Physician Survey to Determine How Dengue Is Diagnosed,Treated and Reported in Puerto Rico

Dengue is a major cause of morbidity in Puerto Rico and is well-known to its physicians. Early case identification and timely initiation of treatment for patients with severe dengue can reduce medical complications and mortality. To determine clinical management and reporting practices, and assess knowledge of dengue and its management, a survey was sent to 2,512 physicians with a medical license in Puerto Rico. Of the 2,313 physicians who received the survey, 817 (35%) completed the questionnaire. Of the respondents, 708 were currently practicing medicine; 138 were board certified (Group 1), 282 were board eligible (Group 2), and 288 had not finished residency (Group 3). Although respondents clinically diagnosed, on average, 12 cases of dengue in the preceding three months, 31% did not report any suspected cases to public health officials while about half (56%) reported all cases. Overall, 29% of respondents correctly identified early signs of shock and 48% identified severe abdominal pain and persistent vomiting as warning signs for severe dengue with the proportion of correct respondents highest in Group 1. Reportedly about sixty percent (57%) appropriately never give corticosteroids or prophylactic platelet transfusions to dengue patients. One third (30%) of respondents correctly identified administration of intravenous colloid solution as the best treatment option for dengue patients with refractory shock and elevated hematocrit after an initial trial of intravenous crystalloids, and nearly one half (46%) correctly identified administration of a blood transfusion as the best option for dengue patients with refractory shock and decreased hematocrit after a trial of intravenous crystalloids. Even though dengue has been endemic in Puerto Rico for nearly 4 decades, knowledge of dengue management is still limited, compliance with WHO treatment guidelines is suboptimal, and underreporting is significant. These findings were used to design a post graduate training course to improve the clinical management of dengue.     

Recombinant Paracoccin Reproduces the Biological Properties of the Native Protein and Induces Protective Th1 Immunity against Paracoccidioides brasiliensis Infection

Background

Paracoccin is a dual-function protein of the yeast Paracoccidioides brasiliensis that has lectin properties and N-acetylglucosaminidase activities. Proteomic analysis of a paracoccin preparation from P. brasiliensis revealed that the sequence matched that of the hypothetical protein encoded by PADG-3347 of isolate Pb-18, with a polypeptide sequence similar to the family 18 endochitinases. These endochitinases are multi-functional proteins, with distinct lectin and enzymatic domains.

Methodology/principal findings

The multi-exon assembly and the largest exon of the predicted ORF (PADG-3347), was cloned and expressed in Escherichia coli cells, and the features of the recombinant proteins were compared to those of the native paracoccin. The multi-exon protein was also used for protection assays in a mouse model of paracoccidioidomycosis.

Conclusions/Significance

Our results showed that the recombinant protein reproduced the biological properties described for the native protein—including binding to laminin in a manner that is dependent on carbohydrate recognition—showed N-acetylglucosaminidase activity, and stimulated murine peritoneal macrophages to produce high levels of TNF-α and nitric oxide. Considering the immunomodulatory potential of glycan-binding proteins, we also investigated whether prophylactic administration of recombinant paracoccin affected the course of experimental paracoccidioidomycosis in mice. In comparison to animals injected with vehicle (controls), mice treated with recombinant paracoccin displayed lower pulmonary fungal burdens and reduced pulmonary granulomas. These protective effects were associated with augmented pulmonary levels of IL-12 and IFN-γ. We also observed that injection of paracoccin three days before challenge was the most efficient administration protocol, as the induced Th1 immunity was balanced by high levels of pulmonary IL-10, which may prevent the tissue damage caused by exacerbated inflammation. The results indicated that paracoccin is the protein encoded by PADG-3347, and we propose that this gene and homologous proteins in other P. brasiliensis strains be called paracoccin. We also concluded that recombinant paracoccin confers resistance to murine P. brasiliensis infection by exerting immunomodulatory effects.     

Titi monkeys (Callicebus spp., Atelidae: Platyrrhini) in the Brazilian state of Rondônia

Five species of titi monkey (Callicebus brunneus, Callicebus caligatus Callicebus cinerascens, Callicebus donacophilus, andCallicebus moloch) were recorded in surveys of primate populations at 26 sites throughout the Brazilian state of Rondônia. The distribution of the two species,C. cinerascens andC. donacophilus (recorded in the state for the first time), appeared to be related to that of non-forest ecosystems, the former in thecerrado woodlands, and the latter in gallery forests of the Guaporé grasslands. The results of the surveys also indicate thatC. brunneus has a more restricted distribution in southern Rondônia than was previously thought, whereasC. moloch is more widespread. However, the ecological factors that determine species distribution in the south of the state remain unclear on the basis of the available data. All species were observed in small social groups of no more than five individuals, which are typical of the genus, generally in the middle and lower forest strate.     

