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The survival of Clostridioides difficile (previously Clostridium difficile) vegetative cells and endospores was compared at different levels of indigenous microflora using autoclaved and unautoclaved dairy composts with different moisture contents (MCs).Both types of composts adjusted to 20, 30 and 40% MCs were inoculated with a suspension of C. difficile that contained both vegetative cells (c. 5-6 log CFU per gram) and endospores (c. 5·0 CFU per gram), and then stored aerobically inside a humidity-controlled chamber at room temperature 22·5 ± 0·8°C for 1 year. The level of indigenous microflora was very stable during the storage after day 7 in both types of compost. The greatest reductions of C. difficile vegetative cell counts occurred during the first 24 h of storage in autoclaved and unautoclaved composts, which had 4·7 and 5·5 log CFU per gram with 20% MC, 1·8 and 2·1 log CFU per gram with 30% MC, and 2·3 and 1·3 log CFU per gram with 40% MC, respectively. Both MC and the duration of storage have significant (P 0·05) effects on the survival of vegetative cells for first 120 days of storage. The slow inactivation of C. difficile vegetative cells at higher MCs during aerobic storage was confirmed by exponentially decaying modelling data during the early stage of aerobic exposure. The reduction of endospore counts (1·0 log CFU per gram) during the storage for both types of compost at all MCs was not significant (P 0·05) except for the autoclaved compost with 30% MC.The highly resistant C. difficile endospores to the unfavourable environmental conditions survived for more than a year while vegetative cells died off exponentially upon the initial aerobic exposure.The long-term survival of C. difficile endospores in contaminated compost may transmit the pathogen to fresh produce, animals or water in pre-harvest conditions.

Drought stress severely limits the growth, development, and productivity of crops, and therefore understanding the mechanisms by which plants respond to drought is crucial. In this study, we cloned a maize NAC transcription factor, ZmNAC49, and identified its function in response to drought stress. We found that ZmNAC49 is localized in the nucleus and has transcriptional activation activity. ZmNAC49 expression is rapidly and strongly induced by drought stress, and overexpression enhances stress tolerance in maize. Overexpression also significant decreases the transpiration rate, stomatal conductance, and stomatal density in maize. Detailed study showed that ZmNAC49 overexpression affects the expression of genes related to stomatal development, namely ZmTMM, ZmSDD1, ZmMUTE, and ZmFAMA. In addition, we found that ZmNAC49 can directly bind to the promoter of ZmMUTE and suppress its expression. Taken together, our results show that the transcription factor ZmNAC49 represses ZmMUTE expression, reduces stomatal density, and thereby enhances drought tolerance in maize.

Mitogen-activated protein kinase (MPK) is a critical regulator of the antioxidant defence system in response to various stimuli. However, how MPK directly and exactly regulates antioxidant enzyme activities is still unclear. Here, we demonstrated that a NAC transcription factor ZmNAC49 mediated the regulation of antioxidant enzyme activities by ZmMPK5. ZmNAC49 expression is induced by oxidative stress. ZmNAC49 enhances oxidative stress tolerance in maize, and it also reduces superoxide anion generation and increases superoxide dismutase (SOD) activity. A detailed study showed that ZmMPK5 directly interacts with and phosphorylates ZmNAC49 in vitro and in vivo. ZmMPK5 directly phosphorylates Thr-26 in NAC subdomain A of ZmNAC49. Mutation at Thr-26 of ZmNAC49 does not affect the interaction with ZmMPK5 and its subcellular localisation. Further analysis found that ZmNAC49 activates the ZmSOD3 expression by directly binding to its promoter. ZmMPK5-mediated ZmNAC49 phosphorylation improves its ability to bind to the ZmSOD3 promoter. Thr-26 of ZmNAC49 is essential for its transcriptional activity. In addition, ZmSOD3 enhances oxidative stress tolerance in maize. Our results show that phosphorylation of Thr-26 in ZmNAC49 by ZmMPK5 increased its DNA-binding activity to the ZmSOD3 promoter, enhanced SOD activity and thereby improved oxidative stress tolerance in maize.