Overview

Lactose Broth does two jobs in a food / water / dairy QC lab:
  • Pre-enrichment — gently revives stressed Salmonella from a sample before you transfer to a selective broth.
  • Presumptive coliform test — coliforms ferment the lactose and produce gas, captured in a Durham tube.

The GMExpression formulation matches the BD Difco 211725 / Oxoid CM0137 / HiMedia M015 reference (beef extract 3 g/L + peptone 5 g/L + lactose 5 g/L = 13 g/L total) and is validated for FDA BAM Chapter 5, USDA-FSIS MLG 4, and APHA Standard Methods 9221B. Under ISO 6579-1:2017 the universal pre-enrichment medium is now Buffered Peptone Water; Lactose Broth is retained for certain dairy-product matrices and for labs running legacy ISO 6579:2002 or FDA-aligned workflows.

Why a non-selective broth as the first step? Cells stressed by freezing, heat, drying, or acidity in food and dairy matrices are damaged at the membrane and ribosome level. Selective broths (Rappaport-Vassiliadis Soya, MKTTn, selenite-cystine) kill those cells before they can recover. The 18–24 h Lactose Broth incubation lets the cells repair first; the selective step then picks them up alive.

How the coliform / Durham gas test works. Lactose is the only fermentable sugar in the broth. E. coli, Klebsiella, Enterobacter, and Citrobacter ferment it to mixed acids + gas within 24–48 h at 35 °C. Trapped gas in an inverted Durham tube is the presumptive positive. Confirm by streak on EMB, MacConkey, or VRBL agar — or by sub-culture into EC broth at 44.5 °C for thermotolerant / faecal coliform confirmation.

We also have

Rappaport-Vassiliadis Soya Broth (selective Salmonella enrichment) · Selenite-Cystine Broth · Brilliant Green Bile 2% Broth (confirmatory coliform) · EC Broth (faecal coliform & E. coli) · Lauryl Sulfate Tryptose Broth (LST) · XLD Agar (Salmonella confirmation) · MacConkey Agar

Package Contents

Standard pack:

  • Mixture — pre-weighed dehydrated Lactose Broth base (beef extract, peptone, lactose) at 13 g/L; supplied in N₂-flushed amber bottles, sized for 10 L of final volume.

Optional supplies:

  • Optional inverted Durham tubes (6 × 50 mm borosilicate, 100 pieces) — for gas-collection coliform / lactose-fermentation assay.

Alternative pack on request:

  • Pre-poured 10 mL tubes (× 100) — sterile, ready-to-use; includes inverted Durham tube where ordered.

Customisation options on request: double-strength formulation (26 g/L) for high-sample-volume MPN dilution; bromocresol-purple-supplemented variant for direct acid-production visualisation; agar-supplemented variant for plate enumeration; cycloheximide 50 mg/L addition for fungal suppression in long-incubation environmental matrices.

Composition — per 1 L of prepared medium

Mixture — Dehydrated base (per 1 L; BD Difco 211725 / Oxoid CM0137 / HiMedia M015 reference)

ComponentConcentrationFunction
Beef extract (defatted bovine muscle infusion)3.0 gAmino acids, B-vitamins, mineral micronutrients; supports repair of sub-lethally injured cells
Peptone (pancreatic digest of animal protein)5.0 gFree amino acids, dipeptides; primary nitrogen source
Lactose monohydrate (β-D-galactopyranosyl-(1→4)-D-glucose · H2O; CAS 64044-51-5; anhydrous form CAS 63-42-3)5.0 g monohydrate (= 13.9 mM; equivalent to 4.75 g/L lactose anhydrous)Sole fermentable carbohydrate; differential substrate for coliforms (lactose-fermenters) vs Salmonella (lactose-non-fermenters). EP/USP-grade lactose monohydrate is the standard pharmacopoeial reference form.

Pre-autoclaving pH: 6.9 ± 0.2 at 25 °C (no adjustment normally required with commercial-grade powder).

Dehydrated total solids: 13.0 g/L. For double-strength formulation (used when an equal volume of liquid sample is added in MPN coliform assays), prepare at 26 g/L.

