Analysis of commercial kidney stone probiotic supplements

Abstract

Objective: To examine the levels of Oxalobacter formigenes in probiotic supplements marketed by PRO-LAB, Ltd, Toronto, Canada, and capsules of Oxalo purchased from Sanzyme Ltd, Hyderabad, India, and to measure the ability of these preparations to degrade oxalate in vitro.

Methods: Probiotic supplements and pure cultures of O. formigenes were cultured in a number of media containing oxalate. Optical density at 595 nm (OD595) was used to measure bacterial growth, and ion chromatography was used to measure loss of oxalate in culture media. O. formigenes-specific and degenerate Lactobacillus primers to the oxalate decarboxylase gene (oxc) were used in polymerase chain reaction (PCR).

Results: Incubating probiotic supplements in different media did not result in the growth of oxalate-degrading organisms. PCR indicated the absence of organisms harboring the oxc gene. Culture and 16S ribosomal ribonucleic acid gene sequencing indicated the PRO-LAB supplement contained viable Lactococcus lactis subsp. lactis (GenBank accession no. KJ095656.1), whereas Oxalo contained several Bacillus species and Lactobacillus plantarum.

Conclusion: The probiotic supplement sold over the Internet by PRO-LAB Ltd and Sanzyme Ltd did not contain identifiable O. formigenes or viable oxalate-degrading organisms, and they are unlikely to be of benefit to calcium oxalate kidney stone patients.

Figure 1

Growth of O. formigenes OxCC13. Representative growth in SBO (●), medium B (- -▲- -), and medium B supplemented with 0.5% yeast (··■··). All media contained 100mM sodium oxalate and 10mM sodium acetate. Starter cultures were grown to end of log growth (OD₅₉₅ 0.1), and then diluted 3/10,000 into each medium to monitor growth over time.

Figure 2

Amplification of oxc. (A) All Oxalobacter strains tested with oxc-specific primers exhibited a band of the expected size (1284 bp). Lanes: 1, molecular weight ladder (bp); 2, OxB; 3, BA-1; 4, Sox4; 5, OxCC13; 6, OxWR; 7, HOxBLS; 8, OxK; 9, HC1; 10, OxGP; 11, no DNA control; 12, purified DNA from CC13. (B) Amplification of oxc using oxc-specific primers (lanes 2–5) and degenerate oxc primers (lanes 6–9, expected size 419 bp) and DNA extracted from ™PRO Lab supplement powder, and BHI cultures of ™PRO Lab supplement. Lanes: 1, molecular weight ladder (bp); 2, BHI ™PRO Lab supplement DNA; 3, ™PRO Lab supplement DNA; 4, no DNA control; 5, purified DNA from CC13; 6, BHI ™PRO Lab DNA; 7, ™PRO Lab DNA; 8, no DNA control; 9, purified DNA from CC13. (C) Amplification of oxc using oxc-specific primers and DNA extracted from Oxalo™ capsule. Lanes: 1, molecular weight ladder (bp); 2, Oxalo™ capsule DNA; 3, purified DNA from OxCC13. (D) Amplification of oxc using degenerate oxc primers and DNA extracted from Oxalo™ capsule. Lanes: 1, molecular weight ladder (bp); 2, Oxalo™ capsule DNA; 3, purified DNA from OxCC13.

Figure 3

Sequence alignment of 16S rRNA from ™PRO Lab supplement powder. This PCR product was 98% identical with GenBank accession no. KJ095656.1, Lactococcus lactis subsp. lactis.