3-O sulfation of syndecan-1 mediated by the sulfotransferase HS3ST3a1 enhances myeloma aggressiveness.

Multiple myeloma is a hematological neoplasm derived from plasma cells invariably developing in the bone marrow (BM). The persisting clinical challenge in MM resides in its high ability to resist drugs as shown by the frequent relapses observed in patients regardless of the treatment applied. In a mouse model of MM, we identified a subpopulation of cells harboring increased resistance to current MM drugs. These cells bound a proliferation inducing ligand (APRIL), a key MM promoting/survival factor. APRIL binding involved the heparan sulfate (HS) chain present on syndecan-1 (SDC-1), and correlated with reactivity to the anti-HS antibody 10e4. 10e4+cells had a high proliferation activity, and were able to form colonies in 3-D cultures. 10e4+ cells were the only cells able to develop in BM after intravenous injection. They also resisted drugs in vivo, since their number increased after treatment in BM. Notably, 10e4+ cells differentiated into 10e4- cells upon in vitro and in vivo expansion. Expression of one sulfotransferase, HS3ST3a1, allowed modification of syndecan-1 to confer reactivity to 10e4 and binding to APRIL. HS3ST3a1 deletion inhibited tumorigenesis in BM. Notably, the two populations coexisted at a variable frequency in the BM of MM patients at diagnosis. In total, our results indicate that 3-O-sulfation on SDC-1 carried out by HS3ST3a1 defines aggressive MM cells, and that targeting of this enzyme could possibly be used to better control drug resistance.

Data on Transfer of Human Coronavirus SARS-CoV-2 from Foods and Packaging Materials to Gloves Indicate That Fomite Transmission Is of Minor Importance.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is mainly transmitted via droplets and aerosols. To evaluate the role of transmission by fomites, SARS-CoV-2-specific data on transfer rates from surfaces to hands and from hands to face are lacking. Here, we generated quantitatively controlled transfer rates for SARS-CoV-2 from food items (lettuce, ham, and vegetarian meat alternative [VMA]) and packaging materials (cardboard and plastic) to gloves using a wet, dry, and frozen viral inoculum and from glove to glove using a wet viral inoculum. For biosafety reasons, the transfer from surfaces to hands and hands to face was simulated by using gloves. The cumulative transfer rate was calculated by using the data from the first transfer experiment, food or packaging material to glove, and combined with the transfer rate obtained from the second transfer experiment from glove to glove. The cumulative transfer rates from lettuce (4.7%) and ham (3.4%) were not significantly different (P > 0.05) but were significantly higher (P < 0.05) than that from VMA ("wet" or "frozen"). The wet cumulative transfer rate from VMA (1.3%) was significantly higher than the cumulative transfer rate from frozen VMA (0.0011%). No transfer from plastic or cardboard was observed with a dry inoculum. The plastic packaging under wet conditions provided the highest cumulative transfer rate (3.0%), while the cumulative transfer from frozen cardboard was very small (0.035%). Overall, the transfer rates determined in this study suggest a minor role of foods or food packaging materials in infection transmission. IMPORTANCE The observation of SARS-CoV-2 RNA in swab samples from frozen fish packages in China, confirmed only once by cell culture, led to the hypothesis that food contaminated with SARS-CoV-2 virus particles could be the source of an outbreak. Epidemiological evidence for fomites as infection source is scarce, but it is important for the food industry to evaluate this infection path with quantitative microbial risk assessment (QMRA), using measured viral transfer rates from surfaces to hands and face. The present study provides transfer data for SARS-CoV-2 from various types of foods and packaging materials using quantitative methods that take uncertainties related to the virus recovery from the different surfaces into consideration. The transfer data from this model system provide important input parameters for QMRA models to assess the risk of SARS-CoV-2 transmission from contaminated food items.

Assessment of Antiviral Coatings for High-Touch Surfaces by Using Human Coronaviruses HCoV-229E and SARS-CoV-2.