Ligninolytic activity of tropical rainforest basidiomycetes

A total of 116 strains of Brazilian tropical rainforest basidiomycetes were evaluated in terms of their ability to oxidize the dye rhemazol brilliant blue R (RBBR) and guaiacol. Laccase and peroxidase activities were detected by the drop test using solutions of α-naphthol and pyrogallol, respectively. RBBR and guaiacol oxidation occurred in 96.6 and 87.1% of the strains tested, respectively. One hundred strains oxidized both substrates. In the drop test, most strains presented laccase (96.6%) and peroxidase (92.2%) activity. The quick screening method used here can be useful to identify ligninolytic fungal strains to be used in various biotechnological applications. This revised version was published online in November 2006 with corrections to the Cover Date.     

Direct measurement of spatial autocorrelation at the community level in four plant communities

Abstract. Four sites were sampled to determine spatial autocorrelation in vegetation at the community level. All were in western New Zealand, but on different substrates and of different physiognomy: a terrace forest, a floodplain forest, a mire and the middle of a logging road. In ‘dissimograms’the four communities all showed steady increases in dissimilarity with distance, but with shoulders in the curve for some sites, which could be related to plant morphology. Most of the increase in dissimilarity occurred over very short distances: less than 0.5 m in the forests, less than 1 m in the mire and less than 2 m in the road. Separate analyses of the woody and herbaceous guilds in the floodplain forest showed that herbaceous dissimilarities remained low at distances up to 20 m, probably because of clonal structure in some species. The mire showed low overall dissimilarity, which is attributed to the uniform substrate and the small species pool. Simulations showed that the approach is capable of indicating structure when it is present. Although the dissimogram was clearest when analysing a simulated grid of patches, other types of simulated patchiness showed dissimograms that were clearly distinguishable from those obtained from the vegetation studied. The almost continuous rise in dissimilarity with distance found in the four sites offers no support to the Hierarchy theory, fitting much more closely the alternative Continuum theory.     

Prevalence of Sleep Apnea and Electrocardiographic Disturbances in Morbidly Obese Patients

Objective: To determine the prevalence of sleep apnea in morbidly obese patients and its relationship with cardiac arrhythmias. Research Methods and Procedures: Fifty‐two consecutive morbidly obese (body mass index ≥ 40 kg/m²) outpatients from the Obesity Clinic of the National Institute of Nutrition Salvador Zubirán underwent two nights of polysomnography with standard laboratory techniques. Electrocardiographic polysomnography signals (Lead II) were evaluated by two experienced cardiologists, and sleep complaints were measured with a standard sleep questionnaire (Sleep Disorders Questionnaire). In order to make comparisons between groups with different severities of sleep‐disordered breathing, we classified the patients in four groups using the apnea‐hypopnea index (AHI): Group 1, AHI 5 < 15 (n = 10); Group 2, AHI 15 < 30 (n = 10); Group 3, AHI 30 < 65 (n = 14); Group 4, AHI ≥ 65 (n = 17). Results: A wide range of sleep‐disordered breathing, ranging from AHI of 2.5 to 128.9 was found. Ninety‐eight percent of the sample (n = 51) had an AHI ≥ 5 (mean = 51 ± 37), and 33% had severe sleep apnea with AHI ≥ 65 with a mean nocturnal desaturation time of <65% over 135 minutes. Electrocardiographic abnormalities were present in 31% of the patients. Cardiac rhythm alterations showed an association with the level of sleep‐disordered breathing and oxygen desaturation. Discussion: We conclude that there is a high prevalence of sleep apnea in morbidly obese patients and that the risk for cardiac arrhythmias increases in this population in the presence of a severe sleep apnea (AHI ≥ 65) with severe oxygen desaturation (Sao ₂ ≤ 65%).     

Bacterial Activity in the Rhizosphere Analyzed at the Single-Cell Level by Monitoring Ribosome Contents and Synthesis Rates

The growth activity of Pseudomonas putida cells colonizing the rhizosphere of barley seedlings was estimated at the single-cell level by monitoring ribosomal contents and synthesis rates. Ribosomal synthesis was monitored by using a system comprising a fusion of the ribosomal Escherichia coli rrnBP1 promoter to a gene encoding an unstable variant of the green fluorescent protein (Gfp). Gfp expression in a P. putida strain carrying this system inserted into the chromosome was strongly dependent on the growth phase and growth rate of the strain, and cells growing exponentially at rates of ≥0.17 h⁻¹ emitted growth rate-dependent green fluorescence detectable at the single-cell level. The single-cell ribosomal contents were very heterogeneous, as determined by quantitative hybridization with fluorescently labeled rRNA probes in P. putida cells extracted from the rhizosphere of 1-day-old barley seedlings grown under sterile conditions. After this, cells extracted from the root system had ribosomal contents similar to those found in starved cells. There was a significant decrease in the ribosomal content of P. putida cells when bacteria were introduced into nonsterile bulk or rhizosphere soil, and the Gfp monitoring system was not induced in cells extracted from either of the two soil systems. The monitoring system used permitted nondestructive in situ detection of fast-growing bacterial microcolonies on the sloughing root sheath cells of 1- and 2-day-old barley seedlings grown under sterile conditions, which demonstrated that it may be possible to use the unstable Gfp marker for studies of transient gene expression in plant-microbe systems.     

