Identification of Microorganisms Dwelling on the 19th Century Lanna Mural Paintings from Northern Thailand Using Culture-Dependent and -Independent Approaches

Abstract

Lanna painting is a unique type of painting in many temples in the Northern Thai region. Similar to most mural paintings, they usually decay over time partly due to the activity of microbes. This study aimed to investigate the microorganisms from two Lanna masterpiece paintings in two temples that differ in the numbers of visitors using both culture-dependent and -independent approaches. The microorganisms isolated from the murals were also tested for the biodeterioration activities including discoloration, acid production and calcium precipitation. Most microorganisms extracted from the paintings were able to discolor the paints, but only fungi were able to discolor, produce acids and precipitate calcium. The microorganism communities, diversity and functional prediction were also investigated using the culture-independent method. The diversity of microorganisms and functional prediction were different between the two temples. Gammaproteobacteria was the predominant group of bacteria in both temples. However, the fungal communities were different between the two temples as Aspergillus was the most abundant genus in the site with higher number of visitors [Buak Krok Luang temple (BK)]. Conversely, mural paintings at Tha Kham temple (TK) were dominated by the Neodevriesia genera. We noticed that a high number of visitors (Buak Krok Luang) was correlated with microbial contamination from humans while the microbial community at Tha Kham temple had a higher proportion of saprotrophs. These results could be applied to formulate a strategy to mitigate the amount of tourists as well as manage microorganism to slow down the biodeterioration process.

Keywords: Lanna mural painting; biodeterioration; microbial community; next generation sequencing.

Figure 1

Buak Krok Luang Temple (A), mural painting of the Buak Krok Luang Temple (B), Tha Kham Temple (C), mural painting of the Tha Kham Temple (D) and sampling areas within the temples, the white frames demonstrate 3 sampling replications in each area (E).

Figure 2

Biodeterioration activity of isolate Aspergillus aculeatinus (A) on crimson red and isolate Aspergillus piperis (C) on malachite green, compare with crimson red control (B) and malachite green control (D).

Figure 3

Acid production of Aspergillus fumigatus (red peak) compared with fumaric acid standard (blue peak) (A) and Calcium formation and EDS analysis of Aspergillus piperis (B).

Figure 4

Microbial community composition of bacteria (A) and fungi (B) from two mural paintings from Buak Krok Luang temple (BK) and Tha Kham temple (TK). The stacked bar-plots were derived from high-throughput sequencing data.

Figure 5

Alpha diversity of bacteria (A) and Fungi (B) of mural paintings according to Chao1, Shannon and Simpson indices in samples collected from Buak Krok Luang Temple (BK) and Tha Kham Temple (TK), black dots and error bar represent the means and Standard deviation of each barplot.

Figure 6

Non-metric Multi-dimensional Scaling (NMDS) with environment parameters of microorganism, bacteria (A) and fungi (B) on mural paintings from the two temples. The red dots represent the samples from Buak Krok Luang temple while the blue dots represent the samples from Tha Kham temple. The green arrows indicate the factors which impact on the microorganism community in each temple.

Figure 7

Ratios of fungal (A) and bacterial (B) communities based on high-throughput sequencing method illustrated in stacked-barplot. Details of each isolate are shown in the table on the right (TK; Tha Kham temple, BK; Buak Krok Luang temple).

Figure 8

Microorganism diversity comparison among different countries. The triangles are communities from tropical countries (Thailand and Brazil), while circles are communities from temperate countries (China, Georgia, Germany, Italy, Russia, Serbia and Spain).