potential discolouration of Arthrobacter agilis, one of the isolates was tested on the French oolitic Savonnières limestone. This stone is extensively used as a replacement for Lede stone (Dewanckele et al., 2014) This was tested by a water run-off test with a similar setup as De Muynck et al. (2009). Arthrobacter agilis was grown in R2A broth at 20 °C, after which it was sprinkled during two hours over six slaps of Savonnières limestone and dried during 24 hours. This was repeated two times more and in between its colour at two spots on the rock was examined with a point measurement using the CM-2600d spectrophotometer.
Results and Discussion
Gypsum crusts were present both in the urban and rural environment. These were thick botryoidal and black in Ghent, while in Berlare those were thinner and laminar with a rusty colour. The occurrence of the gypsum crusts has also been confirmed by the soluble salt analysis (see Table 1 for the anions). In every sample, there was a high concentration
of soluble SO42– ranging between 1 mg/grock and 10 mg/grock. Although highly variable between the samples, overall, the soluble salt content was higher in the City Hall of Ghent compared to the samples of Berlare. This was also the case for the Cl– and NO3–. Sample B6 was the only sample of Berlare with more Cl– and NO3–, because of its sheltered position underneath a windowsill. Nitrite and phosphate were not found or only available in a very low amount. The compatible cations were primarily calcium (0.6−5 mg/grock) and in a lesser extend also potassium and magnesium. Sodium was most of the times slightly present, but could reach a high concentration (about 5 mg/grock for G4) combined with a high amount of soluble chloride. Based on the modelling performed using RUNSALT the accompanying salts (except gypsum), for the samples with high soluble salts content, at 20 °C and low RH were among others carnallite (KCl·MgCl2.6H2O), niter (KNO3), nitromagnesite, (Mg(NO3)2.6H2O), halite (NaCl), nitratine (NaNO3) and Ca(NO3)2. These salts would be in solution at a relative humidity of about 60 % and more. For the other samples, the result of the model was more complex.
Within and underneath the gypsum crusts, 16S rRNA gene sequencing successfully identified the prokaryotic community on the City hall of Ghent and the Castle of Berlare, except for one sample (G1). Sulphur oxidizing, sulphur reducing and nitrogen oxidizers have been identified in some of the samples. Genera of nitrifying bacteria from different taxa were present: Nitrolancea (0.03 % 98in G2), Nitrospira (0.06 % in G2), Nitrosomonadaceae with Nitrosospira (0.002 % in G4, 0.7 % G6 and 0.0003 % in B1) and Nitrosomonas (0.002 % in G6). Archaea have been found in only one sample (G6) belonging to the nitrifying Nitrososphaeraceae.
Table 1: Soluble anions of the different samples in the urban and rural environment (mg/grock).
| Sample | Cl– | NO3– | SO42– |
| G1 | 0.351 | 1.626 | 6.795 |
| G2 | 1.014 | 5.404 | 6.858 |
| G3 | 0.103 | 0.261 | 7.337 |
| G4 | 5.348 | 12.537 | 9.651 |
| G5 | 0.112 | 0.238 | 6.948 |
| G6 | 0.087 | 0.306 | 6.884 |
| B1 | 0.004 | 0.006 | 4.000 |
| B2 | 0.207 | 0.129 | 7.156 |
| B3 | 0.005 | 0.004 | 4.952 |
| B4 | 0.004 | 0.003 | 1.261 |
| B5 | 0.005 | 0.005 | 4.632 |
| B6 | 0.801 | 1.150 | 7.221 |
| B7 | 0.008 | 0.005 | 2.851 |
Some sulphur oxidizing bacteria were found as well, belonging to the “purple non-sulphur” bacteria (PNS). Despite their name, members of this group can oxidize low concentrations of sulphur (Hunter et al., 2009). PNS were abundant in G5 with 16 % of Rhodoplanes. Furthermore, there might be more PNS present in the samples as the order Rhodospirillales has been detected in G4 and G6 with an unclassified chemolithoautotrophic Magnetospira (0.2 % in G4) and with an unclassified genus of the family Rhodospirillaceae (0.3 % in G4). Furthermore, potential PNS in Rhodobacteraceae were represented as well, however again unclassified (0.5 % in G2, 0.002 % in G4, 0.4 % in G6 and 0.0003 % in B1) (Hunter et al., 2009; Williams et al., 2012). Beside sulphur oxidation, some identified bacteria belonged to the sulphur/sulphate reducing bacteria such as Desulfuromonadales (0.005 % in G3) and 4 % Geoalkalibacter in G5, but also Desulfobacteraceae, with Desulfofrigus (0.008 % in G5 and 0.0006 % in B4), Desulforhopalus (0.0006 % in B4) and unclassified genera (0.0009 % in G6, 0.003 % in B4 and 0.003 % in B6) (Greene, Patel and Yacob, 2009; Rosenberg et al., 2013).
There is a strong link between the location and the occurrence of the nitrogen oxidizing and sulphur bacteria. Both are present in selected samples across both locations, but mainly in the City Hall of Ghent, where in one sample they even reach high abundance. There is also a link between the occurrence of the nitrifying and sulphur prokaryotes and the higher soluble salt content of respectively nitrate or sulphate. However, also here there are some exceptions. The occurrence of sulphate reducing bacteria followed the same trend. These bacteria can cause further deterioration (Krumbein and Gorbushina, 2009) but besides inducing gypsum crust formation, they can reduce the sulphate and precipitate calcium carbonate. This process also produces sulphur and H2S (Castanier, Le Métayer-Levrel and Perthuisot, 1999). As one sample in the city (G5) contains both a high concentration of PNS and sulphate reducing bacteria, it is not impossible that both groups are strongly linked together.
These results combine some of the findings of previous research by Mansch and Bock (1998) & Villa et al. (2015). The former found more nitrifying bacteria in the urban environment and the latter more sulphur oxidizing bacteria. It also confirms the findings of Li et al. (2016). However, members of sulphur compound oxidizing genera of which it is known
