Comparative Testing Report on the Detection and Quantification of GM events in compound feedstuff - Comparative testing round: ILC-EURL-GMFF-CT-02/12

The European Union Reference Laboratory for Genetically Modified Food and Feed (EU-RL GMFF (Regulation (EC) No 1829/2003(1)) that is also mandated as EU-RL by Regulation (EC) 882/2006(2) organised a comparative testing round for National Reference Laboratories (NRLs) nominated under Regulation (EC) No 882/2004(2). Participation was open and free of charge for NRLs nominated under Regulation (EC) No 1981/2006(3), for all members of the European Network of GMO Laboratories (ENGL), and for official control laboratories from the EU and third countries. The EU-RL is accredited under ISO 17043 (‘General requirements for proficiency testing’(4)) and this comparative testing round met this ISO Standard(4). The test items were produced in-house from dried leaves of MON 88017 (MON-88Ø17-3) and seeds of soybean event 40-3-2 (MON-Ø4Ø32-6) provided by Monsanto, by spiking a compound feedstuff provided by a Belgian NRL. Participants were required to screen two test items (feedstuff Levels 1 and 2) for the presence of maize events Maize MON 88017, MON 89034 and soybean events 356043, 40-3-2 and MON 89788. Any event detected then had to be quantified. Participants could report the results in mass/mass % or copy/copy % and the EU-RL calculated the robust means (R) of Level 1 and 2 test items accordingly. In addition, "target" values () were assigned by the EU-RL on the basis of its homogeneity study(8) for m/m % data. These values were included in the uncertainty budget. The target standard deviation for CT was fixed by the Advisory Board for Comparative Testing at 0.15 (log10 value) for soybean event 40-3-2 and at 0.20 for maize event MON 88017 based on experience from previous CT rounds. This target standard deviation was used to derive z-scores for the participants’ results. Ninety laboratories from 43 countries registered for this CT round of which 82 from 35 countries returned at least qualitative test results. The results of the qualitative evaluation of the GM content indicated that most of the laboratories correctly detected soybean event 40-3-2 and maize event MON 88017 thus resulting in a very good performance overall. The results of the quantitative evaluation of GM content were found to be satisfactory overall for both events, with 94% of the laboratories submitting results in mass/mass % with a z-score, estimated on the basis of the robust mean, lying within the range of -2 to +2. This percentage decreased to 79% for results expressed in copy/copy %. When asked to repeat experimental work, most of the underperforming laboratories obtained satisfactory results. Only ~57% of participants provided information on measurement uncertainty in a complete and consistent manner, it is apparent therefore that despite the overall satisfactory outcome of this CT round, there is still improvement needed in this crucial area.JRC.I.3-Molecular Biology and Genomic

Comparative Testing Report on the Detection and Quantification of GM Events in Rice Noodles

The European Union Reference Laboratory for Genetically Modified Food and Feed (EU-RL GMFF), accredited under ISO/IEC 17043, organised a comparative testing (CT) round for National Reference Laboratories (NRLs) nominated under Regulation (EC) No 882/2004, with voluntary participation of other official control laboratories. The test items consisted of rice noodles and commercial soybeans spiked with ground powder of soybean event DP-356043-5 in two different concentrations (Level 1 and 2). Participants were required to perform species identification and test for the presence of any GM event in the two test items. Any event detected then had to be quantified. Participants could report the results in mass/mass % or copy/copy % and the EU-RL GMFF calculated the robust means (R) for Level 1 and 2 test items accordingly. The target standard deviation for CT was fixed by the Advisory Board for Comparative Testing at 0.2 for the event, based on the experience of previous CT rounds. The robust means and target standard deviation were used to derive z-scores for the participants’ results. Eighty-eight laboratories from 42 countries registered for this CT round, of which 85 laboratories from 41 countries returned at least qualitative test results. When performing species identification, almost all laboratories correctly identified soybean and rice in both test items, and a few laboratories also detected maize and/or oilseed rape. In total 71 laboratories reported the presence of GM material in the test items, but 14 laboratories failed in this task. All of the 71 laboratories, except eight, correctly identified soybean event DP-356043-5 in the test items. Results of the quantitative evaluation of the GM content were satisfactory for both measurement units, with only two NRLs appointed under Regulation (EC) No 1981/2006 (one measuring in m/m % and one in cp/cp %) obtaining unsatisfactory z-scores (|z| ≥ 2.0) for both test items. Despite the overall satisfactory outcome of this CT round, only 58 % of participants provided information on measurement uncertainty in a complete and consistent manner, and further improvement in this crucial area is needed.JRC.I.3-Molecular Biology and Genomic

