High-content phenotypic screening identifies novel chemistries that disrupt mosquito activity and development.

New classes of chemistries are needed to control insecticide resistant populations of mosquitoes and prevent transmission of vector-borne diseases (VBDs). Organismal screens of chemical collections have played an important role in the search for new vector insecticides and the identification of active ingredients (AIs) that cause rapid mortality of mosquitoes. Advances in image-based screening offer an opportunity to identify chemistries that operate via novel biochemical modes and investigate the range of phenotypes exhibited by mosquitoes following exposure to lethal and sub-lethal chemical dose. An automated, high throughput phenotypic screen (HTS) employing high-content imaging of first instar (L1) Aedes aegypti larvae was developed to identify chemistries associated with mortality and atypical morphological phenotypes. A pilot screen of the Library of Pharmacologically Active Compounds (LOPAC1280) identified 92 chemistries that disrupted larval activity and development, including conventional insecticides and chemistries known to modulate G protein-coupled receptors (GPCRs) and other molecular targets in mammalian systems. Secondary assay series were used to evaluate a selection of chemistries for impacts on mosquito activity, survival and development. Ritodrine hydrochloride reduced mobility of larvae but had no observable effect on survival and development of mosquitoes. High doses of metergoline suppressed larval activity and sub-lethal dose resulted in pupal mortality. Assay data support the utility of phenotypic screening and diverse entomological end-points for discovery of novel insecticidal chemical scaffolds. The insecticide discovery process must consider how multi-modal efficacy spectra contribute to vector and VBD control.

High alpha-1 antitrypsin clearance predicts severity of gut graft-versus-host disease (GVHD) in children.

The clinical evaluation and management of gut GVHD is a significant challenge in pediatric HSCT. It is often difficult to obtain pathological evidence to confirm diagnosis and/or to determine response to treatment. The severity of the disease itself may not be related to just the classic symptom of diarrhea. The objectives of this study were to prospectively evaluate patients with suspected gut GVHD for PLE as measured by AATC in stools at two different times for each patient and to compare the severity of the PLE with the severity of clinical acute gut GVHD. Thirteen patients were suspected of gut GVHD by clinical criteria (diarrhea > 10 mL/kg/24 h); one patient was excluded for being unable to complete the stool collection. Therefore, 12 patients, 10 boys and two girls, were studied. Median stool volume was 27.5 mL/kg/day (range 10.1-109.0).The median age at BMT was 11.1 yr (range 3.9-17.0 yr). All patients had negative stool electron microscopy for viruses and cultures for C. difficile on their first collection. Nine patients (75%) had two 24-h stool collections performed at a median of eight days apart (range 7-14 days). At the time of the first collection, six patients had ≥ stage 2 acute gut GVHD, and at second collection, four patients had ≥ stage 2 gut GVHD and four collections were of non-diarrheal stool (hence treatment response). Median AATC from all 21 collections was 19.0 mL/day (range 3.0-561.0), and levels >22 mL/day indicate the diagnosis of PLE. The four children initially suspected of GVHD but who had a negative biopsy completed a total of five collections with a median AATC of 5.0 mL/day (range 3.0-16.0) vs. a median of 33.5 for the remainder of the collections (range 3-561). Stage of gut GVHD correlated with elevated AATC and with stool volume. AATC > 22 mL/day showed a sensitivity of 70% and specificity of 82% for significant gut GVHD (≥ stage 2). Seven stool collections were taken at ≥ stage 3 gut GVHD; six of those seven patients were positive for PLE. Larger stool volumes were more predictive, and five collections with stool volumes >30 mL/kg/day were positive for PLE. We conclude that a significant positive correlation exists between the severity of PLE and the stage of gut GVHD (p < 0.04), particularly obvious in patients with stages 2-4 GVHD (p = 0.03). Despite the small number of patients recruited, this study emphasizes the need to consider PLE as a useful aspect of the clinical picture. We suggest that in order to see a response to therapy and therefore a decrease in AATC, clinicians should wait at least 2 wk from the initiation of therapy before repeating AATC test. In light of the significant morbidity and mortality associated with ≥ stage 2 gut GVHD, and as an important therapeutic decision for these patients, one may consider evaluating AATC if a biopsy is not an option.

Similarities and mutual understanding: exchange experiences in Malawi for host and guest students.

