Validation of the Thermo Scientific SureTect™ Campylobacter jejuni, C. coli, and C. lari PCR Kit for the Detection of Campylobacter jejuni, C. coli, and C. lari in Raw Poultry and Ready-to-Cook Poultry Products: AOAC Performance Tested MethodSM 012101.

BACKGROUND: The Thermo Scientific SureTect™ Campylobacter jejuni, C. coli, and C. lari PCR Kit is a real-time PCR assay for the detection and differentiation of C. jejuni, C. coli, and C. lari from raw poultry, ready-to-cook poultry products, and environmental samples. OBJECTIVE: The Thermo Scientific SureTect Campylobacter jejuni, C. coli, and C. lari PCR Kit was evaluated for AOAC®Performance Tested MethodsSM certification. METHODS: Inclusivity/exclusivity, matrix studies, product consistency and stability, and robustness testing were conducted to assess the method's performance. In the matrix studies, the method was validated against United States and international reference methods for Campylobacter detection. RESULTS: There were no statistically significant differences found in the matrix studies between the candidate and reference methods when analyzed by probability of detection. All 52 inclusivity strains and none of the 51 exclusivity strains tested were detected by the assay. Robustness testing demonstrated that the assay gave reliable performance with specific method deviations outside of the recommended parameters, and the real-time stability testing demonstrated that there were no statistically significant differences between kit lots, validating the stated shelf life of the kit. CONCLUSION: The data presented support the product claims that the Thermo Scientific SureTect Campylobacter jejuni, C. coli, and C. lari PCR assay is suitable for the detection and differentiation of C. jejuni, C. coli, and C. lari from raw poultry, ready-to-cook poultry products, and environmental samples. HIGHLIGHTS: Presumptive results can be obtained in as little as 23 h. Microaerophilic incubators are not required for enrichment.

Validation of the Thermo Scientific™ SARS-CoV-2 RT-PCR Detection Workflow for the Detection of SARS-CoV-2 from Stainless-Steel Environmental Surface Swabs: AOAC Performance Tested MethodSM 012103.

BACKGROUND: The Thermo Scientific™ SARS-CoV-2 reverse transcription-polymerase chain reaction (RT-PCR) Detection Workflow, packaged with Applied Biosystems™ TaqMan™ 2019-nCoV Assay Kit v1 targets three different SARS-CoV-2 genomic regions in a single RT-PCR reaction. OBJECTIVE: To validate the Thermo Scientific SARS-CoV-2 RT-PCR Workflow, for the detection of SARS-CoV-2 virus on stainless-steel surfaces as part of the AOAC Performance Tested MethodSM Emergency Response Validation program. METHOD: The Applied Biosystems TaqMan 2019-nCoV Assay Kit v1, as part of the Thermo Scientific SARS-CoV-2 RT-PCR Workflow, was evaluated for specificity using in silico analysis of 15 764 SARS-CoV-2 sequences and 65 exclusivity organisms. The Thermo Scientific SARS-CoV-2 RT-PCR Workflow was evaluated in an unpaired study for one environmental surface (stainless steel) and compared to the U.S. Centers for Disease Control and Prevention 2019-Novel Coronavirus RT-PCR Diagnostic Panel, Instructions for Use (Revision 4, Effective 6/12/2020). RESULTS: In silico analysis showed that, of the 15 756 target SARS-CoV-2 genomes analyzed, 99% of the strains/isolates are perfectly matched to at least two of the three assays, and more than 90% have 100% homology to all three assays (ORF1ab, N-gene, S-gene) in the SARS-CoV-2 Kit. None of the 65 non-target strain genomes analyzed showed matching sequences. In the matrix study, the Thermo Scientific SARS-CoV-2 workflow showed comparable detection to the centers of disease control and prevention (CDC) method. CONCLUSIONS: The Thermo Scientific SARS-CoV-2 RT-PCR Workflow is an effective procedure for detection of RNA from SARS-CoV-2 virus from stainless steel. HIGHLIGHTS: The workflow provides equivalent performance results with the two tested RNA extraction platforms and the two tested RT-PCR instruments.