Comparative Testing Report on the Quantification of Maize Line DAS-59122-7 and Oilseed rape Line GT73 (RT73) - Comparative testing round: ILC-EURL-GMFF-CT-01/12

In the frame of Regulation (EC) No 882/2004, the European Union Reference Laboratory for Genetically Modified Food and Feed has the duty to organise comparative testing rounds and to ensure an appropriate follow-up of these activities. This report describes the outcome of the fifth comparative testing round ILC-EURL-GMFF-CT-01/12. Participants had to determine the content of oilseed rape event GT73 and maize event 59122 in two test items denoted genomic DNA levels 1 and 2, containing different GM percentages of both GM events. This comparative testing round was organised in collaboration with the Food Safety and Quality Unit of the Institute for Reference Materials and Measurements (Geel, BE). The test items were produced in-house. The Food Safety and Quality Unit managed the on-line registration and submission of results. A total of 160 laboratories were invited to participate in ILC-EURL-GMFF-CT-01/12. Eighty laboratories from 36 countries returned results, of which 59 were National Reference Laboratories, six were only members of the European Network of GMO Laboratories, three were only Official control laboratories and 12 were laboratories from third countries. Five laboratories including one National Reference Laboratory and four laboratories from third countries did not submit results. In this fifth comparative testing round 92 % to 98 % of participants gained a satisfactory z-score in the range of -2 to +2 for the results expressed in mass/mass % depending on the GM content and the GM event. However, a lower percentage (38 – 93 %) of z-scores within the working range of -2 to +2 was calculated for those participants that expressed the results in copy/copy %.JRC.I.3-Molecular Biology and Genomic

A systems approach to architecting a mission package for LCS support of amphibious operations

The United States Navy andMarine Corps have identified capability gaps in the areas of collecting Intelligence, Surveillance, Reconnaissance, and Targeting (ISR&T) data and employing fires during amphibious operations. The Littoral Combat Ship (LCS) presents an opportunity to deploy specific mission capabilities in the amphibious theater. This paper identifies the operational, functional, and physical architecture of an LCS Amphibious Warfare Mission Package (LAMP) necessary to provide capabilities associated with ISR&T data collect and fires employment. Physical architecture configurations are evaluated using a discrete-event model. Cost estimates for each alternative are presented in order to identify the LAMP architecture that provides the most cost-effective solution for providing capabilities associated with ISR&T data collection and fires employment. This paper concludes by identifying potential LAMP assets that would provide cost-effective support of amphibious operations. Four feasible alternatives are ultimately identified as cost-effective solutions, with LCCEs ranging from $105.49M and$188.22M and providing varying levels of effectiveness in terms of average engagement time and percentage of threats successfully affected.http://archive.org/details/asystemspproacht1094543992Approved for public release; distribution is unlimited

Comparative Testing Report on the Detection and Quantification of GM events in biscuit powder; Comparative testing round: ILC-EURL-GMFF-CT-01/13 - Version b

The European Union Reference Laboratory for Genetically Modified Food and Feed (EU-RL GMFF), accredited under ISO 17043, organised a comparative testing (CT) round for National Reference Laboratories (NRLs) nominated under Regulation (EC) No 882/2004, with voluntary participation of other official control laboratories. The test items consisted of biscuit powder made from conventional maize and wheat grain spiked with powder of 98140 maize and MON 863 X MON 810 maize in different concentrations (Level 1 and 2). Participants were required to perform species identification and test for the presence of maize events 3272, Bt11, Bt176, 59122, GA21, MIR604, MON 810, MON 863, NK603, 1507, MON 88017, MON 89034, 98140 and MIR162 in the two test items. Any event detected then had to be quantified. Results could be reported in mass/mass % or copy/copy % and the EU-RL calculated the robust means (R) of Level 1 and 2 test items accordingly. The target standard deviation for CT was fixed by the Advisory Board for Comparative Testing at 0.20 for all events, based on the experience of previous CT rounds. The robust means and target standard deviation were used to derive z-scores for the participants’ results. Sixty-four laboratories from 28 countries registered for this CT round, of which all but one returned at least qualitative test results. The EU-RL GMFF issued a final report on this CT round in April 2014. After the final report was prepared, it was discovered that the distribution of the quantitative data obtained from laboratories for event MON 863 deviated from normality because some laboratories used the adh1 reference gene (with target amplicon of 70 bp) for quantification of MON 863 maize, which is known to carry a nucleotide polymorphism that may affect amplification. Therefore, the robust means for event MON 863 were re-calculated based on the data from laboratories that had used a reference gene different from adh1-70 bp. The original final report has been re-called; the current version contains the re-calculated z-scores for maize event MON 863. When performing species identification, almost all laboratories correctly identified maize species in each test item. In addition, approximately half of laboratories also identified soybean and a few oilseed rape. The majority of laboratories correctly detected the three GM events included in the Level 1 and Level 2 test samples. Results of the quantitative evaluation of the GM content were compromised by the use of the adh1 gene for MON 863 quantification by a number of laboratories. Forty five laboratories (71 %) obtained satisfactory z-scores (|z| ≤ 2.0) for the three GM events in both test items. Among the remaining 18 laboratories, half had unsatisfactory z-scores for MON 863. Only 46 % of participants provided information on measurement uncertainty in a complete and consistent manner and further improvement in this crucial area is needed.JRC.I.3-Molecular Biology and Genomic