A novel and robust approach to evaluate the antiviral activity of coatings was developed, assessing three commercially available leave-on surface coating products for efficacy against human coronaviruses (HCoVs) HCoV-229E and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The assessment is based on three criteria that reflect real-life settings, namely, (i) immediate antiviral effect, (ii) effect after repeated cleaning of the coated surface, and (iii) antiviral activity in the presence of organic material. The results showed that only a copper compound-based coating successfully met all three criteria. A quaternary ammonium compound-based coating did not meet the second criterion, and a coating based on reactive oxygen species showed no antiviral effect. Moreover, the study demonstrated that HCoV-229E is a relevant SARS-CoV-2 surrogate for such experiments. This new approach allows benchmarking of currently available antiviral coatings and future coating developments to avoid unjustified claims. The deployment of efficient antiviral coatings can offer an additional measure to mitigate the risk of transmission of respiratory viruses like SARS-CoV-2 or influenza viruses from high-touch surfaces. IMPORTANCE SARS-CoV-2, the virus responsible for the coronavirus disease 2019 (COVID-19) pandemic, is transmitted mainly person-to-person through respiratory droplets, while the contribution of fomite transmission is less important than suspected at the beginning of the pandemic. Nevertheless, antiviral-coating solutions can offer an additional measure to mitigate the risk of SARS-CoV-2 transmission from high-touch surfaces. The deployment of antiviral coatings is not new, but what is currently lacking is solid scientific evidence of the efficacy of commercially available self-disinfecting surfaces under real-life conditions. Therefore, we developed a novel, robust approach to evaluate the antiviral activity of such coatings, applying strict quality criteria to three commercially available products to test their efficacies against SARS-CoV-2. We also showed that HCoV-229E is a relevant surrogate for such experiments. Our approach will also bring significant benefit to the evaluation of the effects of coatings on the survival of nonenveloped viruses, which are known to be more tolerant to desiccation and disinfectants and for which high-touch surfaces play an important role.

Surveillance recommendations for patients with Lynch syndrome and FAP : a monocentric study.

BACKGROUND AND STUDY AIMS: The most important causes of hereditary colorectal cancer are Lynch syndrome (LS) and the adenomatous polyposis syndromes (familial adenomatous poly- posis syndrome or FAP, attenuated FAP or AFAP and MUTYH associated polyposis syndrome or MAP). The aim of this study was to investigate whether all patients with a hereditary syndrome within one center receive uniform advice regarding surveillance and treatment. PATIENTS AND METHODS: A retrospective analysis was performed of all electronic patient health records of patients with LS, FAP, AFAP and MAP who received genetic counselling or were followed by a health care specialist at the University Hospital in Ghent. RESULTS: Data from 122 patients were collected. For all patients, recommendations from the medical genetics department were highly consistent. Adherence to their recommendations was good within the center for the management of colon polyps. There was a lack of consistency in the screening and surveillance advice for other tumors in departments other than gastroenterology. Only 33 patients had systematic follow-up consultations to check results and organize surveillance. CONCLUSION: Previously, small studies have suggested that patients with hereditary gastrointestinal cancer syndromes infrequently have surveillance as specified in the guidelines. This study shows almost uniform recommendations and good adherence for surveillance of the colon, but incomplete or contradictory advice for surveillance of other organs. The need for an integrated approach from a multidisciplinary team will only increase in the future, because more families with hereditary cancer are likely to be found due to the increased use of next generation sequencing in cancer diagnostics.

Giant peritoneal loose body : a case report and review of the literature.

We present a case about a 53-year-old man who complained of abdominal pain and constipation. Computed tomography showed a well-described nodular structure of 6cm in size with a central dense core of 0.5cm with compression against the rectosigmoid. The presence of a foreign body was suggested and a diagnostic laparoscopy was performed. Surgery revealed a giant peritoneal loose body measuring 5.5cm in diameter. After the removal, the patient was relieved of his symptoms. Peritoneal loose bodies are usually small and asymptomatic. They are mostly found incidentally during laparotomy. Giant peritoneal loose bodies are a rare entity and diagnosis is difficult. A review of the literature is presented.

