Spot test analysis of microbial contents during composting of kitchen- and garden biowaste: sampling procedures, bacterial reductions, time-temperature relationships, and their relevance for EU-regulations concerning animal by-products.

This study was aimed to collect data and develop methodologies to determine if and how Dutch biowaste composting plants can meet the microbiological requirements set out in EU-Regulations (EC) 1774/2002 and (EC) 1069/2009, and to provide the European Food and Safety Authority (EFSA) with data and analysis for evaluation of these regulations. We examined twenty plant locations and four types of composting technologies, all with forced aeration and without an anaerobic digestion phase. Raw biowaste, material after sanitation and compost were sampled by spot test analysis according to a standard protocol, and according to an additional protocol with enhanced hygienic precautions. Samples were analyzed for Escherichia coli, Enterococcaceae and Salmonella content. The latter protocol resulted in improved bacterial reductions after sanitation, whereas in compost Enterococcus levels but not E. coli levels increased substantially with both protocols, due to more thermo-resistant regrowth. Salmonella presence in compost coincided with low temperatures and increased levels of E. coli and Enterococcus, absence of Salmonella was associated with absence of E. coli (74%), but not with absence of Enterococcus (17%). In compost, E. coli and Salmonella showed a comparable time-temperature inactivation pattern. A pilot study with co-composting of biowaste and poultry manure indicated a similar inactivation pattern for ESBL-containing bacteria. We conclude that the abundance of Enterococcus in compost is caused by regrowth and not by (re)contamination, and that E. coli is a more reliable indicator species for the absence/presence of Salmonella in compost. Compliance with current EU-regulations concerning biowaste composting can be shown by spot test analysis at all examined plants, provided that adequate hygienic precautions are taken during sampling.

Intralipid administration in case of a severe venlafaxine overdose in a patient with previous gastric bypass surgery.

This case describes a patient with a history of bariatric surgery who was admitted to our hospital with a severe venlafaxine intoxication. Due to the altered anatomy of the gastrointestinal tract, the use of oral activated charcoal and large volumes of laxatives to prevent further uptake of venlafaxine was hampered. This resulted in massive absorption and a severe serotonergic syndrome. The patient was successfully treated with intravenous lipid emulsion.

The injection of deaggregated gamma globulins in adult mice induces antigen-specific unresponsiveness of T helper type 1 but not type 2 lymphocytes.

Injection of adult mice with high doses of monomeric human gamma globulins (dHGG) has been previously shown to produce a state of peripheral tolerance in both B and T cells. To gain insight into the mechanism of induction and maintenance of adult tolerance in this model, we have analyzed the pattern of lymphokines produced by control and tolerant animals in response to the tolerogen. The data presented indicate that HGG-specific, interleukin 2 (IL-2)- and interferon gamma (IFN-gamma)-producing T cells (thus referred to as T helper type 1 [Th1] cells) are rendered unresponsive after in vivo administration of soluble HGG. In contrast, antigenic stimulation of T cells isolated from tolerant adult mice leads to increased production of IL-4 in vitro. In vivo challenge of dHGG-treated adult animals with hapten-coupled HGG (p-azophenylarsonate [ARS]-HGG) induced a significant ARS-specific antibody response, suggesting that tolerance induction in this model does not completely abrogate tolerogen-specific Th activity in vivo. In agreement with the in vitro data, hapten-specific antibody response of tolerant animals is characterized by a selective deficiency in the IFN-gamma-dependent IgG2a subclass. Injection of immunogenic forms of HGG into tolerant animals also produced an IL-4-dependent increase in total serum IgE levels, indicative of an increased activity of HGG-specific Th2 cells in these animals. The finding that tolerance induction differentially affects Th subpopulations suggests that crossregulation among lymphocyte subsets may play a role in the induction and/or maintenance of acquired tolerance in adults.

Monoclonal antibodies directed against the sexual binding site of Chlamydomonas eugametos gametes.