BACKGROUND: A Malawian nursing college and a Norwegian university college have agreed to cooperate in facilitating clinical exchanges for nursing students at their respective institutions. This study describes the experiences of students in an innovative exchange. Norwegian and Malawian nursing students shared clinical placement in pairs of two in Malawi for 8 weeks. This study shows that both host and guest students benefit from the clinical placement as it enhances development of clinical competence. METHODS: The design of the study is phenomenological/hermeneutical. The theoretical foundation is based upon Campinha-Bacote's model for the development of cultural competence. All participating nursing students were interviewed. Their stories provided the rationale for the three main categories that are discussed. FINDINGS: Students experience both similarities and differences in practice, but similarities are regarded as the stronger impression. Learning relational skills is the primary learning outcome, but learning how to nurse patients is also an important outcome. During the exchange period all students developed cultural competence. This way of organizing shared placements for guest and host students from different countries is valuable for all students. It also met the curricular demands in both countries.

"Candidatus Paraporphyromonas polyenzymogenes" encodes multi-modular cellulases linked to the type IX secretion system.

BACKGROUND: In nature, obligate herbivorous ruminants have a close symbiotic relationship with their gastrointestinal microbiome, which proficiently deconstructs plant biomass. Despite decades of research, lignocellulose degradation in the rumen has thus far been attributed to a limited number of culturable microorganisms. Here, we combine meta-omics and enzymology to identify and describe a novel Bacteroidetes family ("Candidatus MH11") composed entirely of uncultivated strains that are predominant in ruminants and only distantly related to previously characterized taxa. RESULTS: The first metabolic reconstruction of Ca. MH11-affiliated genome bins, with a particular focus on the provisionally named "Candidatus Paraporphyromonas polyenzymogenes", illustrated their capacity to degrade various lignocellulosic substrates via comprehensive inventories of singular and multi-modular carbohydrate active enzymes (CAZymes). Closer examination revealed an absence of archetypical polysaccharide utilization loci found in human gut microbiota. Instead, we identified many multi-modular CAZymes putatively secreted via the Bacteroidetes-specific type IX secretion system (T9SS). This included cellulases with two or more catalytic domains, which are modular arrangements that are unique to Bacteroidetes species studied to date. Core metabolic proteins from Ca. P. polyenzymogenes were detected in metaproteomic data and were enriched in rumen-incubated plant biomass, indicating that active saccharification and fermentation of complex carbohydrates could be assigned to members of this novel family. Biochemical analysis of selected Ca. P. polyenzymogenes CAZymes further iterated the cellulolytic activity of this hitherto uncultured bacterium towards linear polymers, such as amorphous and crystalline cellulose as well as mixed linkage ß-glucans. CONCLUSION: We propose that Ca. P. polyenzymogene genotypes and other Ca. MH11 members actively degrade plant biomass in the rumen of cows, sheep and most likely other ruminants, utilizing singular and multi-domain catalytic CAZymes secreted through the T9SS. The discovery of a prominent role of multi-modular cellulases in the Gram-negative Bacteroidetes, together with similar findings for Gram-positive cellulosomal bacteria (Ruminococcus flavefaciens) and anaerobic fungi (Orpinomyces sp.), suggests that complex enzymes are essential and have evolved within all major cellulolytic dominions inherent to the rumen.

[Immunisation strategies for the management of severe acute respiratory syndrome (SARS)]. / Impfstrategien bei Schwerem Akutem Respiratorischem Syndrom (SARS).

Most patients with severe acute respiratory syndrome (SARS) develop antibodies against the SARS coronavirus and survive the infection. This suggests that active or passive immunisation might be an effective option in preventing or treating SARS. Therefore, the development of SARS vaccination strategies belongs to the most important targets of SARS research. The present study analyses data-bases for the current knowledge on vaccination strategies. Both, passive and active immunisation protocols are presently being developed. Passive immunisation with sera from surviving patients leads to partial success. Whereas the passive immunisation exhibits a promising therapeutic tool, only active immunisation can successfully prevent infection. A number of approaches has been used on the basis of inactivated SARS coronaviruses, recombinant subunits, recombinant DNA, and viral vectors. However, all recently developed candidates need to be evaluated critically before clinical use. The so-called "antibody-dependent enhancement" can improve viral uptake into host cells resulting in increased viral load and exacerbation of disease.

The burden of chemotherapy-induced nausea and vomiting in children receiving hematopoietic stem cell transplantation conditioning: a prospective study.

