Salmonella Serogroup Detection in Poultry Meat Samples by Examining Multiple Colonies from Selective Plates of Two Standard Culture Methods

Abstract

This study aims to determine the serogroup profiles of randomly collected 46 chicken meat and 15 turkey meat samples, following the U.S. Food and Drug Administration's (FDA) Bacteriological Analytical Manual Chapter 5: Salmonella and International Organization for Standardization (ISO) Method 6579 culture methods. The total number of poultry meat samples with more than one serogroup isolated by the FDA and ISO culture methods were 10 (37.0%) and 21 (77.8%) of 27, respectively. Presence of multiple serogroups per sample was more frequently observed in chicken meat samples than in turkey meat samples. The profile of Salmonella serogroup isolates of chicken meat samples in descending order were serogroups D and E4 (15.8%), B and C2 (8.8%), C1 (5.3%), G (3.5%), and E1 and F (1.7%). The serogroup distribution of turkey meat sample isolates were serogroups B (27.2%), E4 (18.2%), and C2 (9.1%). On the basis of our findings that a selective plate in Salmonella culture method can harbor more than one serogroup, and that the FDA and ISO methods could detect different serogroups from chicken and turkey meats, we suggest screening multiple suspect colonies from each plate, if possible, and considering the collective and comparative use of the FDA and ISO culture methods and/or including several selective and differential media to ensure the detection of Salmonella and the possible detection of multiple serogroups from samples.

Introduction

F oodborne salmonellosis remains as a major public health problem worldwide (Poppe, 1999; Bell and Kyriakides, 2002; DEFRA, 2002). Salmonella enterica serovars, as the etiological agents of foodborne salmonellosis, are carried by poultry and contaminate their meat, which is recognized as one of the major vehicles of human infections (Poppe, 1999). To reduce the incidence of Salmonella infections in humans due to consumption of contaminated poultry meat, the first step is to detect these pathogens by rapid and reliable methods (Malorny et al., 2007; O'Regan et al., 2008; Woods et al., 2008; Hadjinicolaou et al., 2009; Lee et al., 2009), together with internationally recognized culture methods for previously reported reasons (Schrank et al., 2001; Eyigor et al., 2002; Hoorfar et al., 2003; Lofstrom et al., 2004; Bohaychuk et al., 2007). Although time consuming, these culture methods are indispensable, as they are the only tools to determine complete Salmonella serovar profile of the sample for epidemiological investigations of foodborne salmonellosis and for further studies such as the detection of S. enterica serovars. In Salmonella culture methods, it is particularly important to search for as many Salmonella serovars/serogroups as possible to represent the actual serovar/serogroup distribution, because more than one serogroup/serovar type can be encountered in a food sample. Several studies (Eblen et al., 2006; Berrang et al., 2009; Dione et al., 2009; Miranda et al., 2009) indicated selecting a minimum of two typical colonies per selective plate in their Salmonella culture methods, but their purpose for doing so was to find the possible Salmonella isolate from the plate, consequently from the sample. Therefore, the aims of the aforementioned studies were not to search for more than one isolate in the plate or in the sample, or to determine whether a sample carried more than one Salmonella serovar/serogroup. The same issue is also unclear as indicated in some guidelines (ISO, 2002; FDA, 2007; USDA, 2008).

Recently, although not a food sample example, we were able detect three S. enterica serovars from a single eye inoculum of a diseased layer chicken (Carli et al., 1996), by picking five colonies from selective plates, as recommended by Waltman (1999). Also, a study by Bosilevac et al. (2009), reporting prevalence of Salmonella in commercial ground beef in the United States, informed as extra data that four samples had two different Salmonella serotypes. In this context, we suspected this instance could be encountered in food samples as well and so planned this study to determine if a poultry meat sample would contain more than one Salmonella serogroup. During our study, we came across a technical report by Fedorka-Cray et al. (2009) indicating more colony selection from one plate would increase the chance of determining more than one serogroup in a sample.

In the present study, presence of more than one Salmonella serogroup and the serogroup profiles in chicken meat and turkey meat samples was examined by following the U.S. Food and Drug Administration's (FDA) Bacteriological Analytical Manual Chapter 5: Salmonella (FDA, 2007) and International Organization for Standardization (ISO) Method 6579 (ISO, 2002) culture methods.