Review: norovirus prevalence in Belgian, Canadian and French fresh produce: a threat to human health?

Foodborne viruses, especially noroviruses (NoV), are increasingly reported as the cause of foodborne outbreaks. NoV outbreaks have been reported linked to fresh soft red fruits and leafy greens. Belgium, Canada and France were the first countries to provide data about the prevalence of NoV on fresh produce. In total, 867 samples of leafy greens, 180 samples of fresh soft red fruits and 57 samples of other types of fresh produce (tomatoes, cucumber and fruit salads) were analyzed. Firstly, the NoV detection methodology, including virus and RNA extraction, real-time RT-PCR and quality controls were compared among the three countries. In addition, confirmation and genotyping of the NoV strains was attempted for a subset of NoV positive samples using conventional RT-PCR targeting an alternative region followed by sequencing. Analysis of the process control showed that 653, 179 and 18 samples of the leafy greens, soft red fruits and other fresh produce types were valid for analysis based on the recovery of the process control. NoV was detected by real-time RT-PCR in 28.2% (N=641), 33.3% (N=6) and 50% (N=6) of leafy greens tested in Canada, Belgium and France, respectively. Soft red fruits were found positive by real-time RT-PCR in 34.5% (N=29) and 6.7% (N=150) of the samples tested in Belgium and France, respectively. 55.5% (N=18) of the other fresh produce types, analyzed in Belgium, were found NoV positive by real-time RT-PCR. Conventional RT-PCR resulted in an amplicon of the expected size in 19.5% (52/266) of the NoV positive samples where this assay was attempted. Subsequent sequencing was only successful in 34.6% (18/52) of the suspected amplicons obtained by conventional RT-PCR. From this study, using the described methodology, NoV genomes were frequently detected in fresh produce however sequence confirmation was not successful for the majority of the samples tested. Infection or outbreaks were rarely or not known to be related to the NoV positive samples. With the increase in sensitivity of the detection methodology, there is an increasing concern about the interpretation of positive NoV results by real-time amplification. Strategies to confirm the results by real-time RT-PCR should be developed in analogy with the detection of microbial pathogens in foods. Detection might indicate contact with NoV in the fresh produce chain. Consequently, a potential risk for infection cannot be excluded but the actual risk from RT-PCR NoV positive produce is still unknown. Studies should be designed determining the probability of infection related to the presence or levels of NoV genomic copies.

Parallel habitat specialization within the wolf spider genus Hogna from the Galápagos.

Within most island archipelagos, such as the Galápagos, similar ecological gradients are found on geographically isolated islands. Species radiations in response to these ecological gradients may follow different scenarios being (i) a single habitat specialization event followed by secondary colonization of each ecotype on the different islands or (ii) repeated and parallel habitat specialization on each island separately. This latter scenario has been considered less likely as gene flow might hamper such ecotypic differentiation. At least for the Galápagos, the extent to which this process is involved in species radiations remains yet poorly understood. Within the wolf spider genus Hogna, seven species are described that can be divided into three different ecotypes based on general morphology and habitat preference i.e. species that inhabit the pampa vegetation in the highlands, species that occur in coastal dry habitats and one generalist species. Comparison of the species phylogeny based on one mitochondrial (COI) and one nuclear (28S) gene fragment convincingly demonstrates that 'pampa' and 'coastal dry' species evolved in parallel on the islands Santa Cruz and San Cristóbal. Despite the observation that allozymes analysis indicated that each species forms a distinct genetic cluster, phylogenetic divergence within these species complexes was very low and paraphyletic and most likely due to hybridization rather than incomplete lineage sorting, as demonstrated for the Santa Cruz species complex. This suggests that within-island speciation occurred under low levels of gene flow. Species phylogeny in general did not follow the progression of island emergence as a molecular clock analysis suggested that island endemic species may have diverged after as well as before the emergence of the islands. This represents the first clear example of parallel and within-island speciation because of habitat specialization on the Galápagos and that such divergence most likely occurred under historic gene flow.