An old drug and different ways to treat cutaneous leishmaniasis: Intralesional and intramuscular meglumine antimoniate in a reference center,Rio de Janeiro,Brazil

BackgroundTreatment of cutaneous leishmaniasis (CL) remains challenging since the drugs currently used are quite toxic, thus contributing to lethality unrelated to the disease itself but to adverse events (AE). The main objective was to evaluate different treatment regimens with meglumine antimoniate (MA), in a reference center in Rio de Janeiro, Brazil.MethodologyA historical cohort of 592 patients that underwent physical and laboratory examination were enrolled between 2000 and 2017. The outcome measures of effectiveness were epithelialization and complete healing of cutaneous lesions. AE were graded using a standardized scale. Three groups were evaluated: Standard regimen (SR): intramuscular (IM) MA 10–20 mg Sb⁵⁺/kg/day during 20 days (n = 46); Alternative regimen (AR): IM MA 5 mg Sb⁵⁺/kg/day during 30 days (n = 456); Intralesional route (IL): MA infiltration in the lesion(s) through subcutaneous injections (n = 90). Statistical analysis was performed through Fisher exact and Pearson Chi-square tests, Kruskal-Wallis, Kaplan-Meier and log-rank tests.ResultsSR, AR and IL showed efficacy of 95.3%, 84.3% and 75.9%, with abandonment rate of 6.5%, 2.4% and 3.4%, respectively. IL patients had more comorbidities (58.9%; p = 0.001), were mostly over 50 years of age (55.6%), and had an evolution time longer than 2 months (65.6%; p = 0.02). Time for epithelialization and complete healing were similar in IL and IM MA groups (p = 0.9 and p = 0.5; respectively). Total AE and moderate to severe AE that frequently led to treatment interruption were more common in SR group, while AR and IL showed less toxicity.Conclusions/SignificanceAR and IL showed less toxicity and may be good options especially in CL cases with comorbidities, although SR treatment was more effective. IL treatment was an effective and safe strategy, and it may be used as first therapy option as well as a rescue scheme in patients initially treated with other drugs.     

PcoB is a defense outer membrane protein that facilitates cellular uptake of copper

Copper (Cu) is one of the most abundant trace metals in all organisms, involved in a plethora of cellular processes. Yet elevated concentrations of the element are harmful, and interestingly prokaryotes are more sensitive for environmental Cu stress than humans. Various transport systems are present to maintain intracellular Cu homeostasis, including the prokaryotic plasmid‐encoded multiprotein pco operon, which is generally assigned as a defense mechanism against elevated Cu concentrations. Here we structurally and functionally characterize the outer membrane component of the Pco system, PcoB, recovering a 2.0 Å structure, revealing a classical β‐barrel architecture. Unexpectedly, we identify a large opening on the extracellular side, linked to a considerably electronegative funnel that becomes narrower towards the periplasm, defining an ion‐conducting pathway as also supported by metal binding quantification via inductively coupled plasma mass spectrometry and molecular dynamics (MD) simulations. However, the structure is partially obstructed towards the periplasmic side, and yet flux is permitted in the presence of a Cu gradient as shown by functional characterization in vitro. Complementary in vivo experiments demonstrate that isolated PcoB confers increased sensitivity towards Cu. Aggregated, our findings indicate that PcoB serves to permit Cu import. Thus, it is possible the Pco system physiologically accumulates Cu in the periplasm as a part of an unorthodox defense mechanism against metal stress. These results point to a previously unrecognized principle of maintaining Cu homeostasis and may as such also assist in the understanding and in efforts towards combatting bacterial infections of Pco‐harboring pathogens.     

Plasmodium berghei leucine-rich repeat protein 1 downregulates protein phosphatase 1 activity and is required for efficient oocyst development

Protein phosphatase 1 (PP1) is a key enzyme for Plasmodium development. However, the detailed mechanisms underlying its regulation remain to be deciphered. Here, we report the functional characterization of the Plasmodium berghei leucine-rich repeat protein 1 (PbLRR1), an orthologue of SDS22, one of the most ancient and conserved PP1 interactors. Our study shows that PbLRR1 is expressed during intra-erythrocytic development of the parasite, and up to the zygote stage in mosquitoes. PbLRR1 can be found in complex with PbPP1 in both asexual and sexual stages and inhibits its phosphatase activity. Genetic analysis demonstrates that PbLRR1 depletion adversely affects the development of oocysts. PbLRR1 interactome analysis associated with phospho-proteomics studies identifies several novel putative PbLRR1/PbPP1 partners. Some of these partners have previously been characterized as essential for the parasite sexual development. Interestingly, and for the first time, Inhibitor 3 (I3), a well-known and direct interactant of Plasmodium PP1, was found to be drastically hypophosphorylated in PbLRR1-depleted parasites. These data, along with the detection of I3 with PP1 in the LRR1 interactome, strongly suggest that the phosphorylation status of PbI3 is under the control of the PP1–LRR1 complex and could contribute (in)directly to oocyst development. This study provides new insights into previously unrecognized PbPP1 fine regulation of Plasmodium oocyst development through its interaction with PbLRR1.