Materials and Methods

Samples

Forty-six chicken wing (comprised of 8 wings per package) and 15 turkey neck (comprised of 4 necks per package) retail meat samples belonging to six major brands, which were randomly collected from nine supermarkets, were transferred on ice to the laboratory. Samples in their original packages were individually repacked in polyethylene bags to prevent cross contamination during purchase and transfer. The analysis of all samples was initiated immediately after transfer to the laboratory as follows: All package contents of the retail meat purchased was placed into one sterile bag, and meat pieces were thoroughly massaged for 3 min from outside so that surfaces (skin and meat parts and any protruding bone parts) of all the samples contact completely to maintain a sufficiently mixed unit sample. Then, from this mixed sample, ∼60 g of meat was taken and minced into smaller pieces using sterile techniques. Two equal 25-g parts, each of which was placed into separate sterile stomacher bags, were used as samples for FDA and ISO preenrichments.

Culture

Standard preenrichments (lactose broth [Oxoid; CM0137] for FDA and buffered peptone water [Oxoid; CM1049] for ISO) and primary enrichments (tetrathionate broth [Oxoid; CM029] and Rappaport Vassiliadis R10 broth [Beckton Dickinson; 218581] for FDA, and Muller–Kauffmann tetrathionate–novobiocin broth [Oxoid; CM1048] and Rappaport Vassiliadis soya peptone broth [Oxoid; CM0866] for ISO) were applied as indicated in the FDA (2007) and ISO (2002) methods. Selective plating was performed from each of the primary enrichment broths of both methods onto xylose lysine deoxycholate agar (Oxoid; CM0469) and xylose lysine tergitol-4 agar (Beckton-Dickinson; 223420), and the plates were incubated at 35°C for 24 h. Five randomly selected suspect Salmonella colonies from each selective plate were picked and streaked onto individual MacConkey agar (MAC; Beckton-Dickinson; 212123) plates. After incubation at 37°C for 24 h, one colony from each MAC plate was subjected to both biochemical identification by API 20E (Biomerieux; 20100) and serological identification using Salmonella group-specific antisera (Beckton-Dickinson).

Serogrouping

One colony from each MAC plate was subjected to slide agglutination test with 10 μL of the following commercially available antisera for serogrouping: Salmonella O Antiserum Poly A (somatic groups A, B, D, E1 [E2, E3], E4, L); Salmonella O Antiserum Poly B (somatic groups C1, C2, F, G, H); Salmonella O Antiserum factor 4; Salmonella O Antiserum factor 5; Salmonella O Antiserum factor 9; Salmonella O Antiserum factor 12; Salmonella O Antiserum factor 14; Salmonella O Antiserum Group C1 (factors 6, 7); Salmonella O Antiserum Group C2 (factors 6, 8); Salmonella O Antiserum Group E1 (factors 3, 10); Salmonella O Antiserum Group E4 (factors 1, 3, 19) (Beckton Dickinson) were used according to antigenic formulae of the Salmonella serovars indicated in the White-Kauffmann-Le Minor Scheme (Grimont and Weill, 2007).

Results

When the FDA and ISO culture methods are considered together, two or more Salmonella isolates belonging to different serogroups were identified in 27 of 61 poultry meat samples (44.3%), with 2, 3, 4, and 5 serogroups (multiple) with the following rates: 66.7%, 18.5%, 11.1%, and 3.7% (Table 1).

Table 1.

Number of Serogroups in Chicken Meat and Turkey Meat Samples

		Number of samples with one or more different serogroups/total number of isolates (%)
Sample type (total number of isolates/total number of samples)	Method	One serogroup	Two different serogroups together	Three different serogroups together	Four different serogroups together	Five different serogroups together
Chicken meat (22/46)	FDA and ISO	0/22 (0.0)	14/22 (63.6)	4/22 (18.2)	3/22 (13.6)	1/22 (4.6)
	FDA	6/22 (27.3)	5/22 (22.7)	2/22 (9.1)	1/22 (4.6)	0/22 (0.0)
	ISO	6/22 (27.3)	10/22 (45.5)	4/22 (18.2)	1/22 (4.6)	1/22 (4.6)
Turkey meat (5/15)	FDA and ISO	0/5 (0.0)	4/5 (80.0)	1/5 (20.0)	0/5 (0.0)	0/5 (0.0)
	FDA	3/5 (60.0)	2/5 (40.0)	0/5 (0.0)	0/5 (0.0)	0/5 (0.0)
	ISO	0/5 (0.0)	4/5 (80.0)	1/5 (20.0)	0/5 (0.0)	0/5 (0.0)
Total (27/61)	FDA and ISO	0/27 (0.0)	18/27 (66.7)	5/27 (18.5)	3/27 (11.1)	1/27 (3.7)
	FDA	9/27 (33.3)	7/27 (25.9)	2/27 (7.4)	1/27 (3.7)	0/27 (0.0)
	ISO	6/27 (22.2)	14/27 (51.9)	5/27(18.5)	1/27 (3.7)	1/27 (3.7)