This prospective study describes chemotherapy-induced nausea and vomiting (CINV) in children (4-18 years) receiving their first hematopoietic stem cell transplant. Emetic episodes, nausea severity (assessed using a validated, self-report nausea severity assessment tool) and antiemetic administration were documented from the start of conditioning until 24 h after the last conditioning agent was administered (acute) and for a further 7 days (delayed). Relationships between CINV control and parenteral nutrition (PN) use and acute gut GvHD (aGvHD) were explored. Fifty-nine children (4.6-17.4 years) were evaluable. Complete chemotherapy-induced vomiting (CIV; acute: 24%; delayed 22%) and chemotherapy-induced nausea (CIN; acute 7%; delayed 12%) control rates were low. Few children experienced complete CINV control (no vomiting/retching and no nausea) during the acute (5%) or delayed phases (12%). Children experiencing complete acute or delayed CIN control or complete delayed CIV control were more likely to have received: a lower proportion of their total energy requirement as PN at the end of the delayed phase (P<0.036) and PN for a shorter time (P<0.044). Low patient numbers did not permit evaluation of the association between gut aGvHD and CINV control. Effective and safe interventions aimed at improving CINV control in children are required.

Novel Syntrophic Populations Dominate an Ammonia-Tolerant Methanogenic Microbiome.

Biogas reactors operating with protein-rich substrates have high methane potential and industrial value; however, they are highly susceptible to process failure because of the accumulation of ammonia. High ammonia levels cause a decline in acetate-utilizing methanogens and instead promote the conversion of acetate via a two-step mechanism involving syntrophic acetate oxidation (SAO) to H2 and CO2, followed by hydrogenotrophic methanogenesis. Despite the key role of syntrophic acetate-oxidizing bacteria (SAOB), only a few culturable representatives have been characterized. Here we show that the microbiome of a commercial, ammonia-tolerant biogas reactor harbors a deeply branched, uncultured phylotype (unFirm_1) accounting for approximately 5% of the 16S rRNA gene inventory and sharing 88% 16S rRNA gene identity with its closest characterized relative. Reconstructed genome and quantitative metaproteomic analyses imply unFirm_1's metabolic dominance and SAO capabilities, whereby the key enzymes required for acetate oxidation are among the most highly detected in the reactor microbiome. While culturable SAOB were identified in genomic analyses of the reactor, their limited proteomic representation suggests that unFirm_1 plays an important role in channeling acetate toward methane. Notably, unFirm_1-like populations were found in other high-ammonia biogas installations, conjecturing a broader importance for this novel clade of SAOB in anaerobic fermentations. IMPORTANCE The microbial production of methane or "biogas" is an attractive renewable energy technology that can recycle organic waste into biofuel. Biogas reactors operating with protein-rich substrates such as household municipal or agricultural wastes have significant industrial and societal value; however, they are highly unstable and frequently collapse due to the accumulation of ammonia. We report the discovery of a novel uncultured phylotype (unFirm_1) that is highly detectable in metaproteomic data generated from an ammonia-tolerant commercial reactor. Importantly, unFirm_1 is proposed to perform a key metabolic step in biogas microbiomes, whereby it syntrophically oxidizes acetate to hydrogen and carbon dioxide, which methanogens then covert to methane. Only very few culturable syntrophic acetate-oxidizing bacteria have been described, and all were detected at low in situ levels compared to unFirm_1. Broader comparisons produced the hypothesis that unFirm_1 is a key mediator toward the successful long-term stable operation of biogas production using protein-rich substrates.

Isolation and characterization of hemolymph antifreeze proteins from larvae of the longhorn beetle Rhagium inquisitor (L.).

We describe a simple and effective procedure to isolate antifreeze proteins (AFPs) from the hemolymph of larvae of the longhorn beetle Rhagium inquisitor, and present some characteristics of their structures. Several AFPs were isolated from the hemolymph of this species by heat and acid extraction followed by cation exchange. The hemolymph contains at least six AFPs ranging in size from 12.5 to 12.8 kDa. Of these, three were separated to purity by the ion exchange step, as indicated by mass spectrometry. The remaining three forms were further separated by size exclusion chromatography, but could not be isolated to purity. All AFPs in the hemolymph of this species appears to have isoelectric points above 8.00. The dominant form, RiAFP(H4), was purified by the ion exchange step. Its amino acid composition reveals a lower level of cysteine and a higher level of threonine, arginine, alanine and glycine than seen in other insect AFPs. Its trypsin fingerprint does not match that of any known protein. It interacts with ice both in the anionic and cationic state.

Urinary symptomatology in younger men.