Indicative chemistry

ParameterValueNotes
Final pH after autoclaving6.9 ± 0.2 at 25 °CWithin BD Difco 211725 / Oxoid CM0137 / HiMedia M015 published specification of 6.9 ± 0.2
ColourLight straw-yellow, clearNo precipitate; slight Maillard browning normal
Lactose stabilityStable at 121 °C × 15 minAvoid > 20 min — lactose caramelises with peptone (browning)
Osmotic strength~ 80 mOsmHypotonic — favours injured-cell recovery and repair
SelectivityNone (non-selective)By design — for resuscitation step only

→Culturomics and Media Navigator

Not sure which pre-enrichment + selective enrichment + confirmatory agar fits your matrix? The GMExpression Culturomics and Media Navigator works both ways:

  • Start from a target pathogen — Salmonella, E. coli O157:H7, Cronobacter sakazakii, Listeria monocytogenes — and see the full ISO/FDA-aligned medium panel.
  • Start from a matrix type — raw chicken, dairy powder, leafy greens, bottled water, dried egg, chocolate — and see the validated medium sequence.

The Navigator cross-references Lactose Broth with the rest of the food / water QC catalogue: RVS, MKTTn, Selenite-Cystine, Brilliant Green Bile, EC, LST, XLD, EMB, MacConkey, and VRBL.

Use and Applications

  • FDA Bacteriological Analytical Manual (BAM) Chapter 5 — Salmonella. Pre-enrichment of dry foods (chocolate, milk powder, nut butter, dried egg, dried herbs); 25 g sample in 225 mL Lactose Broth (1:10 ratio), 35 °C × 24 h before transfer to Rappaport-Vassiliadis Soya (RVS) and Tetrathionate (TT) selective enrichment broths. Lactose Broth is the FDA BAM Chapter 5 reference pre-enrichment medium for the matrix categories listed.
  • ISO 6579-1:2017 — legacy / matrix-specific pre-enrichment. The current ISO 6579-1:2017 main procedure specifies Buffered Peptone Water as the universal non-selective pre-enrichment medium; Lactose Broth was the pre-enrichment medium under earlier ISO 6579:2002 editions and is retained as a validated alternative for certain dairy-product workflows and laboratories operating under legacy method validations. For new ISO 6579-1:2017 method development, see our companion Buffered Peptone Water product.
  • APHA Standard Methods 9221B — Standard Total Coliform Multiple-Tube Fermentation. Single-strength Lactose Broth (10 mL) inoculated with 1 mL water sample, or double-strength (10 mL of 2× LB) inoculated with 10 mL sample; incubated 35 °C × 24–48 h; positive tubes show gas in Durham tube within 48 h.
  • USDA-FSIS MLG 4.10 / 4.11 — Salmonella in poultry and meat. Lactose Broth as the pre-enrichment step before selective transfer.
  • Dairy product testing (IDF / ISO 6611 framework). Lactose Broth pre-enrichment for Salmonella in skim milk powder, infant formula, whey powder; lactose in the medium is also the fermentable substrate for incidental yoghurt-culture lactobacilli (which can mask Salmonella signal — use selective enrichment promptly).
  • Bottled water and water-supply quality monitoring. Pre-enrichment of stressed coliforms from chlorinated, UV-treated, or filtered water before MPN enumeration.
  • Cosmetics, pharmaceutical, and personal-care product safety testing (USP <62> / EP 2.6.13). Pre-enrichment for Salmonella testing of non-sterile manufactured products.
  • Environmental swab testing (food-processing surfaces, retail-deli surfaces, slaughter-line environment). Pre-enrichment from sponge / swab samples in 100–200 mL Lactose Broth.
  • Presumptive coliform / faecal-coliform identification. Subculture positive Lactose Broth tubes into EC broth (44.5 °C) for thermotolerant / faecal coliform confirmation, or into Brilliant Green Bile Broth for total coliform confirmation.

