Microbiome characterization and nitrogen transformation on Angkor sandstone monuments

Abstract

Stone cultural heritages are threatened by deterioration caused by natural and anthropogenic processes, among which microorganisms are involved. Metagenomic analysis of the Angkorian temple Preah Vihear was carried out by applying whole-genome shotgun sequencing (WGS) techniques. Results revealed phylogenetic groups and potential metabolic reactions of geo-microbiological cycles in the microbiome with the presence of sulfur- and ammonia-oxidizing microorganisms retrieved from this metagenomic dataset indicating acid-producing processes contributing to the deterioration of sandstone cultural heritage. At the same time, the accumulation of nitrate has been widely observed at different Angkor monuments. The relatively low activity of denitrification and the inactive of the anaerobic ammonium oxidation (Anammox) biochemical reactions on Angkor sandstone monuments were confirmed by 15N isotope amendment, incubation, and quantification. The nitrate production by ammonia/ammonium oxidation cannot be balanced by its removal of denitrification and Anammox on Angkor monuments resulting in a net accumulation. In this study, N transformation microorganisms, namely ammonia oxidizing bacteria (AOB), archaea (AOA), and complete ammonia oxidizing bacteria (Comammox) in stone-dwelling microbiome were also further examined from three Angkor monuments by DNA-based metagenomics and RNA-based functional gene reverse-transcriptional (RT)-qPCR quantification. The metagenomic results showed the AOA and Comammox were abundant ammonia oxidizers together with the dissimilatory nitrate reduction to ammonium (DNRA) which could drive internal recycling between ammonia/ammonium and nitrate to support the stone-dwelling microbiome. The activity of this internal cycle is further supported by the significantly high enrichment of stable isotope 15N of nitrate in the microbial biofilms on the surface of Angkor monuments. And the expression of amoA genes by RNA-based RT-qPCR confirmed that AOAs were much more active than AOB in the microbiome on these Angkor monuments. Bioinformatics meta-analysis was further conducted on metagenomes of stone monuments in different climatic regions to obtain the distribution characteristics of key community members in stone-dwelling microbiome on stone deterioration processes under different climatic conditions of the world. The results of this large data analysis are in good agreement with those obtained in Cambodia and further expanded insights were also enlightened for other climate types. Higher microbial diversity is found in dry regions than tropic, but the high abundance of AOAs was detected in humid regions. Angkor sandstone monuments attract millions of visitors from around the world and are an important contributor to the local economy. The growth of tourism poses public health-related risks, because these popular sites become vectors for mixing microorganisms from humans, animals, and the ecosystem to amplify pathogenicity and resistome diffusivity. The microbiome and resistome profiling of selected geo-ecologically variable Angkor monuments revealed that pathogens and resistome of the microbiome are impacted by anthropogenic influences. A group of antibiotic-resistant genes (ARGs) with both high trans-ecological niche and trans-spatial dispersal ability was found on these monuments, suggesting a latent health-related risk of pathogens and antibiotic resistance genes transmission on the Angkor monuments with a high density of tourism activity.