Quantitative procedure for enumeration of bifidobacteria

Abstract

A membrane filter technique has been developed for the enumeration of bifidobacteria in natural aquatic environments. The technique is quantitative, selective, and differential. The medium (YN-6) contains: yeast extract, 2.0 g; agar, 1.5 g; polypeptone peptone, 1.0 g; vitamin-free Casamino Acids, 0.8 g; sodium chloride, 0.32 g; and L-cysteine hydrochloride, 0.003 g; in 100 ml of deionized water. The medium is adjusted to pH 7.0 before autoclaving. Nalidixic acid (80 micrograms/ml), neomycin sulfate (2.5 micrograms/ml), and bromcresol green (300 micrograms/ml) are included as selective and differential agents. After incubation for 48 h at 37 degrees C in an anaerobic environment, Gram-stained smears from green, glistening, smooth entire colonies are examined microscopically for typical bifidobacterial morphology. No significant difference in recoveries was observed when YN-6 was compared with reinforced clostridial agar, using bifidobacteria freshly isolated from feces and raw sewage. Using this technique with aquatic and fecal samples, less than 9% false-positive and 8% false-negative isolates were observed. These results indicated that the medium was able to satisfactorily recover organisms from a variety of situations.

The persistence of bifidobacteria populations in a river measured by molecular and culture techniques

“…Bifidobacterium has been suggested as a microbial indicator in water, as this genus fulfils most of the requirements for being a faecal indicator (Resnick and Levin 1981b;Mara and Oragui 1983;Bahaka et al 1993;Beerens 1998;Sinton et al 1998;Lynch et al 2002;Nebra et al 2003). Moreover, some species are associated with human faeces, whereas others have an exclusively animal origin.…”

Sorbitol-fermenting bifidobacteria as indicators of diffuse human faecal pollution in estuarine watersheds

“…Although colonial morphology facilitates distinguishing bifids from other bacteria that grow on media designed for detection of bifidobacteria, i.e. bacteroides, clostridia, eubacteria, lactococci, lactobacilli, enterococci, streptococci ( Resnick & Levin 1981a; Shah 1989; Hartemink et al . 1996 ; Silvi et al .…”

Selective medium for isolation and enumeration of Bifidobacterium spp

“…were made in parallel on RCA. This rich, nonselective medium served as a reference, as reported previously (11).…”

“…Recently, two selective and differential media have been described for the isolation of Bifidobacterium spp. : (11) and (9). However, as has been reported previously (9), YN-6 medium is inhibitory to some species of Bifidobacterium and lacks the desired selectivity to assess a real estimation of the number of viable cells of Bifidobacterium spp.…”