RESUMEN
AIM: To develop a novel validated method for the isolation of Bifidobacterium animalis ssp. lactis BB-12 (BB-12) from faecal specimens and apply it to studies of BB-12 and Lactobacillus rhamnosus GG (LGG) recovered from the healthy human gastrointestinal (GI) tract. METHODS AND RESULTS: A novel method for isolating and enumerating BB-12 was developed based on its morphologic features of growth on tetracycline-containing agar. The method identified BB-12 correctly from spiked stool close to 100% of the time as validated by PCR confirmation of identity, and resulted in 97-104% recovery of BB-12. The method was then applied in a study of the recovery of BB-12 and LGG from the GI tract of healthy humans consuming ProNutrients® Probiotic powder sachet containing BB-12 and LGG. Viable BB-12 and LGG were recovered from stool after 21 days of probiotic ingestion compared to baseline. In contrast, no organisms were recovered 21 days after baseline in the nonsupplemented control group. CONCLUSIONS: We demonstrated recovery of viable BB-12, using a validated novel method specific for the isolation of BB-12, and LGG from the GI tract of healthy humans who consumed the probiotic supplement. SIGNIFICANCE AND IMPACT OF THE STUDY: This method will enable more detailed and specific studies of BB-12 in probiotic supplements, including when in combination with LGG.
Asunto(s)
Bifidobacterium animalis/aislamiento & purificación , Tracto Gastrointestinal/microbiología , Lacticaseibacillus rhamnosus/fisiología , Probióticos/administración & dosificación , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Antibacterianos , Bifidobacterium animalis/clasificación , Bifidobacterium animalis/genética , Bifidobacterium animalis/fisiología , Suplementos Dietéticos , Heces/microbiología , Femenino , Voluntarios Sanos , Humanos , Lacticaseibacillus rhamnosus/genética , Lacticaseibacillus rhamnosus/aislamiento & purificación , Masculino , Persona de Mediana Edad , Tetraciclina , Adulto JovenRESUMEN
BACKGROUND: Helicobacter pylori produces Hpn, a 60-amino acid, histidine-rich protein that avidly binds nickel and zinc ions, and NixA, a high-affinity nickel transporter in the cytoplasmic membrane. We tested the hypothesis that Hpn and NixA govern susceptibility to metal ions in H. pylori. MATERIALS AND METHODS: Hpn-negative mutants of four H. pylori strains were constructed by standard allelic exchange techniques to yield isogenic Hpn+/Hpn-deficient pairs. A metal concentration that inhibited growth by 50% (IC50) was calculated for Ni2+, Zn2+, Cu2+, and Co2+ by comparing OD600 of cultures in metal-supplemented and control media. RESULTS: Among all four pairs of isogenic strains, the tolerance for Ni2+ was reduced significantly (p <.001) in the Hpn mutants; the mean IC50 value for wild-type strains was 1.9 mM; for the mutant, it was 0.8 mM. In contrast, growth inhibition by Zn2+ was identical within the fours pairs, as was Cu2+ and Co2+ tolerance in one pair tested. We also found that deletion of the hpn gene increases susceptibility to therapeutic forms of bismuth by testing a mutant and wild-type pair with ranitidine bismuth citrate, bismuth citrate, and four antibiotics. Minimal inhibitory concentrations of ranitidine bismuth citrate dropped from 9.2 to 2.3 microg/ml, and those of bismuth citrate dropped from 7.4 to 3.2 microg/ml (p <.05 for both comparisons), while susceptibility to the antibiotics was unaffected. Disruption of the nixA gene encoding the specific Ni2+ transport protein of H. pylori did not change susceptibility to bismuth. CONCLUSION: We concluded that bacteria lacking Hpn, cultured in vitro, are more susceptible than is the wild type to bismuth and Ni2+.
Asunto(s)
Proteínas Bacterianas , Bismuto/farmacología , Proteínas Portadoras/metabolismo , Proteínas de Transporte de Catión , Helicobacter pylori/efectos de los fármacos , Proteínas de la Membrana/metabolismo , Metales Pesados/farmacología , Proteínas/metabolismo , Proteínas Portadoras/genética , Farmacorresistencia Microbiana , Infecciones por Helicobacter/microbiología , Helicobacter pylori/genética , Helicobacter pylori/crecimiento & desarrollo , Humanos , Proteínas de la Membrana/genética , Pruebas de Sensibilidad Microbiana , Compuestos Organometálicos/farmacología , Proteínas/genética , Ranitidina/análogos & derivados , Ranitidina/farmacologíaRESUMEN
Helicobacter pylori is killed in vitro by polyoxyethylene acyl esters and ethers similar to simethicone emulsifiers in therapeutic antifoams. The MBC of these compounds for Helicobacter pylori was less than 20 micrograms/ml, while other gram-negative bacteria were unaffected by much higher concentrations of up to 50 mg/ml.