Fungal colonization can severely damage artifacts. Nematode endosymbiotic bacteria exhibit good prospects in protecting artifacts from fungal damage. We previously found that supernatant from the fermentation of nematode endosymbiotic bacterium, Xenorhabdus bovienii, is effective in inhibiting the growth of Fusarium solani NK-NH1, the major disease fungus in the Nanhai No.1 Shipwreck. Further experiments proved that X. bovienii produces volatile organic compounds (VOCs) that inhibit NK-NH1. Here, using metabolomic analysis, GC–MS, and transcriptomic analysis, we explored the antifungal substances and VOCs produced by X. bovienii and investigated the mechanism underlying its inhibitory effect against NK-NH1. We show that X. bovienii produces several metabolites, mainly lipids and lipid-like molecules, organic acids and derivatives, and organoheterocyclic compounds. The VOCs produced by X. bovienii showed two specific absorption peaks, and based on the library ratio results, these were predicted to be of 2-pentanone, 3-(phenylmethylene) and 1-hexen-3-one, 5-methyl-1-phenyl. The inhibition of F. solani by VOCs resulted in upregulation of genes related to ribosome, ribosome biogenesis, and the oxidative phosphorylation and downregulation of many genes associated with cell cycle, meiosis, DNA replication, and autophagy. These results are significant for understanding the inhibitory mechanisms employed by nematode endosymbiotic bacteria and should serve as reference in the protection of artifacts.

Nanhai No. 1, a shipwreck in the Southern Song Dynasty, China, has a history of more than 800 years. It was salvaged in 2007 and is now on display in the Guangdong Maritime Silk Road Museum. Due to the fact that the hull is a wooden cultural relic and exposed to the air, the biological corrosion and biodegradation caused by microorganisms are key problems of hull protection. At present, the antimicrobial agent Euxyl^® K100 (isothiazolinone) has a significant antimicrobial effect in the field, but it has a certain negative impact on the environment and archeologists. In order to reduce the use of chemical antimicrobial agents, we evaluated the inhibitory effects of cinnamaldehyde on the main destructive microorganisms of Nanhai No. 1. Cinnamaldehyde is the main active component of cinnamon, and has broad-spectrum antimicrobial properties. The paper diffusion method, gas diffusion method and minimum inhibitory concentration experiment were used to detect the inhibitory effects of cinnamaldehyde on the main microorganisms of Nanhai No. 1. We found that cinnamaldehyde had significant inhibitory effects on Bacillus tequilensis NK-NH5, Bacillus megaterium NK-NH10, Bacillus velezensis NK-NH11, Bacillus sp. NK-NH15, Bacillus sp. NK-NH16, Bacillus sp. NK-NH17, Fusariumsolani NK-NH1 and Scedosporiumapiospermum NK.W1-3. At the same time, cinnamaldehyde had more inhibitory effects on fungi than bacteria. Finally, we verified that cinnamaldehyde can effectively inhibit the growth of microorganisms in water, for storing the scattered wood blocks of the Nanhai No. 1 hull through laboratory simulation experiments. Cinnamaldehyde, as an environment-friendly antimicrobial agent, is of great significance to protecting water-saturated wooden relics from microbial corrosion and degradation in the future.

The Nanhai No. 1 was a wooden merchant ship of the Southern Song Dynasty, which wrecked and sank in the South China Sea, Yangjiang City, Guangdong Province, China. The Nanhai No. 1 shipwreck was salvaged as a whole in 2007 and began to be excavated in 2013. During the archaeology excavation, some of the hull wood fell off the hull. These waterlogged archaeological woods (WAW) were immersed in the buffer containing EDTA-2Na and isothiazolinone K100 for moisture stabilization, preliminary desalination, and microbial inhibition. We evaluated the properties of these WAW through testing the chemical components (including lignin, holocellulose, and ash content) of the wood, and monitoring the iron element content, anion and cation content in the buffer. At the same time, the microbial composition in the desalination buffer was also detected. The results showed that the holocellulose content in these WAW were much lower than in fresh wood. The ash content in these WAW decreased after desalination treatment. The iron element content, anion and cation content in the buffer were high and kept at a certain level after desalination treatment. At the same time, the problem of biodegradation in the buffer should be paid attention to. The comprehensive protection of WAW requires to combine wood properties and microbial problems. This study provides a reference for the protection of WAW from the Nanhai No. 1 shipwreck and other similar historical wood.

To clarify the microbial contamination and wood degradation risk of the Nanhai No. 1 shipwreck hull and verify on-site antibacterial agent effectiveness, microbial samples were collected and analyzed via SEM, metagenomic sequencing, bacterial isolation, enzyme activity detection, and antibacterial experiments. The results showed that Actinomycetota was the dominant phylum, and Brachybacterium, Microbacterium, and Brevibacterium were the dominant genera. Seven bacterial strains were isolated and purified, among which Brevibacterium sp. (NH.SH-B6) had the strongest wood degradation ability, possessing cellulase, LiP, MnP, and Lac activities. When cultured with hull wood as the sole carbon source, LiP was the dominant degrading enzyme of NH.SH-B6, and its maximum enzyme activity was achieved under the optimal conditions of pH = 7, 10% NaCl, 1000 mg/L FeSO₄, and no PEG400 added. 50 mg/mL cinnamaldehyde and 0.5% isothiazolinone K100 had good inhibitory effects on the isolated bacteria, and bacterial proliferation was due to incomplete antibacterial agent spraying. This study clarifies the microbial degradation risk of the Nanhai No. 1 shipwreck hull and provides a scientific basis for optimizing the on-site protection strategy of the shipwreck.