The efficacy of preservation methods to inactivate foodborne viruses.

During the last decade an increased incidence of infections and outbreaks attributed to foodborne viruses, in particular noroviruses (NoV), was observed world wide. The awareness of the presence of viruses on food emphasized the need to acquire knowledge regarding the effect of preservation methods upon viruses. Most foodborne viruses cannot be cultured in the laboratory, which hinders studies of their stability in food. Cultivable surrogate viruses, genetically related to the human infecting strains, are taken as a substitute to define inactivation rates. The last years, the number of survival and inactivation studies using various surrogate viruses increased. In this review, state-of-the-art information regarding the efficacy of preservation methods to reduce the level of viruses on food is compiled. In the first place, the effect of preservation methods establishing microbial growth inhibition (chilling, freezing, acidification, reduced water activity and modified atmosphere packaging) upon foodborne viruses is described. Secondly, the use of preservation methods establishing microbial inactivation such as heat treatment, high hydrostatic pressure processing and irradiation to eliminate viruses is discussed. In the third place, the efficacy of decontamination methods on fresh produce and purification procedures applied on live bivalve shellfish to reduce the viral load is included. These studies indicate that viruses persist well on chilled, acidified, frozen foods and foods packed under modified atmosphere or in dried conditions. Intervention strategies inducing microbial inactivation are required to achieve a 3 log reduction of the level of viruses. Decontamination of fresh produce reduces viruses with a maximum of 1 to 2 log while purification of live bivalves is not adequate to prevent viral outbreaks. It was noted that the effect of a particular food preservation method is dependent upon the virus tested and type of food.

Reported foodborne outbreaks due to noroviruses in Belgium: the link between food and patient investigations in an international context.

The Belgian data for foodborne norovirus (NoV) outbreaks became available for the first time with the introduction of an extraction and detection protocol for NoV in the National Reference Laboratory for foodborne outbreaks in September 2006. In 2007, 10 NoV foodborne outbreaks were reported affecting 392 persons in Belgium. NoV became the most detected agent in foodborne outbreaks followed by Salmonella (eight foodborne outbreaks). The major implicated foods were sandwiches (4/10), where food handlers reported a history of gastroenteritis in two outbreaks. A food handler was implicated in the limited number of Belgian NoV outbreaks which is in accord with internationally recorded data. Forty foodborne and waterborne outbreak events due to NoV, epidemiological and/or laboratory confirmed, from 2000 to 2007 revealed that in 42.5% of the cases the food handler was responsible for the outbreak, followed by water (27.5%), bivalve shellfish (17.5%) and raspberries (10.0%).

Development of a liquid chromatographic assay for an anti-HIV tablet containing lamivudine, zidovudine and TMC278.HCL.

A liquid chromatographic method was developed to analyse a tablet containing three anti-human immunodeficiency virus (HIV) compounds: lamivudine, zidovudine and a compound with the code name TMC278.HCl. Due to the presence of UV absorbing chromophores in the three active components, a single LC method with UV detection was developed. A Hypersil BDS C(18) column was used as stationary phase and the assay was performed with gradient elution using mobile phases containing acetonitrile, 0.2M potassium dihydrogen phosphate and water. The sample pretreatment is performed by treating the formulation with dimethyl sulfoxide-water (1:1) followed by filtration. After method development, the influence of the different chromatographic parameters on the separation, the interference of other active compounds and excipients, the repeatability and the linearity were investigated. The method was shown to be robust, selective, linear and repeatable. Finally, the content of the compounds in the tablet was determined.

The reduction of murine norovirus 1, B. fragilis HSP40 infecting phage B40-8 and E. coli after a mild thermal pasteurization process of raspberry puree.

