The role of ZIP transporters and group F bZIP transcription factors in the Zn-deficiency response of wheat (Triticum aestivum).

Understanding the molecular basis of zinc (Zn) uptake and transport in staple cereal crops is critical for improving both Zn content and tolerance to low-Zn soils. This study demonstrates the importance of group F bZIP transcription factors and ZIP transporters in responses to Zn deficiency in wheat (Triticum aestivum). Seven group F TabZIP genes and 14 ZIPs with homeologs were identified in hexaploid wheat. Promoter analysis revealed the presence of Zn-deficiency-response elements (ZDREs) in a number of the ZIPs. Functional complementation of the zrt1/zrt2 yeast mutant by TaZIP3, -6, -7, -9 and -13 supported an ability to transport Zn. Group F TabZIPs contain the group-defining cysteine-histidine-rich motifs, which are the predicted binding site of Zn2+ in the Zn-deficiency response. Conservation of these motifs varied between the TabZIPs suggesting that individual TabZIPs may have specific roles in the wheat Zn-homeostatic network. Increased expression in response to low Zn levels was observed for several of the wheat ZIPs and bZIPs; this varied temporally and spatially suggesting specific functions in the response mechanism. The ability of the group F TabZIPs to bind to specific ZDREs in the promoters of TaZIPs indicates a conserved mechanism in monocots and dicots in responding to Zn deficiency. In support of this, TabZIPF1-7DL and TabZIPF4-7AL afforded a strong level of rescue to the Arabidopsis hypersensitive bzip19 bzip23 double mutant under Zn deficiency. These results provide a greater understanding of Zn-homeostatic mechanisms in wheat, demonstrating an expanded repertoire of group F bZIP transcription factors, adding to the complexity of Zn homeostasis.

A comparison of precipitation and filtration-based SARS-CoV-2 recovery methods and the influence of temperature, turbidity, and surfactant load in urban wastewater.

Wastewater-based epidemiology (WBE) has become a complimentary surveillance tool during the SARS-CoV-2 pandemic. Viral concentration methods from wastewater are still being optimised and compared, whilst viral recovery under different wastewater characteristics and storage temperatures remains poorly understood. Using urban wastewater samples, we tested three viral concentration methods; polyethylene glycol precipitation (PEG), ammonium sulphate precipitation (AS), and CP select™ InnovaPrep® (IP) ultrafiltration. We found no major difference in SARS-CoV-2 and faecal indicator virus (crAssphage) recovery from wastewater samples (n = 46) using these methods, PEG slightly (albeit non-significantly), outperformed AS and IP for SARS-CoV-2 detection, as a higher genome copies per litre (gc/l) was recorded for a larger proportion of samples. Next generation sequencing of 8 paired samples revealed non-significant differences in the quality of data between AS and IP, though IP data quality was slightly better and less variable. A controlled experiment assessed the impact of wastewater suspended solids (turbidity; 0-400 NTU), surfactant load (0-200 mg/l), and storage temperature (5-20 °C) on viral recovery using the AS and IP methods. SARS-CoV-2 recoveries were >20% with AS and <10% with IP in turbid samples, whilst viral recoveries for samples with additional surfactant were between 0-18% for AS and 0-5% for IP. Turbidity and sample storage temperature combined had no significant effect on SARS-CoV-2 recovery (p > 0.05), whilst surfactant and storage temperature combined were significant negative correlates (p < 0.001 and p < 0.05, respectively). In conclusion, our results show that choice of methodology had small effect on viral recovery of SARS-CoV-2 and crAssphage in wastewater samples within this study. In contrast, sample turbidity, storage temperature, and surfactant load did affect viral recovery, highlighting the need for careful consideration of the viral concentration methodology used when working with wastewater samples.

An analysis of 45 large-scale wastewater sites in England to estimate SARS-CoV-2 community prevalence.

Accurate surveillance of the COVID-19 pandemic can be weakened by under-reporting of cases, particularly due to asymptomatic or pre-symptomatic infections, resulting in bias. Quantification of SARS-CoV-2 RNA in wastewater can be used to infer infection prevalence, but uncertainty in sensitivity and considerable variability has meant that accurate measurement remains elusive. Here, we use data from 45 sewage sites in England, covering 31% of the population, and estimate SARS-CoV-2 prevalence to within 1.1% of estimates from representative prevalence surveys (with 95% confidence). Using machine learning and phenomenological models, we show that differences between sampled sites, particularly the wastewater flow rate, influence prevalence estimation and require careful interpretation. We find that SARS-CoV-2 signals in wastewater appear 4-5 days earlier in comparison to clinical testing data but are coincident with prevalence surveys suggesting that wastewater surveillance can be a leading indicator for symptomatic viral infections. Surveillance for viruses in wastewater complements and strengthens clinical surveillance, with significant implications for public health.