FDA, U.S. Food and Drug Administration; ISO, International Organization for Standardization.

When the FDA and ISO culture methods were examined individually, single serogroup isolation from chicken meat and turkey meat samples by the FDA method were 6 (27.3%) and 3 (60.0%), respectively. Six chicken meat samples (27.3%) were also found to harbor one serogroup by the ISO method, whereas there was no single serogroup isolation from turkey meat samples by this method. More than one serogroup was isolated from 8 of 22 (36.4%) chicken and 2 of 5 (40.0%) turkey meat samples by the FDA method, respectively, whereas these numbers were 16 of 22 (72.7%) and 5 of 5 (100.0%) by the ISO method for the same sample types (Table 1).

The total number of poultry samples with more than one serogroup isolated by the FDA and ISO methods were 10 (37.0%) and 21 (77.8%) of 27, respectively. The number of Salmonella serogroups detected after isolation from chicken meat by the FDA method were five samples (22.7%) with 2, two samples (9.1%) with 3, and one sample (4.6%) with 4 serogroups identified, whereas only two turkey meat samples were found to have 2 (40.0%) different serogroups by the FDA method. The number of Salmonella serogroups detected after isolation from chicken meat by the ISO method were 10 samples (45.5%) with 2, 4 samples (18.2%) with 3, 1 sample (4.6%) with 4, and 1 sample (4.6%) with 5 serogroups identified, whereas the number of Salmonella serogroups detected from turkey meat by the ISO method were four (80.0%) with 2 serogroups and one (20.0%) with 3 serogroups (Table 1).

The profiles of Salmonella serogroups within serogrouped isolates of chicken meat samples in descending order were serogroups D and E4 (15.8%), B and C2 (8.8%), C1 (5.3%), G (3.5%), and E1 and F (1.7%) (Table 2). The serogroup distribution within serogrouped isolates of turkey meat samples was serogroups B (27.2%), E4 (18.2%), and C2 (9.1%) (Table 2).

Table 2.

Serogrouping Results of Salmonella Isolates According to Poultry Meat Sample Types

	Number of isolates (%)
Serogroup	Chicken meat	Turkey meat	Total
Group B	5 (8.8)	3 (27.2)	8 (11.8)
Group C1	3 (5.3)	0 (0.0)	3 (4.4)
Group C2	5 (8.8)	1 (9.1)	6 (8.8)
Group D	9 (15.8)	0 (0.0)	9 (13.2)
Group E1	1 (1.7)	0 (0.0)	1 (1.5)
Group E4	9 (15.8)	2 (18.2)	11 (16.2)
Group F	1 (1.7)	0 (0.0)	1 (1.5)
Group G	2 (3.5)	0 (0.0)	2 (2.9)
Not serogrouped	22 (38.6)	5 (45.5)	27 (39.7)
Total	57 (100.0)	11 (100.0)	68 (100.0)

The total number of chicken and turkey meat serogroup isolates were 26 and 7, and 47 and 11 by the FDA and ISO methods, respectively (Table 3). Within serogrouped isolates, the distribution of chicken meat isolates obtained by the FDA method was 3 serogroup B, 8 serogroup D, and 3 serogroup E4 for chicken meat. There were two serogroup B turkey meat isolates by the FDA method. There were no C1, C2, E1, F, and G serogroup isolations by the FDA method in either sample types. Serogroups C1 (3), C2 (5), E1 (1), F (1), and G (2) were isolated only by the ISO method in chicken meat samples. Also, serogroups C2 (1) and E4 (2) were obtained only by the ISO method in turkey meat samples. The number of serogroup D isolates from 22 chicken meat samples by the FDA and ISO methods were 8 (36.4%) and 4 (18.2%), respectively. Also, the number of serogroup E4 isolates from these samples were 3 (13.6%) by the FDA method and 8 (36.3%) by the ISO method (Table 3).