OBJECTIVES: We surveyed a "population" of younger men (20 to 49 years old) for lower urinary tract symptomatology and for symptomatology associated with prostatitis. METHODS: A National Guard unit was surveyed by mail with a 58-question urinary symptom questionnaire. Surveys were returned anonymously by mail. RESULTS: International Prostate Symptom Score (IPSS) of 8 or greater was seen in 5% of men in their 20s and rose to 15% of those in their 40s. Approximately 5% (0% to 7%) reported a history of prostatitis. Caffeine caused symptoms in 2% to 13%, while exercise and smoking were not associated with symptoms. Individual prostatitis symptoms were only seen occasionally across this age group. CONCLUSIONS: As measured by the IPSS, urinary symptoms increased during the 20 to 49-year age period. A history of prostatitis in much less common than most nonpopulation studies suggest.

Saccharin aversions in hamsters as a result of nicotine injections.

Golden Syrian hamsters (males, N = 70) showed dose-related conditioned taste aversion (CTA) when saccharin drinking was followed by delayed nicotine injections. Baseline consisted of measuring amounts consumed after 20 minutes of daily access to tap water. Measures were taken for five days. The hamsters were then conditioned by offering them saccharin solution (0.1%, w/v) for 20 minutes; afterwhich a 30 minute delay was imposed. Subsequent to the delay, groups of 10 animals were treated as follows: nicotine injection (1.0, 3.0, or 9.0 mg/kg, IP), saline injection, lithium chloride injection (2% body weight of a 0.15 M solution), sham injection, or left in their cages as handling/stress controls. Following two recovery days with plain water available for 20 minutes, all animals were tested for CTA by offering them saccharin solution. Dose-related CTA was demonstrated in the nicotine animals as measured by a decrease in saccharin consumption compared to drinking measures obtained from animals injected with saline. Lithium chloride produced the same degree of CTA as 9 mg/kg of nicotine, and the aversions had extinguished in all groups by the third test day.

Conditioned saccharin aversions in rats as a result of cutaneous nicotine or intraperitoneal nicotine administered in divided doses.

Nicotine base was used in a conditioned taste aversion (CTA) paradigm to avert male Sprague-Dawley rats to saccharin solution (0.1%, w/v). Experiments investigated different dose routes of nicotine administration and duration of action as determinants in nicotine-induced CTA. In Experiment 1 nicotine was injected intraperitoneally (IP) at doses of 0.5, 1.0, or 3.0 mg/kg 30 min after drinking saccharin solution. Using a two-bottle choice test, no CTA was observed, although all nicotine animals showed obvious symptoms of malaise including seizures in the highest dose group. Experiment 2 showed dose-related CTA when nicotine (10.0, 30.0, or 50.0 mg/kg) was cutaneously applied 30 min following saccharin drinking. Experiment 2B showed that the aversions were due to associative rather than nonassociative factors such as sensitization or enhanced neophobia. In Experiment 3, the following group treatments were begun 30 min after saccharin drinking to distribute identical total nicotine doses over an extended period of time: One IP injection of 2.0 mg/kg nicotine (in a saline vehicle) and four injections of saline solution, three injections of 0.67 mg/kg nicotine and two injections of saline, five injections of 0.40 mg/kg nicotine, or five injections of saline. All injections were spaced 30 min apart. Compared with saline-injected controls, CTA occurred in the rats receiving either three or five injections of nicotine but the group receiving one injection did not differ from the control group. There was no difference in CTA between the groups receiving three or five injections.

Urban anglers' perception of risk from contaminated fish.

The Newark Bay Complex includes the Newark Bay, tidal portions of the Hackensack River, Passaic River, Arthur Kill, and Kill van Kull. It is a highly industrialized urban area including five counties and more than 20 local governments with a large racially-mixed population of more than 3 million people. In 1982, research conducted by the New Jersey Department of Environmental Protection (NJDEP) showed elevated levels of chemical contaminants in five species of fish and one type of crab in the Newark Bay Complex. Subsequently, the State of New Jersey adopted advisories to guide citizens on safe consumption practices for fish and crabs. Since then, fish consumption advisories have been issued primarily through the Fish and Game Digest, a publication distributed by the state to licensed anglers. However, anglers in the Complex are not required to have a fishing license because the waters are marine. Therefore, most anglers in this area do not receive advisory information. To gain greater insight into the information sources and risk perceptions of urban anglers, a survey was conducted of 300 anglers at 26 fishing and crabbing locations in the Newark Bay Complex during the summer and early fall of 1995. The objectives of the study were to learn anglers': (1) knowledge of fish consumption advisories; (2) belief in the advisories; (3) perception of how safe fish are to eat; (4) sources for information about fish and fishing; and (5) sources for information on fish consumption advisories. The study concluded that while 60% had heard about advisories, they either did not believe or were unconcerned about health effects from eating contaminated species. In addition, the most used source for information about fish and fishing was other fishermen, while newspapers were selected as a source for information about community news, health, and food safety.