Pasteurization processes of raspberry puree are nowadays limited to short times and rather low temperatures to maintain flavor and nutritional quality. Norovirus (NoV) outbreaks associated with raspberries highlight the need to determine the survival of NoV on this type of soft fruit. Therefore, resistance of murine norovirus 1 (MNV-1), a surrogate for human NoV, B. fragilis HSP40 infecting phage B40-8, and E. coli towards mild pasteurization was tested. Raspberry puree heat treated at 65 degrees C for 30s showed a 1.86, 2.77, and 3.89 log reduction of, respectively, MNV-1, E. coli, and B40-8. Heating at 75 degrees C for 15s established a 2.81 log reduction of MNV-1 while a 3.44 and 3.61 log reduction of B40-8 and E. coli was observed. No supplementary lethal effect of holding the heat-treated raspberry puree at 4 degrees C overnight was noticed. B40-8 failed to be useful as a tool to monitor NoV inactivation during mild pasteurization processes. Moreover, <3 log reductions of MNV-1 were observed suggesting that upon high initial contamination load, infectious NoV particles may remain on mildly pasteurized raspberry puree.

Evaluation of two viral extraction methods for the detection of human noroviruses in shellfish with conventional and real-time reverse transcriptase PCR.

AIMS: Comparison of two viral extraction methods in order to establish a sensitive and simple detection method for human noroviruses (NV) in bivalve shellfish. METHODS AND RESULTS: A direct RNA extraction method and an alkaline virus elution-concentration method were tested on artificially contaminated mussels. The latter used an alkaline buffer and polyethylene glycol (PEG) to isolate and concentrate the virus particles from shellfish. In both methods Trizol was used to release RNA. The final RNA extracts were amplified and detected with conventional and real-time reverse transcriptase PCR. The direct RNA extraction method was not able to detect low inoculation levels. However, the virus elution-concentration method was more sensitive. CONCLUSIONS: The alkaline elution-PEG concentration method followed by Trizol effectively removed inhibitory components and fulfilled the demands to be a useful tool for routine testing of shellfish for NV detection. SIGNIFICANCE AND IMPACT OF THE STUDY: Because of the lack of standardized methods to detect NV in shellfish, many 'in-house' extraction methods are used in practice. A comparison of these methods aims to determine a simple, rapid and sensitive method that could be a candidate method for screening suspected shellfish.

Itraconazole formulation studies of the melt-extrusion process with mixture design.

Itraconazole is a poorly water soluble compound. One method to increase the aqueous solubility of itraconazole is through formation of a solid dispersion. The purpose of this study is to develop a 40% w/w itraconazole formulation through solid dispersion formation, using hydroxypropyl-beta-cyclodextrin (HP-beta-CD) and hydroxypropylmethyl-cellulose (HPMC) as mixture components. The solid dispersion was obtained by melt-extrusion using a twin-screw corotating melt extruder. A D-optimal mixture design was applied for the development of the optimal itraconazole formulation. The itraconazole fraction varied between 20% w/w and 50% w/w in the mixture design and the HPMC and HP-beta-CD fractions varied between 10% w/w and 60% w/w. The itraconazole formulation was optimized by producing clear extrudates, minimizing the torque, and maximizing the glass transition temperature and the apparent itraconazole solubility in 0.1 N HCl. Regression models were developed for the torque, glass transition temperature, and apparent solubility of itraconazole. High itraconazole fraction in the mixture promoted a better melt processing (minimizes torque). High HPMC fraction (>33% w/w) resulted in clear extrudates, indicating a solid dispersion and resulted in high glass transition temperature of the melt. High HP-beta-CD fraction resulted in increased apparent itraconazole solubility in 0.1 N HCl. The optimal itraconazole formulation consisted of 45% w/w HPMC and 15% HP-beta-CD w/w.

Using deepest regression method for optimization of fluidized bed granulation on semi-full scale.

