FDA updates the BAM method for Cronobacter (September 2026 edition): secondary enrichment in CSB broth for powdered infant formula, a procedure for environmental samples and a new qPCR probe
The US Food and Drug Administration (FDA) has published the September 2026 edition of Chapter 29 of the Bacteriological Analytical Manual (BAM) — the method for the detection and isolation of Cronobacter spp. in powdered infant formula (PIF) and environmental samples. The method page is current as of 16 September 2026; the previous edition of the chapter dated from November 2023.
- Enrichment: the number of enrichment culture aliquots subjected to centrifugation was reduced from three to one. As an equally acceptable approach for PIF, secondary enrichment with a corresponding DNA extraction procedure was added — 0.1 ml of the buffered peptone water (BPW) culture is transferred to 10 ml of CSB broth and incubated at 41.5 ± 1 °C for 24 ± 2 h. According to FDA, secondary enrichment in CSB helps isolate Cronobacter from samples with high levels of background flora, and qPCR after the two-step BPW–CSB enrichment yields results one day later than qPCR performed directly after BPW enrichment and centrifugation.
- Procedures for enrichment, qPCR screening and cultural confirmation of environmental samples were added (a swab in 9 ml BPW, a sponge in 90 ml BPW). The qPCR, which targets part of the macromolecular synthesis operon spanning the 3′ end of the rpsU gene and the 5′ end of the dnaG gene, gained a preferred double-quenched Cronobacter probe, an equally acceptable approach to preparing the internal amplification control and a higher annealing temperature.
- The required and optional chromogenic agars were updated — in the 2026 edition’s analysis flowchart these are DFI agar (36 ± 1 °C), CCI agar and R&F agar (41.5 ± 1 °C), with optional ESIA agar (44 ± 1 °C) and R&F agar at 36 ± 1 °C, each for 24 ± 2 h — as were the inclusivity and exclusivity tables: the method was tested on 50 Cronobacter strains representing seven species and on 40 strains of 34 non-Cronobacter species.
- The method combines real-time PCR screening with a cultural component that can be used on its own, without qPCR; a positive qPCR result is always confirmed by the cultural method. Pure cultures are confirmed by qPCR and VITEK 2 or by qPCR alone. The November 2023 edition had, among other things, removed the use of the SmartCycler for qPCR and the option of using RAPID ID 32E for cultural confirmation, and added optional CCI and ESIA agars.