Effects of arginine and ornithine on strength, lean body mass and urinary hydroxyproline in adult males.

Twenty-two adult males participated in a 5 week progressive strength training program. One half the subjects received the amino acids L-arginine and L-ornithine and the other half, a placebo. The study used a double blind protocol so that subjects as well as investigators had no knowledge of which substances were being administered. Dosages amounted to 2 grams or 1 gram each of L-arginine and L-ornithine, and 600 mg of calcium and 1 gram of Vitamin C as placebos. These supplements were taken orally for a total of 25 administrations. Following the short term strength program using progressively high intensities, tests were taken for total strength (TS), lean body mass (LBM) and urinary hydroxyproline (UH). The results from ANOVA showed that subjects who were taking the arginine-ornithine combination scored significantly higher in TS and LBM (p less than .05), and significantly lower in UH (p less than .05), than subjects on placebos. It was concluded that arginine and ornithine taken in prescribed doses can, in conjunction with a high intensity strength training program, increase TS and LBM in a relatively short period of time. Arginine and ornithine also aid in recovery from chronic stress by quelling tissue breakdown as evidenced by lower UH levels.

Structural characteristics of a novel antifreeze protein from the longhorn beetle Rhagium inquisitor.

Antifreeze proteins (AFPs) are characterized by their capacity to inhibit the growth of ice and are produced by a variety of polar fish, terrestrial arthropods and other organisms inhabiting cold environments. This capacity reflects their role as stabilizers of supercooled body fluids. The longhorn beetle Rhagium inquisitor is known to express AFPs in its body fluids. In this work we report on the primary structure and structural characteristics of a 12.8 kDa AFP from this beetle (RiAFP). It has a high capacity to evoke antifreeze activity as compared to other known insect AFPs and it is structurally unique in several aspects. In contrast to the high content of disulfide bond-formation observed in other coleopteran AFPs, RiAFP contains only a single such bond. Six internal repeat segments of a thirteen residue repeat pattern is irregularly spaced apart throughout its sequence. The central part of these repeat segments is preserved as TxTxTxT, which is effectively an expansion of the TxT ice-binding motif found in the AFPs of several known insect AFPs.

Consulting for substance abuse: mental disorders among adolescents and their parents.

Studies conducted outside of Scandinavia indicate that most adolescents with substance misuse problems suffer from co-morbid mental disorders. The present study assessed the mental health of adolescents seeking help for substance misuse problems in a large Swedish city. Parents' mental health was also examined. The sample included 97 girls with their 90 mothers and 52 fathers, and 81 boys with their 72 mothers and 37 fathers. The adolescents completed a diagnostic interview, either the Kiddie-SADs or the Structured Clinical Interview for DSM-IV (SCID) depending on their age. Their parents underwent diagnostic interviews with the SCID. Ninety per cent of the girls and 81% of the boys met criteria for at least one disorder other than substance misuse, and on average, they suffered from three other disorders, most of which had onset before substance misuse began. Almost 80% of the mothers and 67% of the fathers met criteria for at least one mental disorder other than alcohol and drug-related disorders. The findings concur with those reported from studies conducted in North America. The results suggest that in Sweden mental disorders are not being identified and effectively treated among some children and young adolescents who subsequently abuse alcohol and/or illicit drugs. Adolescents who consult for substance abuse problems require assessments and treatment by mental health professionals.

[Pharmacotherapy of severe acute respiratory syndrome (SARS)]. / Pharmakotherapie bei Schwerem Akutem Respiratorischem Syndrom (SARS).

Severe acute respiratory syndrome (SARS) constitutes the first new infectious disease of the current millennium. It is caused by the novel SARS-Coronavirus (SARS-CoV). SARS is related to a high morbidity and mortality and first appeared during an epidemic in 2002 - 2003. To date no specific therapy against the SARS-CoV is available. Due to the rapid spread of SARS during the epidemics in 2002 - 2003, randomised and controlled multicentre studies were not performed. Therefore, general guidelines have not been developed. Since the outbreak, scientists have been testing potential antiviral substances using in vitro and animal models. This study analyses the presently available in vitro and in vivo data on the pharmacotherapy of SARS.