Improving photosynthesis for algal biofuels: toward a green revolution.

Biofuels derived from marine algae are a potential source of sustainable energy that can contribute to future global demands. The realisation of this potential will require manipulation of the fundamental biology of algal physiology to increase the efficiency with which solar energy is ultimately converted into usable biomass. This 'photosynthetic solar energy conversion efficiency' sets an upper limit on the potential of algal-derived biofuels. In this review, we outline photosynthetic molecular targets that could be manipulated to increase the efficiency and yield of algal biofuel production. We also highlight modern 'omic' and high-throughput technologies that might enable identification, selection and improvement of algal cell lines on timescales relevant for achieving significant contributions to future energy solutions.

PIF3 is a repressor of chloroplast development.

The phytochrome-interacting factor PIF3 has been proposed to act as a positive regulator of chloroplast development. Here, we show that the pif3 mutant has a phenotype that is similar to the pif1 mutant, lacking the repressor of chloroplast development PIF1, and that a pif1pif3 double mutant has an additive phenotype in all respects. The pif mutants showed elevated protochlorophyllide levels in the dark, and etioplasts of pif mutants contained smaller prolamellar bodies and more prothylakoid membranes than corresponding wild-type seedlings, similar to previous reports of constitutive photomorphogenic mutants. Consistent with this observation, pif1, pif3, and pif1pif3 showed reduced hypocotyl elongation and increased cotyledon opening in the dark. Transfer of 4-d-old dark-grown seedlings to white light resulted in more chlorophyll synthesis in pif mutants over the first 2 h, and analysis of gene expression in dark-grown pif mutants indicated that key tetrapyrrole regulatory genes such as HEMA1 encoding the rate-limiting step in tetrapyrrole synthesis were already elevated 2 d after germination. Circadian regulation of HEMA1 in the dark also showed reduced amplitude and a shorter, variable period in the pif mutants, whereas expression of the core clock components TOC1, CCA1, and LHY was largely unaffected. Expression of both PIF1 and PIF3 was circadian regulated in dark-grown seedlings. PIF1 and PIF3 are proposed to be negative regulators that function to integrate light and circadian control in the regulation of chloroplast development.

Light signalling pathways regulating the Mg-chelatase branchpoint of chlorophyll synthesis during de-etiolation in Arabidopsis thaliana.

Precise regulation of tetrapyrrole synthesis is critical for plant survival when seedlings first emerge into the light. At this time there is a massive increase in demand for chlorophyll to drive the assembly of the photosynthetic apparatus. To understand how this demand is met we have followed the expression of genes encoding the chelatase enzymes at the branchpoint between chlorophyll and heme synthesis. Dark-grown Arabidopsis thaliana seedlings were transferred to continuous white, red, far-red or blue light and the expression of eight tetrapyrrole pathway genes was followed using real-time RT-PCR. Our results show that the CHLH gene encoding the H subunit of Mg-chelatase was induced by light under all conditions with an initial peak after 2-4 h light. The other Mg-chelatase subunit genes CHLI and CHLD and the ferrochelatase genes FC1 and FC2 were not strongly regulated at the level of transcript abundance, but the Mg-chelatase regulator GUN4 had an expression profile almost identical to that observed for CHLH. The CHLM gene encoding Mg-protoporphyrin IX methyltransferase, the next enzyme in the pathway, was also light regulated, but showed a very different pattern of expression. Using photoreceptor mutants it was demonstrated that regulation of CHLH and GUN4 is primarily under the control of phytochromes A and B with some input from the cryptochromes. Induction of CHLH and GUN4 under red and far-red light was also compromised in the phytochrome-signalling mutants, fhy1 and fhy3. These results establish GUN4 as a major target of photoreceptor regulation during the earliest stages of de-etiolation.