Comparative Testing Report on the Detection and Quantification of Soybean 40-3-2 and Maize MON 88017 Comparative testing round: ILC-EURL-GMFF-CT-02/12 - Preliminary report

In the frame of Regulation (EC) No 882/2004, the European Union Reference Laboratory for Genetically Modified Food and Feed has the duty to organise comparative testing rounds and to ensure an appropriate follow-up of these activities. This preliminary report describes the outcome of the sixth comparative testing round ILC-EURL-GMFF-CT-02/12. Participants were required to screen two test items denoted compound feedstuff levels 1 and 2, for the presence of maize events Maize MON 88017, MON 89034 and soybean events 356043, 40-3-2 and MON 89788. Any events detected were then to be quantified. The test items were produced in-house. The EU-RL GMFF managed the on-line registration and submission of results. A total of 160 laboratories were invited to participate in ILC-EURL-GMFF-CT-02/12. Participants could report the results of the exercise either in mass/mass % or in copy/copy %. Overall, in this sixth comparative testing round results expressed in m/m % were characterised by a higher percentage of z-scores lying within the working range of -2 to +2 with respect to results reported in cp/cp % (mean 94.37 % versus 78.76 %, estimated only on z-scores calculated on the basis of the robust means). However, The differences in the percentages of z-scores within the working range between m/m % and cp/cp % results for maize event MON88017 (mean 96.08 % versus 71.57 % for m/m % and cp/cp % respectively, estimated only on Z calculated on the basis of the robust means) appear more relevant than what can be seen for soybean event 40-3-2 (mean 92.66 % versus 85.95 % for m/m % and cp/cp % respectively, estimated only on z-scores calculated on the basis of the robust means). Moreover, a relevant difference between the assigned value and the robust mean for level 2 concentration of maize event MON 88017 was observed. Such a difference occurred because of a substantial under-estimation of the actual concentration level by 43 out of 51 laboratories (i.e. 84.31 %) reporting results in m/m % format.JRC.I.3-Molecular Biology and Genomic

A Novel Reduced-Intensity Conditioning Regimen for Unrelated Umbilical Cord Blood Transplantation in Children with Nonmalignant Diseases

Reduced-intensity conditioning (RIC) regimens have the potential to decrease transplantation-related morbidity and mortality. However, engraftment failure has been prohibitively high after RIC unrelated umbilical cord blood transplantation (UCBT) in chemotherapy-naïve children with nonmalignant diseases (NMD). Twenty-two children with a median age of 2.8 years, many with severe comorbidities and prior viral infections, were enrolled in a novel RIC protocol consisting of hydroxyurea, alemtuzumab, fludarabine, melphalan, and thiotepa followed by single UCBT. Patients underwent transplantation for inherited metabolic disorders (n = 8), primary immunodeficiencies (n = 9), hemoglobinopathies (n = 4) and Diamond Blackfan anemia (n = 1). Most umbilical cord blood (UCB) units were HLA-mismatched with median infused total nucleated cell dose of 7.9 × 10(7)/kg. No serious organ toxicities were attributable to the regimen. The cumulative incidence of neutrophil engraftment was 86.4% (95% confidence interval [CI], 65% to 100%) in a median of 20 days, with the majority sustaining > 95% donor chimerism at 1 year. Cumulative incidence of acute graft-versus-host disease (GVHD) grades II to IV and III to IV by day 180 was 27.3% (95% CI, 8.7% to 45.9%) and 13.6% (95 CI, 0% to 27.6%), respectively. Cumulative incidence of extensive chronic GVHD was 9.1% (95% CI, 0% to 20.8%). The primary causes of death were viral infections (n = 3), acute GVHD (n = 1) and transfusion reaction (n = 1). One-year overall and event-free survivals were 77.3% (95% CI, 53.7% to 89.8%) and 68.2% (95% CI, 44.6% to 83.4%) with 31 months median follow-up. This is the first RIC protocol demonstrating durable UCB engraftment in children with NMD. Future risk-based modifications of this regimen could decrease the incidence of viral infections. (www.clinicaltrials.gov/NCT00744692)