Compatible Microorganisms

Resuscitation of sub-lethally stressed pathogens (intended primary use)

  • Salmonella enterica subsp. enterica — all serovars including ser. Typhimurium (ATCC 14028), Enteritidis (ATCC 13076), Choleraesuis, Newport, Heidelberg, Infantis, Montevideo
  • Salmonella enterica subsp. arizonae, diarizonae, houtenae, indica
  • Salmonella bongori
  • Escherichia coli (ATCC 25922) — including pathogenic STEC, EHEC, ETEC, EIEC, EAEC strains (with EC broth follow-up at 44.5 °C)
  • Cronobacter sakazakii (ATCC 29544) — historical "Enterobacter sakazakii"; ISO/TS 22964 pre-enrichment in modified LST

Coliform group (presumptive lactose-fermentation positive)

  • Escherichia coli (ATCC 25922) — gas-positive in 24 h at 35 °C and 44.5 °C
  • Klebsiella pneumoniae (ATCC 13883), K. oxytoca (ATCC 49131), K. variicola
  • Enterobacter cloacae (ATCC 13047), E. aerogenes (now Klebsiella aerogenes; Tindall & Sutton 2017 IJSEM 67:502), E. asburiae, E. hormaechei
  • Citrobacter freundii (ATCC 8090), C. koseri (ATCC 27028)
  • Serratia marcescens (lactose-positive variable; gas may be weak)
  • Hafnia alvei (slow lactose fermenter)

Not selective against (background flora)

  • Pseudomonas spp., Aeromonas spp. — grow but do not ferment lactose; no gas in Durham tube
  • Proteus, Providencia, Morganella — lactose non-fermenters; may cause swarming; transfer to selective medium promptly
  • Streptococcus, Enterococcus, lactic acid bacteria — grow but the medium is not designed for them
  • Yeasts and moulds — supplement with cycloheximide 50 mg/L for fungal suppression if matrix contamination is heavy

Preparation

1Weigh. Use 13.0 g of Mixture A for 1 L of single-strength broth, or 26.0 g for 1 L of double-strength broth. A pre-weighed sachet for 5 L (= 65 g) is supplied; pour into the bottle.
2Suspend & dissolve. Add the powder to ~950 mL of Milli-Q water (Type 1 ultrapure, > 18 MΩ·cm at 25 °C) at room temperature. Swirl gently for 2–3 min until fully suspended. Heat briefly on a stirrer-hot-plate to 70–80 °C with mild stirring until completely dissolved (5 min).
3pH check. The pH should be 6.9 ± 0.2 at 25 °C. No adjustment is normally required. If pH is < 6.7, adjust upwards with 0.1 M NaOH; if > 7.1, adjust downwards with 0.1 M HCl.
4Bring to final volume. Make up to 1000 mL with Milli-Q water.
5Dispense. For tubed format: dispense 10 mL into 16 × 150 mm tubes containing an inverted 6 × 50 mm Durham tube. For bottled bulk format (Salmonella pre-enrichment): dispense 225 mL into 500 mL autoclavable bottles. Cap one-quarter turn loose for autoclaving.
6Autoclave. 121 °C × 15 min (103 kPa). Do not exceed 15 min. Excess autoclaving caramelises lactose with the peptone (Maillard reaction), producing a dark amber discoloration and depressing the recoverable lactose pool. Slow exhaust to avoid bumping.
7Cool. Allow to cool to room temperature. Verify Durham tubes are inverted and completely filled with broth (no air bubble at the closed end — if a bubble is present, re-autoclave to displace).
8Sterility QC. Incubate one tube per lot at 35 °C × 24 h before release. Verify no gas formation (lactose pool intact) and no turbidity.
9Inoculation — Salmonella pre-enrichment (FDA BAM Chapter 5). Add 25 g of solid food sample (or 25 mL of liquid sample) to 225 mL pre-warmed (35 °C) Lactose Broth in a stomacher bag or 500 mL bottle. Homogenise for 2 min in a Stomacher 400. Incubate at 35 °C ± 1 °C for 24 ± 2 h. (Note: for ISO 6579-1:2017 workflows, Buffered Peptone Water at 18 ± 2 h, 37 °C ± 1 °C is the currently specified pre-enrichment — see our BPW product page.)
10Inoculation — coliform MPN. Inoculate 5 tubes of 10 mL single-strength LB with 1 mL water sample each (5 × 1 mL series); or 5 tubes of 10 mL double-strength LB with 10 mL sample each (5 × 10 mL series). Incubate at 35 °C ± 0.5 °C and observe gas at 24 h and 48 h. Refer to APHA Standard Methods Table 9221:I for MPN calculations.
11Sub-culture from positive tubes. For Salmonella: 0.1 mL of pre-enrichment into 10 mL RVS broth (41.5 °C, 24 h) and 1.0 mL into 10 mL MKTTn broth (37 °C, 24 h); plate onto XLD and BPLS / BGA. For coliforms: loopful into Brilliant Green Bile 2% Broth (35 °C, 48 h) and EC Broth (44.5 °C, 24 h) for confirmation.