Table 3.

Serogroups from Poultry Meat Isolates Based on U.S. Food and Drug Administration and International Organization for Standardization Methods

		Method
		FDA					ISO
No.	Sample ID	LTD ^a	LTT4 ^a	LR₁₀D ^a	LR₁₀T4 ^a	Total	BMD ^a	BMT4 ^a	BRD ^a	BRT4 ^a	Total	FDA and ISO ^b
1	C1^c	^d				0	C2		C2, NS	NS	2	2
2	C2					0	NS	NS	C2, NS		2	2
3	C3					0	G, NS		G, NS	G	2	2
4	C4					0	E4, NS	E4, NS	E4		2	2
5	C5					0	E4, NS	C2, B, E1, NS	E4, B	E4, NS	5	5
6	C6					0	NS		E4	E4	2	2
7	C7		D, NS			2	NS				1	2
8	C8		NS			1	C1	F, C1	E4	E4	3	4
9	C9					0	E4		E4, NS	E4	2	2
10	C10					0	C1, NS, E4		E4, B		4	4
11	C11		NS			1		C1, NS	C1, NS, G	C1	3	3
12	C22	NS	E4, B			3	NS		NS, E4	B, NS	3	3
13	C23	NS	B	E4	B	3	NS	E4	E4, NS		2	3
14	C24	E4, NS, D	B			4	B, NS	E4	B	B	3	4
15	C27	NS				1	C2		C2	C2	1	2
16	C28	NS	D	D	D	2	D	D	C2	C2	2	3
17	C29		D	D	D	1	NS	NS	NS		1	2
18	C31	NS				1	D		NS		2	2
19	C39	D	D	NS		2			NS		1	2
20	C41	NS	D	D		2		D	NS	D	2	2
21	C45	NS	NS	D		2			D	D	1	2
22	C46	D				1			NS		1	2
1	T2	NS	B			2	NS		B	B	2	2
2	T3	NS				1	NS	NS, E4	C2	C2	3	3
3	T4	NS		B		2	NS		B	B	2	2
4	T5	NS				1	B	B	B, NS	B	2	2
5	T11	NS				1			NS, E4		2	2

Bacteriological method used: first, second, and third letters are initials for preenrichment, primary enrichment, and selective plate for FDA (L, lactose broth; T, tetrathionate broth; R_10, Rappaport Vassiliadis R10 Broth; D, xylose lysine deoxycholate agar; T4, xylose lysine tergitol-4 agar) and ISO (B, buffered peptone water; M, Mueller–Kauffmann tetrathionate novobiocin broth; R, Rappaport Vassiliadis soya peptone broth).

Same serogroup results from same sample is counted once.

C, Chicken meat; T, Turkey meat.

Empty cells indicate plates not examined because of confluent growth/overgrowth by contaminant flora or absence of colony with typical morphology; C2, serogroup C2; NS, not serogrouped with available antisera; G, serogroup G; E4, serogroup E4; B, serogroup B; E1, serogroup E1; C1, serogroup C1; F, serogroup F; D, serogroup D.

Discussion

Our findings showed that chicken meat and turkey meat samples differed in both their diversity and their profile of Salmonella serogroup distribution. For example, serogroups D and E4 are the most prevalent serogroups in chicken meat samples, whereas serogroup B was the most common in turkey meat samples, as parallel to the previous reports for chicken meat (Hernandez et al., 2005; Maharjan et al., 2006; Little et al., 2008; Zaidi et al., 2008) and turkey meat (Foley et al., 2008; Little et al., 2008; Nayak and Stewart-King, 2008; Cook et al., 2009; Oloya et al., 2009). Another noticeable Salmonella serogroup difference between chicken and turkey meat samples was that serogroups C1, D, E1, F, and G were isolated only from chicken meat samples, whereas none of these serogroups was encountered in any of the turkey meat samples tested.

Even though the number of samples tested was limited, results obtained showed presence of more diverse Salmonella serogroups with higher numbers in chicken meat samples compared with turkey meat samples. This indicates that chicken meat could be a major Salmonella reservoir for human infections as also mentioned by others (Capita et al., 2003; Huong et al., 2006; Foley et al., 2008; Little et al., 2008; Nayak and Stewart-King, 2008; Cook et al., 2009; Oloya et al., 2009).

