Modified Semisolid Rappaport Vassiliadis Medium (MSRV) ISO

It is a modification of Rappaport Vassiliadis enrichment broth for detecting motile Salmonella spp. in feces, food products and environmental samples. In this medium the main detection is based on the motility and ability of Salmonella to migrate through the selective medium ahead of competing motile microorganisms, therefore producing opaque halos of growth.

The mobility of other microorganisms is inhibited by selective agents (such as magnesium chloride, malachite green oxalate and novobiocin) as well as by the temperature of incubation at 42 °C.

Tryptose and acid casein peptone provide nitrogen, vitamins, minerals and amino acids essential for growth. Sodium chloride supplies essential electrolytes for transport and osmotic balance. Magnesium chloride and malachite green oxalate are inhibitory to organisms other than Salmonella spp. Novobiocin is a selective agent that inhibits Gram-positive bacteria and avoids the development of Proteus.

This medium is not suitable for the detection of non-motile strains of Salmonella, whose presence is very low (1%).

It is recommended to conduct serological and biochemical tests for Salmonella species confirmation.

Preparation

Suspend 31.6 grams of the medium in one liter of distilled water. Mix well and dissolve by heating with frequent agitation. Boil for one minute until complete dissolution. AVOID OVERHEATING. DO NOT AUTOCLAVE. Dispense into Petri plates.

According to ISO 6579

For detection of Salmonella spp. in food, animal feed, animal faeces, and environmental samples according to ISO 6579:

1. Pre-enrichment in non-selective liquid medium: inoculate the Buffered Peptone Water (LC-1402) with the sample or dilutions, and incubate at 34-38 °C for 18 ± 2 h.
2. Enrichment in/on selective media: inoculate, with the culture obtained in the pre-enrichment stage, the Rappaport Soy Broth (Vassiliadis) (LC-1174) or the Modified Semisolid Rappaport Vassiliadis Medium (MSRV), and the MKTTN Broth (LC-1173). The Rappaport Soy Broth and the Modified Semisolid Rappaport medium are incubated at 41.5 °C for 24 ± 3 h, and the MKTTN Broth at 34-38 °C for 24 ± 3 h.
3. Plating out on selective solid media: from the selective enriched cultures, inoculate two selective isolation agars: XLD agar (LC-1274) and any other selective medium complementary to XLD agar (Salmonella Chromogenic Agar (LC-1122), Brilliant Green Agar (LC-1143), Bismuth Sulfite Agar (LC-1011), DCLS Agar (LC-1045), Desoxycholate Citrate Agar (LC-1067), Hektoen Enteric Agar (LC-1030), Salmonella Shigella Agar (LC-1064) and XLT4 Agar (LC-1159)). Incubate the XLD plates inverted at 34-38 °C for 24 ± 3 h. Incubate the second selective medium in accordance with the manufacturer's instructions.
4. Confirmation: subculture colonies of presumptive Salmonella and confirm their identity by biochemical and serological tests.

Note: according to Annex D of ISO 6579-1:2017, for the detection of enterica subspecies enterica serovars Typhi and Paratyphi, Selenite Cystine Broth (LC-1220) should be added as a selective enrichment medium and Bismuth Sulfite Agar (Wilson Blair) should be selected as a second selective medium (LC-1011).

According to ISO 19250

For detection of Salmonella spp. in water samples according to ISO 19250:

1. Pre-enrichment in non-selective medium: inoculate the Buffered Peptone Water (LC-1402) with the sample or dilutions, and incubate at 34-38 °C for 18 ± 2 h.
2. Enrichment in selective media: inoculate, with the culture obtained in the pre-enrichment stage, the Rappaport Soy Broth (Vassiliadis) (LC-1174) and the MKTTN Broth (LC-1173). The Rappaport Soy Broth is incubated at 41.5 ± 1 °C and the MKTTN Broth at 34-38 °C, both of them for 24 ± 3 hours.
3. Plating out on selective solid media: from the selective enriched cultures, inoculate two selective isolation agars: XLD agar (LC-1274) and any other selective medium complementary to XLD agar (for instance, Brilliant Green Agar (LC-1143) or Bismuth Sulfite Agar (LC-1011)). Incubate the XLD plates inverted at 34-38 °C for 24 ± 3 hours. Incubate the second selective medium in accordance with the manufacturer's instructions.
4. Confirmation: subculture colonies of presumptive Salmonella and confirm their identity by biochemical and serological tests.

• De SMEDT et al.; Rapid Salmonella Detection in Foods by Motility Enrichment on a Modified Semi-Solid Rappaport-Vassiliadis Medium. J. Food Protect. Vol. 49, 7; 510-514 (1986).
• De SMEDT, a. BOLDERDIJK, R.F.; Dynamics of Salmonella Isolation with Modified Semi-Solid Rappaport-Vassiliadis Medium. J. Food Protect. Vol. 50, 8; 658-661 (1987).
• ISO 6579 Microbiology of food and animal feeding stuffs - Horizontal method for the detection of Salmonella spp. Amendment 1: Broader range of incubation temperatures, amendment to the status of Annex D, and correction of the composition of MSRV and SC.