Critical control points

  • Autoclaving time. 15 min is the upper bound. 18–20 min is acceptable for large-volume autoclaves with slow heat-up, but only if validated by pH and visual-colour QC. Above 20 min, Maillard browning and lactose caramelisation will progressively shift the pH downward (toward ~6.5) and reduce the recoverable lactose concentration; sensitive coliform strains may show false-negative gas results.
  • Durham tube air-bubble check. After autoclaving, every Durham tube must be inspected for residual air bubble. A retained air bubble at the closed end will be misread as a false-positive gas signal at 24 h.
  • Pre-warming. The pre-enrichment medium should be pre-warmed to 35–37 °C before sample addition; cold-shock of injured cells reduces recovery by 0.5–1.0 log. Applies to both FDA BAM ch. 5 (35 °C target) and legacy ISO 6579 workflows (37 °C target).
  • Sample-to-medium ratio. Always use 1:10 (sample : Lactose Broth). Higher dilution reduces inhibitor / interferent dilution; lower dilution overloads the recoverable nutrients.

Cautions

Lactose caramelisation on overlong autoclaving. The combination of reducing sugar (lactose) and free amino groups (peptone) undergoes Maillard browning at 121 °C. Mild straw-amber colour is normal; dark amber → brown indicates over-processing. Standardise to 15 min at 121 °C and verify post-autoclave pH ≥ 6.7 as the release criterion.
Non-selective medium — secondary contamination risk. Lactose Broth contains no inhibitors and will support the growth of nearly all environmental bacteria including spore-forming Bacillus, Pseudomonas, swarming Proteus, and yeasts. For environmental samples with heavy background flora, transfer to selective enrichment within the validated time window (16–20 h ISO, 24 h FDA). Do not extend Lactose Broth incubation beyond 48 h.
Lot-to-lot lactose source variability. Commercial lactose (β-lactose vs α-lactose anhydrate vs lactose monohydrate) differs in solubility kinetics and dissolved-sugar concentration. GMExpression sources EP-grade lactose monohydrate. Verify hydrate form on the CofA before customer use; the 5.0 g/L specification is for the monohydrate form (= 4.75 g/L anhydrous-equivalent).
False-positive gas from Aeromonas at low temperature. Some environmental Aeromonas spp. produce gas from lactose at 35 °C but not at 44.5 °C. Always confirm positive tubes by sub-culture to EC broth (44.5 °C) for thermotolerant / faecal coliform certification, and to MacConkey or EMB for morphological identification.
Bovine-derived components and biosecurity. Beef extract is bovine-derived. Although fully denatured during processing, EU, Japan, and Canada (CFIA) customs may apply TSE/BSE prion-risk checks. GMExpression uses the established Australian DAFF EX188M zoosanitary certificate pipeline; documentation packs available via support@gmexpression.com.
Storage of poured tubes. Pre-poured Lactose Broth tubes have a 30-day refrigerated shelf life. Refrigeration is required because lactose / peptone broth at room temperature supports significant background growth from airborne contaminants within 5–7 days even when capped. Use first-in-first-out stock rotation.

Storage and Expiry · Safety

  • Dehydrated powder (Mixture A): store sealed at 15–30 °C in original packaging away from direct sunlight. Shelf life 36 months from manufacture.
  • Prepared broth, screw-cap tubes / bottles, 2–8 °C: 30 days from preparation.
  • Prepared broth, 15–25 °C (ambient): 5 days from preparation; recommended for same-day or next-day use only.
  • Double-strength broth: identical storage as single-strength.
  • Inverted Durham tubes (borosilicate glass): indefinite shelf life; sterilised in situ with the broth.

