the ε-amino group of Lys8 contributed to discrimination in blocking NR2B- and NR2A-containing NMDARs. In contrast to previous findings for Tyr5 substitutions in other conantokins, conRl-B[L5Y] showed potencies on the four NR2 NMDA receptor subtypes that were similar to those of the native conRl-B. When delivered into the brain, conRl-B was active in suppressing seizures in the model of epilepsy in mice, consistent with NR2B-containing NMDA receptors being potential targets for antiepileptic drugs. Circular dichroism experiments confirmed that the helical conformation of conRl-B is stabilized by divalent metal ions. Given the clinical applications of NMDA antagonists, conRl-B provides a potentially important pharmacological tool for understanding the differential roles of NMDA receptor subtypes in the nervous system. This work shows the effectiveness of coupling molecular phylogeny, chemical synthesis, and pharmacology for discovering new bioactive natural products.     

Costs of twins in free-ranging white-bellied spider monkeys (Ateles belzebuth belzebuth) at Tinigua National Park, Colombia

A female spider monkey (Ateles belzebuth belzebuth) that gave birth to twins was studied during 13 months, and her activity budget and diet were compared to those of females with single offspring and females with no offspring to assess selective pressures that could influence litter size. We recorded qualitative information on the development and social interactions of the twins and three other single infants. Emi, the female that had the twins, had the highest proportion of resting time and the lowest proportion of feeding and moving time compared to other adult females and males. Emi also had the lowest average daily travel distance and relied more heavily on flowers and leaves than other group members who included a higher proportion of fruits in their diets. These results suggest that twins are energetically costly to spider monkeys because of the direct energy allocated into raising and carrying the twins and, also, because these costs have direct effects in the ability of a mother to obtain high-quality resources (e.g., fruits). The twins had a slower rate of physical and social development as well as a smaller body size compared to same-aged singletons by the end of the study. Thus, the extended periods of maternal care and the costs associated with rearing and carrying offspring are some of the factors that influence the evolution and maintenance of a litter size of one in most anthropoid primates.     

Both the stimulation and inhibition of root hair growth induced by extracellular nucleotides in Arabidopsis are mediated by nitric oxide and reactive oxygen species

Root hairs secrete ATP as they grow, and extracellular ATP and ADP can trigger signaling pathways that regulate plant cell growth. In several plant tissues the level of extracellular nucleotides is limited in part by ectoapyrases (ecto-NTPDases), and the growth of these tissues is strongly influenced by their level of ectoapyrase expression. Both chemical inhibition of ectoapyrase activity and suppression of the expression of two ectoapyrase enzymes by RNAi in Arabidopsis resulted in inhibition of root hair growth. As assayed by a dose-response curve, different concentrations of the poorly hydrolysable nucleotides, ATPγS and ADPβS, could either stimulate (at 7.5–25 μM) or inhibit (at ≥ 150 μM) the growth rate of root hairs in less than an hour. Equal amounts of AMPS, used as a control, had no effect on root hair growth. Root hairs of nia1nia2 mutants, which are suppressed in nitric oxide (NO) production, and of atrbohD/F mutants, which are suppressed in the production of H₂O₂, did not show growth responses to applied nucleotides, indicating that the growth changes induced by these nucleotides in wild-type plants were likely transduced via NO and H₂O₂ signals. Consistent with this interpretation, treatment of root hairs with different concentrations of ATPγS induced different accumulations of NO and H₂O₂ in root hair tips. Two mammalian purinoceptor antagonists also blocked the growth responses induced by extracellular nucleotides, suggesting that they were initiated by a receptor-based mechanism.     

Universal definition of loss to follow-up in HIV treatment programs: a statistical analysis of 111 facilities in Africa, Asia, and Latin America

Background

Although patient attrition is recognized as a threat to the long-term success of antiretroviral therapy programs worldwide, there is no universal definition for classifying patients as lost to follow-up (LTFU). We analyzed data from health facilities across Africa, Asia, and Latin America to empirically determine a standard LTFU definition.

Methods and Findings

At a set “status classification” date, patients were categorized as either “active” or “LTFU” according to different intervals from time of last clinic encounter. For each threshold, we looked forward 365 d to assess the performance and accuracy of this initial classification. The best-performing definition for LTFU had the lowest proportion of patients misclassified as active or LTFU. Observational data from 111 health facilities—representing 180,718 patients from 19 countries—were included in this study. In the primary analysis, for which data from all facilities were pooled, an interval of 180 d (95% confidence interval [CI]: 173–181 d) since last patient encounter resulted in the fewest misclassifications (7.7%, 95% CI: 7.6%–7.8%). A secondary analysis that gave equal weight to cohorts and to regions generated a similar result (175 d); however, an alternate approach that used inverse weighting for cohorts based on variance and equal weighting for regions produced a slightly lower summary measure (150 d). When examined at the facility level, the best-performing definition varied from 58 to 383 d (mean = 150 d), but when a standard definition of 180 d was applied to each facility, only slight increases in misclassification (mean = 1.2%, 95% CI: 1.0%–1.5%) were observed. Using this definition, the proportion of patients classified as LTFU by facility ranged from 3.1% to 45.1% (mean = 19.9%, 95% CI: 19.1%–21.7%).

Conclusions

Based on this evaluation, we recommend the adoption of ≥180 d since the last clinic visit as a standard LTFU definition. Such standardization is an important step to understanding the reasons that underlie patient attrition and establishing more reliable and comparable program evaluation worldwide. Please see later in the article for the Editors'' Summary