Monoclonal antibodies were raised against the mt- sexual agglutinin of Chlamydomonas eugametos gametes. Those that blocked the agglutination site were selected. They were divided into two classes dependent upon whether they gave a weak (class A) or clear positive (class B) reaction with mt- flagellar membranes in an ELISA and an indirect immunofluorescence test using glutaraldehyde-fixed mt- gametes. Class A antibodies were shown to be specific for the agglutinin in an extract of mt- gametes, based on results from immunoblotting, immunoprecipitation, affinity chromatography, and the absence of a reaction with nonagglutinable cells. Surprisingly, class A mAbs also recognized two mt+ glycoproteins, one of which is the mt+ agglutinin. Class B antibodies were shown to bind to several glycoproteins in both mt- and mt+ gametes, including the mt- agglutinin. Fab fragments from class A mAbs blocked the sexual agglutination process, but those from class B did not, even though the parent antibody did. We conclude that the class A epitope lies in or close to the agglutination site of the mt- agglutinin, whereas the class B epitope lies elsewhere on the molecule. We also conclude that the mt- agglutinin is the only component on the mt- flagellar surface directly involved in agglutination. Class A mAbs were found to elicit several reactions displayed by the mt+ agglutinin. They bound to the mt- agglutinin on gamete flagella and induced most of the reactions typical of sexual agglutination, with the exception of flagellar tip activation. None of these reactions was induced by Fab fragments. High concentrations of class A mAbs completely repressed the sexual competence of live mt- gametes, but low concentrations stimulated cell fusion.

Everolimus pharmacokinetics and its exposure-toxicity relationship in patients with thyroid cancer.

BACKGROUND: Everolimus is a mTOR inhibitor used for the treatment of different solid malignancies. Many patients treated with the registered fixed 10 mg dose once daily are in need of dose interruptions, reductions or treatment discontinuation due to severe adverse events. This study determined the correlation between systemic everolimus exposure and toxicity. Additionally, the effect of different covariates on everolimus pharmacokinetics (PK) was explored. METHODS: Forty-two patients with advanced thyroid carcinoma were treated with 10 mg everolimus once daily. Serial pharmacokinetic sampling was performed on days 1 and 15. Subsequently, a population PK model was developed using NONMEM to estimate individual PK values used for analysis of an exposure-toxicity relationship. Furthermore, this model was used to investigate the influence of patient characteristics and genetic polymorphisms in genes coding for enzymes relevant in everolimus PK. RESULTS: Patients who required a dose reduction (n = 18) due to toxicity at any time during treatment had significant higher everolimus exposures [mean AUC0-24 (SD) 600 (274) vs. 395 (129) µg h/L, P = 0.008] than patients without a dose reduction (n = 22). A significant association between everolimus exposure and stomatitis was found in the four-level ordered logistic regression analysis (P = 0.047). The presence of at least one TTT haplotype in the ABCB1 gene was associated with a 21 % decrease in everolimus exposure. CONCLUSION: The current study showed that dose reductions and everolimus-induced stomatitis were strongly associated with systemic everolimus drug exposure in patients with cancer. Our findings confirm observations from another study in patients with cancer and show us that everolimus is a good candidate for individualized dosing in patients with cancer. CLINICALTRIAL. GOV NUMBER: NCT01118065.

Beneficial Effects of the mTOR Inhibitor Everolimus in Patients with Advanced Medullary Thyroid Carcinoma: Subgroup Results of a Phase II Trial.

Objective. Until recently, advanced medullary thyroid cancer (MTC) had few treatment options except surgery. The mTOR inhibitor everolimus has shown encouraging results in neuroendocrine tumors. As part of a prospective phase II study, we analyzed the safety and efficacy of everolimus in advanced MTC. Methods. Seven patients with per RECIST 1.1 documented advanced MTC were included and received everolimus 10 mg daily. The primary objective was determining treatment efficacy. Secondary endpoints included progression-free survival (PFS), overall survival (OS), toxicity, and pharmacokinetics (PK). Results. Median follow-up duration was 28 weeks (17-147). Five patients (71%) showed SD, of which 4 (57%) showed SD >24 weeks. Median PFS and OS were 33 (95%CI: 8-56) and 30 (95%CI: 15-45) weeks, respectively. Toxicity was predominantly grade 1/2 and included mucositis (43%), fatigue (43%), and hypertriglyceridemia (43%). Four MTCs harbored the somatic RET mutation c.2753T>C, p.Met918Thr. The best clinical response was seen in a MEN2A patient. PK characteristics were consistent with phase I data. One patient exhibited extensive toxicity accompanying elevated everolimus plasma concentrations. Conclusions. This study suggests that everolimus exerts clinically relevant antitumor activity in patients with advanced MTC. Given the high level of clinical benefit and the relatively low toxicity profile, further investigation of everolimus in these patients is warranted.