Safety notes. Lactose Broth contains no Schedule-listed pathogens or controlled reagents in the dehydrated form. The medium is bovine-derived (beef extract). Handle prepared broth and inoculated cultures at BSL-2 when working with Salmonella serovars or pathogenic E. coli. Autoclave all inoculated medium before discard (121 °C × 30 min recommended for tubed cultures). SDS available on request.

References

  1. ISO 6579-1:2017. Microbiology of the food chain — Horizontal method for the detection, enumeration and serotyping of Salmonella — Part 1: Detection of Salmonella spp. Geneva: International Organization for Standardization.
  2. FDA Bacteriological Analytical Manual (current edition). Chapter 5: Salmonella. U.S. Food and Drug Administration, College Park, MD.
  3. APHA, AWWA, WEF. (current edition). Standard Methods for the Examination of Water and Wastewater, 24th ed., Method 9221 — Multiple-Tube Fermentation Technique for Members of the Coliform Group. Washington, DC: American Public Health Association.
  4. USDA-FSIS. Microbiology Laboratory Guidebook (MLG) 4.10 / 4.11 — Isolation and Identification of Salmonella from Meat, Poultry, Pasteurized Egg, and Catfish Products. Athens, GA: U.S. Department of Agriculture Food Safety and Inspection Service.
  5. USP <62> — Microbiological Examination of Nonsterile Products: Tests for Specified Microorganisms. United States Pharmacopeial Convention.
  6. European Pharmacopoeia 2.6.13 — Microbiological Examination of Non-Sterile Products: Test for Specified Microorganisms. EDQM, Council of Europe.
  7. BD Difco™ Lactose Broth product information sheet, BD 211725.
  8. Oxoid Manual, CM0137 Lactose Broth.
  9. HiMedia M015 Lactose Broth technical sheet.
  10. Andrews WH, Hammack TS. (current edition). Salmonella. In: Bacteriological Analytical Manual, FDA, College Park MD (BAM Chapter 5).
  11. D'Aoust JY, Sewell AM, Daley E, Greco P. (1992). Pre-enrichment conditions for the effective recovery of Salmonella in foods and feed ingredients. Journal of Food Protection 55: 88–92.

