Food microbiological analysis is highly variable, given the living status of the analyte and the biological nature of the reagents used. In addition the analytical results are closely related to the method's principle, in particular with conventional Pasteur microbiology and for enumeration methods. Food-borne pathogens represent a direct health hazard for consumer's health. For these reasons, there is a clear need to define common reference methods, properly validated, as well as to get assurance on the quality of commercial test kits which can be used in alternative to these reference methods.
Standardization can be considered to be the preferred mean, since based on consensus, to harmonize reference methods at international level. Standardization structures at international level in ISO and at European level in CEN in food microbiology are at first presented. Some methodological aspects are introduced, as well as the different types of standards developed: reference methods, general aspects. The current programme of work is overviewed. A special hint is given to the position of novel technologies, as well as to the degree of validation of these Standard reference methods.
Reference methods being most of the time based on conventional microbiology, they require a skilled staff and are labor-intensive, long to perform due to the time needed by the bacteria to grow in/on culture media. Thus a large range of alternative methods, based on recent technologies such as ELISA or PCR, have been developed to get results within a shorter interval of time or to reduce work-load through automation. Manufacturers have commercialized a large range of such test kits, but their users need guarantees on their validity, thus the development of validation schemes operated by third-party organizations. To have a common reference technical protocol to conduct these validations, CEN has developed a standard for the validation of alternative methods in food microbiology, adopted by ISO, EN ISO 16140. This standard is presented, as well as its current revision by ISO. The validation/certification schemes are then shortly introduced.
In a final part, we consider the position of Standard reference methods and of validated alternative methods in the context of laboratory accreditation, based on ISO 17025 Standard, and in the European regulatory frame of Regulation 2073/2005 on microbiological criteria in foodstuffs.

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To establish an efficient method for detection and isolation of Salmonella in feed and food manufacturing plant, we evaluated the delayed secondary enrichment (DSE) method, which has already been acknowledged in the field of poultry hygiene. In order to evaluate the detection rate by the increase of pre-enrichment culture and selective enrichment broth, the oilmeal samples (n=182) were examined using different volume (10 ml and 90 ml) of the Hajna-tetrathionate broth (HTT). As a result, the sample with different result was only two (1.1%). The detection sensitivities of these 2 methods were same (p<0.05). We examined oilseed (n=34) and swab samples (n=92) of environment in the oilmeal manufacturing plant. The DSE method alone showed the highest rate of Salmonella detection at 14.3% (18/126), followed by the HTT-DHL method and HTT-CHROMagar Salmonella method, both at 12.7% (16/126). Moreover, O antigen group of isolates detected by the DSE method was different from those of HTT-DHL method in 7 samples. It was found that the detection sensitivity of the DSE method was high. Our findings suggest that O antigen group of isolates detected by the DSE method may differ from those of conventional enrichment methods. We conclude that DSE method is a simple and efficient method for epidemiological investigations of Salmonella in environments, such as food manufacturing plants.

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From April 2002 to March 2008, we collected a total of 210 Salmonella enterica serovar Infantis (Salmonella Infantis) isolates from domestic chicken meat sold by retail stores in Yokohama, Japan. We examined the isolates for antimicrobial susceptibilities to 13 antibiotics (ampicillin, cephalothin, cefotaxime, streptomycin, kanamycin, gentamicin, tetracycline, chloramphenicol, sulfisoxazole, sulfamethoxazole-trimethoprim, nalidixic acid, ciprofloxacin, and fosfomycin). One hundred ninety-four isolates (92.4%) were resistant to at least one antibiotics. Among antibiotics tested, resistance rates against sulfisoxazole, streptomycin, and kanamycin were high.
None of the Salmonella Infantis isolates in this study were resistant to ciprofloxacin. Seven isolates were found to be resistant to ampicillin and cephalothin and 4 isolates were resistant to ampicillin, cephalothin and cefotaxime. Of the 11β-lactamase producing isolates, 5 were found to produce extended-spectrum β-lactamase (ESBL). As a result of the bla gene screening, bla_TEM-20, bla_TEM-52 and bla_CTX-M-2 were detected in 1, 3 and 1 ESBL-producing isolate. In the remaining 6 isolates, bla_CMY-2 was detected. Six of the 11 isolates were originated from chicken meat products that were shipped from the same corporation in a couple of years.
Salmonella Infantis producing TEM-20 type ESBL was detected for the first time in this study.

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