Analysis of two pharmacologically predicted endothelin B receptor subtypes by using the endothelin B receptor gene knockout mouse.

1. This study was performed to clarify whether the endothelin (ET) receptor subtypes mediating two pharmacologically heterogeneous response to ETH receptor agonists in normal mice are the product(s) of a single ETB receptor gene. 2. Vasodilator responses to sarafotoxin S6c (S6c) in the thoracic aorta and contractile responses to ET-1 and IRL1620 in the stomach were examined in tissues from normal and ETB receptor gene knockout mice, in the absence and presence of an ETA receptor antagonist, BQ-123, or an ETA/ETB receptor antagonist, PD142893. 3. In the normal mouse aorta precontracted with phenylephrine, S6c (0.1-100 nM) caused concentration-dependent relaxations (pD2 = 8.4). BQ-123 had no effect on these responses. However, PD142893 almost abolished the relaxations induced by 0.1-300 nM S6c. 4. In aortae taken from ETB receptor gene knockout mice, S6c up to 1 microM failed to cause relaxations, confirming that ETB receptors are involved in mediating this response. 5. In normal mouse gastric fundus, 0.1 nM-1 microM ET-1, S6c or IRL1620 caused dose-dependent, BQ-123-insensitive contractions, which were much more resistant to PD142893 than S6c-induced relaxations of the aorta. The pD2 values for S6c in the absence and presence of PD142893 (10 microM) were 8.12 +/- 0.11 and 7.70 +/- 0.11, respectively. 6. In the gastric fundus of the ETB receptor gene knockout mouse, S6c and IRL1620 caused no contractions. ET-1 (0.1 nM-1 microM) caused contractions sensitive to both BQ-123 and PD142893, indicating that only ETA receptors mediate ET-1-induced contractions of the knockout mouse gastric fundus. 7. Since both the PD142893-sensitive vasodilator response of the aorta and the PD142893-resistant contractile response of the gastric fundus to S6c were completely absent in the ETB receptor gene knockout mouse, we conclude that the two pharmacologically heterogeneous responses to S6c are mediated by receptors derived from the same ETB receptor gene.

Creative infection control.

In spite of the widespread use of educational programmes, there still remains a low staff knowledge and non-compliance of infection control policies. In order to overcome these problems we have attempted to increase interest and raise awareness of infection control by introducing a motivational programme termed Creative Infection Control. Some of the activities are illustrated and are loosely structured, and rely largely on humour and intergroup competition. These methods are based on sound psychological principles with an emphasis on adult-learning theory. Objective measures indicate that such programmes can significantly influence infection control outcomes.

OM197-MP-AC induces the maturation of human dendritic cells and promotes a primary T cell response.

Dendritic cell (DC) maturation is critical for the induction of antigen-specific T lymphocyte responses and may be essential for the development of human vaccines relying on T cell immunity. We investigated the effects on human DC of OM-197, a synthetic pseudodipeptide derived from amino acids, linked to three fatty acid chains and devoid of endotoxin properties. OM-197 upregulated the expression of HLA-DR, CD80, CD86, CD83, CD40 and CD54 at the surface of myeloid DC naturally present in blood as well as of DC generated in vitro from monocytes using IL-4 and GM-CSF. OM-197 also induced the release of IL-12 and TNF-alpha from DC. Finally, DC incubated with OM-197 after pulsing with hepatitis B surface antigen (HBs Ag) induced in vitro expansion of IFN-gamma-secreting HBs Ag-specific CD4(+) T lymphocytes from naive individuals. Taken together, these data identify OM-197 as a potential vaccine adjuvant for the induction of Th1-type responses.

The periodate oxidation of sucrose in aqueous N,N-dimethylformamide.

Sucrose has been oxidized with sodium periodate in 0-50% aqueous N,N-dimethylformamide (DMF). In 50% aqueous DMF the reaction is selective for the glucose ring, yielding a dialdehyde. The increased selectivity is not due to conformational factors but is ascribed to the dissociation of water from cyclic periodate ester species which makes the reaction via the acyclic ester on fructose unfavourable.

