Ion chromatography

Radioactive sulfate was used to determine the sulfate-reduction rates (SRR) in camel feces. Feces were placed in sterile 5 mL syringes sealed with butyl rubber stoppers, which received aliquots (300 µL) of Na₂³⁵SO₄ (3 µCi ‘Perkin-Elmer’, Waltham, MA, USA) by injection through the butyl rubber stopper. The syringes were incubated in the dark at 37 °C, for 24 h followed by the addition of 1 mL of 2M KOH to terminate the reaction and fix sulfide. Radioactivity was measured in the acid volatile sulfide (AVS), H₂S and FeS, and chromium-reducible sulfur (CRS) fractions, which included pyrite, and elemental and organic sulfur, as previously described [25 (link),26 (link)]. Sulfate concentration was analyzed by ion chromatography (Dionex). The average SRR and standard deviation were calculated from triplicate incubations.

Medium pH was measured using a standard bench top pH meter. Nitrate concentrations were measured using ion chromatography (Dionex, Sunnyvale, CA), respectively, using previously described protocols [34 (link)].

The content of elements (C, H, N) was determined with a Yanaco CHN CORDER MT-6 Elemental Analyzer, and a Dionex Ion Chromatography was used to analyze the content of sulfur.

The abilities of isolates to grow via the disproportionation of thiosulfate (10 mM), elemental sulfur (1% wt/vol), and sulfite (1, 5 mM) were tested with the MMJS medium in the absence or presence of Fe (III). All of the experiments were performed in duplicate. In addition, appropriate control experiments were performed in order to differentiate biotic and abiotic reactions. Serum bottles were homogenized via manual shaking, and aliquots of the cultures were sampled using a syringe that was nitrogen-flushed prior to sampling. The samples were processed immediately for further analyses in order to minimize oxygen exposure. Bacterial growth was measured via direct cell counts using a phase-contrast microscope (Eclipse 80i, Nikon). Before counting, the iron precipitates were dissolved using a dithionite solution (5% in 0.3 M acetic acid) (57 (link)). The dissolved sulfide concentration was determined spectrophotometrically using the methylene blue method, according to Cline (1969) (58 (link)). Thiosulfate and sulfate were analyzed via ion chromatography (Dionex, USA) after separation on an IonPac AG19 column (4 × 250 mm) with NaOH (20 mM) as the eluent at a flow rate of 1 mL/min. Fe (III) reduction was monitored by quantitatively measuring Fe (II) using the ferrozine method (59 (link)). The samples for the assay were prepared as described previously (60 (link)).

The monosaccharide composition of LBP was determined by ion chromatography [58 (link)]. Then, 5 mg of LBP was precision weighed in ampoan ule bottle, 2 mL 3 mol/L trifluoroacetic acid as added, and the mixture was hydrolyzed at 120 °C for 3 h. The acid hydrolysis solution was transferred to the tube and dried by nitrogen blowing. Next, 5 mL of deionized water was added, dissolved and mixed. Then, 50 μL was added to 950 μL deionized water and centrifuged at 12,000 rpm for 5 min. The supernatant was analyzed by ion chromatography (Thermo Fisher, Shanghai, China). Chromatographic conditions: chromatographic column: Dionex Carbopac^TM PA20 (3 × 150 mm); mobile phase: A: H₂O; B: 15 mM NaOH:100 mM NaOAC (1:1); flow rate: 0.3 mL/min; sample size: 5 μL; column temperature: 30 °C; detector: Electrochemical detector.
The results were compared with 16 monosaccharide standards (fucose (Fuc), aminogalactose hydrochloride (GalN), rhamnose (Rha), arabinose (Ara), glucosamine hydrochloride (GlcN), galactose (Gal), glucose (Glu), xylose (Xyl), mannose (Man), fructose (Fru), ribose (Rib), galactose aldehyde acid (GalA), glucuronic acid (GlcA), N-acetyl-D-glucosamine (GlcNAc), guluronic acid (GulA), and mannose aldehyde acid (ManA)) to determine the monosaccharide species and molar ratio.