This study applied the deepest regression method to estimate the granule size of unsuccessful fluidized bed granulation runs. This study uses data from a previous study [Int. J. Pharm. 220 (2001) 149] on optimization of fluidized granulation process, wherein 8 of the 30 runs did not succeeded due to overwetting of the powder bed. The "complete data" (the observed and the estimated granule size by the depth regression method) were used to develop two regression models for the granule size: an empirical model based on the process variables (inlet air temperature, inlet airflow rate, spray rate, and inlet air humidity) and a fundamental model based on the powder bed moisture content and the relative droplet size. The regression models based on the incomplete data from the previous study and the regression models of the "complete data" were comparable in the sense that the contour plots based on the respective models and the predicted granule size were comparable.

Scaling up of the fluidized bed granulation process.

The scaling up of a fluidized bed granulation process from small scale to production scale is often done empirically in the pharmaceutical industry. In this study, a more practical and systematic method was developed in order to achieve a similar granule size in the scaled up fluid bed. The scaling up is based on the relative droplet size, and the powder bed moisture content at the end of the spraying cycle. The present study describes the scaling up of the fluidized granulation process from small (5 kg scale) to medium (30 kg scale) and to production fluid bed scale (120 kg scale). The granulation process is scaled up with as target a geometric mean granule size of 400 microm. First, the effect of the relative droplet size on the granule size was investigated in the different fluid beds. The effect of the change in relative droplet size on the granule size was different for each fluid bed. Second, experimental design is applied on the small and the medium fluid scale, and regression models for the granule size are proposed in order to scale up the granulation process on the small to medium scale. The granulation process was also successful by scaling-up to the large fluid bed, considering only the relative droplet size.

High initial efficacy of full-spectrum therapy for nocturnal enuresis in children and adolescents.

OBJECTIVE: To investigate the initial efficacy and predictive factors of full-spectrum therapy in the treatment of children and young adolescents with nocturnal enuresis (NE). PATIENTS AND METHODS: Combined therapy for NE comprises an enuresis alarm, bladder training, motivational therapy and pelvic floor muscle training, and is more effective than each of the components alone or than medical intervention. A total of 60 children and adolescents (aged 4-20 years) with NE were treated once a week with full-spectrum therapy for a maximum of 6 months. RESULTS: Overall the therapy was successful (14 consecutive dry nights) in 52 of 60 patients. At 30 days the cure rate was 33%, after 60 days 72% and after 98 days, 87%. The remaining 13% did not achieve 14 consecutive dry nights; seven patients improved, having fewer dry nights/week. One patient discontinued the treatment because of lack of motivation. In children with an initial maximum bladder capacity less than normal for age, the capacity increased from 53% of the normal maximum bladder capacity in week 1 to 88% at the end of treatment. Neither age, gender, sleep arousal, bladder capacity, family history and pathophysiological profile had any association with the success rate. CONCLUSION: The short-term success rate of full-spectrum therapy for NE is high. Age, gender, sleep arousal, bladder capacity, family history and pathophysiological profile of enuresis are unrelated to the success of the intervention.

Hypericin-based fluorescence diagnosis of bladder carcinoma.

OBJECTIVE: To determine the use of hypericin instillation for the fluorescent detection of papillary bladder cancer and carcinoma in situ. PATIENTS AND METHODS: Eighty-seven patients with papillary bladder cancer and/or carcinoma in situ received instillations with 40 mL of an 8 micromol/L hypericin solution for at least 2 h. Fluorescent excitation with blue light was effective for up to 16 h, and biopsies were examined by fluorescence microscopy. RESULTS: There were no side-effects reported, no photobleaching and all papillary lesions fluoresced red. The sensitivity and specificity for detecting carcinoma in situ was 94% and 95%, respectively. An interval of 4 months is recommended after BCG instillations before using this test. Fluorescence microscopy showed that hypericin was selectively localized in the epithelium. CONCLUSIONS: Hypericin-induced fluorescence has a high sensitivity and specificity for detecting bladder cancer. After 4 months there are few false-positive results in patients treated with BCG.