Single crystals of inulin.

Lamellar crystals of inulin were grown by crystallizing sharp fractions of low molecular weight inulin from dilute aqueous ethanol solutions. The crystals were analyzed using three-dimensional electron diffraction and X-ray powder diagrams. Two crystalline polymorphs were observed, depending on the hydration conditions: a hydrated form which indexed on an orthorhombic unit cell, with space group P2(1)2(1)2(1) and with cell dimensions of a = 1.670 nm, b = 0.980 nm and c (chain axis) = 1.47 nm, together with a pseudo-hexagonal semi-hydrated form with unit cell parameters a = 1.670 nm, b = 0.965 nm and c (chain axis) = 1.44 nm. These parameters, together with the density data, indicate that inulin crystallizes along a pseudo-hexagonal six-fold symmetry with an advance per monomer of 0.24 nm. The difference between the hydrated and the semi-hydrated unit cells does not seem to correspond to any change in the conformation of inulin, but rather to a variation in water content.

Midazolam as a phenotyping probe to predict sunitinib exposure in patients with cancer.

PURPOSE: Patients treated with sunitinib show substantial inter-patient variability in drug exposure (~30-40 %), which is largely unexplained. Since sunitinib is metabolized by cytochrome P450(CYP)3A4, variability in the activity of this enzyme may explain a considerable proportion of this inter-patient variability. Midazolam is widely used as a phenotyping probe to assess CYP3A4-activity. The objective of this study was to prospectively evaluate the relationship between midazolam and sunitinib exposure. Additionally, the correlation between sunitinib trough levels and exposure and the influence of sunitinib on midazolam exposure was determined. METHODS: Thirteen patients treated with sunitinib in a 4 weeks "on"-2 weeks "off" regimen received twice 7.5 mg midazolam; once with and once without sunitinib. Steady-state sunitinib, its active metabolite SU12662 and midazolam exposures were determined. RESULTS: A significant correlation between midazolam exposure (AUC(0-7h)) and steady-state sunitinib and sunitinib + SU12662 exposure (AUC(0-24h)) was found (p = 0.006 and p = 0.0018, respectively); midazolam exposure explained 51 and 41 % of the inter-patient variability in sunitinib and sunitinib + SU12622 exposure. Furthermore, C trough was highly correlated (r(2) = 0.94) with sunitinib AUC(0-24h). Sunitinib decreased midazolam exposure with 24 % (p = 0.034). CONCLUSION: Midazolam exposure is highly correlated with sunitinib exposure and explains a large proportion of the observed inter-patient variability in sunitinib pharmacokinetics. Consequently, midazolam could be used to identify patients that are at risk of under- or overtreatment, respectively, at the start of sunitinib therapy. Moreover, sunitinib and sunitinib + SU12662 trough levels are highly correlated with drug exposure and can thus be used in clinical practice to individualize sunitinib therapy. The decrease in midazolam exposure by sunitinib needs further investigation.

Sutureless and glue-free conjunctival autograft in pterygium surgery: a case series.

AIMS: Foreign materials used in ocular surface surgery may lead to local complications such as discomfort, scarring, or infection. Plasma-derived products such as fibrin glue may produce possible hypersensitivity reactions whereas the risk of viral transmission remains. We describe a simple method of achieving conjunctival autograft adherence during pterygium surgery avoiding potential complications associated with the use of fibrin glue or sutures. METHODS: After pterygium excision and fashioning of the autologous conjunctival graft, the recipient bed is encouraged to achieve natural haemostasis and relative dessication before graft placement. Excessive haemorrhage in the graft bed is tamponaded. Graft adherence and positioning is examined 20 min after surgery. RESULTS: A total of 15 eyes of 12 patients (mean (SD) age 73.7 (11.2) years), 8 females underwent SGF autologous conjunctival graft post-pterygium excision. Mean graft area was 24(1.5)mm(2). Mean follow-up time was 9.2 (2.2) months. Cosmesis was excellent in all cases and visual acuity improved in one patient. There were no intra- or post-operative complications requiring further treatment. CONCLUSION: This simple technique for pterygium surgery may prevent potential adverse reactions encountered with the use of foreign materials and in this small series provided safe and comparable results to current methods.