Frequently Asked Questions

Q1. Why pre-enrich Salmonella in a non-selective medium first, instead of going straight to selective enrichment?
Salmonella cells recovered from food, water, and dairy products are frequently sub-lethally stressed: freeze-thaw injury (frozen foods), heat injury (pasteurised dairy), acid injury (mayonnaise, fermented foods), desiccation injury (spice powders, infant formula), and osmotic injury (cured meats) all damage membrane integrity, ribosomes, and proton-motive force. Selective enrichment broths (RVS, MKTTn, selenite-cystine) contain inhibitors (high osmotic pressure, novobiocin, brilliant green, selenium) that target the Salmonella cell envelope. An injured cell exposed directly to selective enrichment will die before it can recover. The 16–20 h pre-enrichment in non-selective Lactose Broth allows the cell to repair membrane and ribosomal damage and re-enter exponential growth before being challenged by the selective inhibitors.
Q2. Why lactose specifically, when Salmonella doesn't ferment lactose?
The lactose in Lactose Broth is not there for Salmonella — it is there for the historical secondary use of the medium as a coliform-detection broth (lactose-fermenting Enterobacteriaceae produce acid and gas, captured in a Durham tube). For the Salmonella pre-enrichment use, the lactose is largely ignored — Salmonella metabolises peptone amino acids instead. The choice of Lactose Broth as the standard pre-enrichment broth is historical: it was the universally available non-selective broth in the mid-20th century when the FDA BAM and APHA methods were codified. Modern alternatives such as Buffered Peptone Water (the universal pre-enrichment specified in ISO 6579-1:2017) and Universal Pre-enrichment Broth (UPB) give equivalent or slightly better recovery; Lactose Broth remains in the FDA BAM Chapter 5 catalogue for compatibility with legacy method validations and specific dairy / chocolate / dried-egg matrices.
Q3. Can I use Buffered Peptone Water (BPW) instead of Lactose Broth?
Yes — for new method development aligned with ISO 6579-1:2017, Buffered Peptone Water is the specified universal non-selective pre-enrichment medium. Lactose Broth is retained as a validated alternative under FDA BAM Chapter 5 and earlier ISO 6579:2002 editions, particularly for dairy products with a strong legacy validation base (skim milk powder, infant formula, cheese, milk chocolate). BPW is non-selective, lactose-free, and pH-buffered at 7.0 with phosphate; it gives marginally better recovery for desiccation-stressed cells. For coliform / Durham-gas assays (APHA SM 9221, FDA BAM ch. 4), Lactose Broth remains the reference medium — BPW has no fermentable sugar. The Navigator's "medium-by-matrix" decision-tree will guide selection.
Q4. Why does my Lactose Broth turn dark brown after autoclaving?
Maillard browning between the reducing sugar (lactose) and the free amino groups of peptone. Mild straw-amber browning is normal. Dark amber to brown indicates one of: (a) autoclave time > 15 min, (b) autoclave temperature exceeding 121 °C (check spore-strip calibration), (c) lactose hydrate form different from spec (alpha-lactose anhydrate browns faster than monohydrate), or (d) iron contamination of water (use Milli-Q water, > 18 MΩ·cm at 25 °C). Verify the pH at 25 °C after autoclaving — if pH < 6.7, discard the lot.
Q5. Can I autoclave Lactose Broth in plastic bottles?
Yes for polypropylene (PP) bottles rated for 121 °C autoclave cycles. Do not use polystyrene (PS) or low-density polyethylene (LDPE) — they will deform. Glass borosilicate (Schott Duran, Pyrex) is the gold standard. Always leave caps one-quarter turn loose during autoclaving for pressure equilibration; tighten only after the load has cooled below 80 °C.
Q6. Why do I get false-positive gas from a non-coliform background?
Most commonly: Aeromonas hydrophila, Aeromonas caviae, and some Vibrio spp. can produce gas from lactose at 35 °C but not at 44.5 °C — they are not coliforms by the operational definition (faecal coliforms / E. coli). The standard confirmation is sub-culture from gas-positive tubes into EC Broth at 44.5 °C × 24 h; only gas-positive at 44.5 °C qualifies as a thermotolerant / faecal coliform / E. coli. Additionally, some non-fermenting environmental flora can introduce a retained air bubble in the Durham tube — always verify by repeated incubation and confirmatory streak to EMB or MacConkey.
Q7. How long should I incubate Lactose Broth pre-enrichment for Salmonella?
FDA BAM Chapter 5 specifies 24 ± 2 h at 35 °C ± 1 °C for Lactose Broth pre-enrichment of the matrices in scope. Earlier ISO 6579:2002 editions specified 16–20 h at 37 °C ± 1 °C for Lactose Broth; ISO 6579-1:2017 has since replaced Lactose Broth with Buffered Peptone Water as the universal non-selective pre-enrichment medium (18 ± 2 h at 37 °C ± 1 °C). Do not exceed 24 h — extended pre-enrichment allows non-pathogen background flora to overtake the Salmonella population and reduces the apparent positive rate. The full sample-to-result timeline is typically: pre-enrichment 18–24 h → selective enrichment 24 h → selective plating 24 h → confirmation 24 h → biochemical / serotyping 24 h = 5–6 days total for confirmed Salmonella identification.
Q8. Can I use Lactose Broth for E. coli O157:H7 detection?
Lactose Broth is not the preferred medium for E. coli O157:H7. The standard pre-enrichment for STEC / O157:H7 per FDA BAM Chapter 4A and USDA-FSIS MLG 5B is modified TSB with novobiocin (mTSB+N) or Buffered Peptone Water with vancomycin / cefsulodin / cefixime (mBPW+v). Lactose Broth lacks the selective inhibitors needed for O157:H7 differential enrichment, and O157:H7 is lactose-fermentation variable (some isolates ferment slowly, some not at all — making the Durham gas test unreliable for this serotype).