One other finding in our study is several marked differences observed between the FDA and ISO culture methods' Salmonella serogroup profile detection in poultry meat samples. The major contrast we noticed was that the serogroup D isolation by the FDA method (8/22; 36.4%) was twice higher than by the ISO method (4/22; 18.2%) in chicken meat samples. Another notable distinction for E4 serogroup isolates were as follows: they were detected more commonly in chicken meat by the ISO method than by the FDA method, and only by the ISO method in turkey meat samples. As it is well known, very important and frequently encountered poultry meat-related Salmonella serovars such as Enteritidis and Senftenberg (Capita et al., 2003; Foley et al., 2008; Little et al., 2008; Zaidi et al., 2008; Eyigor et al., 2009) reside in serogroups D and E4, respectively, and our results showed that only the use of both of these methods for a particular sample would give a more reliable serogroup profile. All these findings and the isolation of serogroups C1, C2, E1, F, and G from both poultry meat types only by the ISO method indicates a possible relation between the culture method used and the specific serogroups isolated from that sample. This, therefore, would suggest a bias toward isolating specific serogroups and underestimating other serogroups, even if they are present in the sample, if serogroup isolation is relied upon one culture method.

We observed that the ISO culture method showed a better performance than the FDA culture method in detecting several serogroups from both chicken and turkey meat samples. As the same selective plate types were used in both methods, we suspect that this difference in performances of the methods could be related either to preenrichment or more likely to primary enrichment ingredients, incubation temperature/time combinations used, and the suitability of the method for these particular sample types as also indicated by others (Arroyo and Arroyo, 1995; Waltman, 1999; Hammack et al., 2001; Feldsine et al., 2003; Schönenbrücher et al., 2008).

We found that more than one serogroup isolation per sample by the ISO method was higher than by the FDA method in both meat types. Also, five different serogroups were detected only by the ISO method. The BRD line of the ISO method in Table 3 seemed to be relatively more successful in detecting at least one serogroup per sample except for sample C7, which was found to harbor serogroup D by the LTT4 line of the FDA method as shown in the same table. This shows again that we cannot prefer one culture method over another if we would like to perceive the overall serogroup distribution in the samples tested. Instead, we have to include different selective enrichment media with different differential characteristics as previously indicated (Waltman, 1999).

The overall evaluation of our findings revealed two main facts for Salmonella serogroup isolation from poultry meat samples. The first one is that it is not uncommon for a selective plate of a specific Salmonella culture method to easily harbor colonies belonging to more than one serogroup/serovar. The second fact is that the FDA and ISO methods enabled us to detect different serogroups from chicken and turkey meats, because of differences in preenrichments and/or primary enrichments favoring/selecting for better growth/survival of certain serogroups.

Conclusion

As a conclusion, we suggest screening multiple suspect colonies from each plate, if possible, and considering the collective and comparative use of the FDA and ISO culture methods and/or including several selective and differential media to ensure the detection of Salmonella and the possible detection of multiple serogroups from samples.

Footnotes

Acknowledgments

This work was funded by the Scientific and Technical Research Council of Turkey (TUBİTAK, Project TOVAG-106O666). The authors thank Özlem Zengin for technical assistance.

Disclosure Statement

No competing financial interests exist.

References

Arroyo

, Arroyo

. Efficiency of different enrichment and isolation procedures for the detection of Salmonella serotypes in edible offal. J Appl Bacteriol, 1995; 79:360–367.

Bell

, Kyriakides

. Salmonella: A Practical Approach to the Organism and Its Control in Foods. Oxford: Blackwell, 2002; 26–27.

Berrang

, Bailey

, Altekruse

, Shaw

, Patel

, Meinersmann

, Fedorka-Cray

. Prevalence, serotype, and antimicrobial resistance of Salmonella on broiler carcasses postpick and postchill in 20 U.S. processing plants. J Food Prot, 2009; 72:1610–1615.

Bohaychuk

, Gensler

, McFall

, King

, Renter

. A real time PCR assay for the detection of Salmonella in a wide variety of food and food-animal matrices. J Food Prot, 2007; 70:1080–1087.

Bosilevac

, Guerini

, Kalchayanand

, Koohmaraie

. Prevalence and characterization of Salmonellae in commercial ground beef in the United States. Appl Environ Microbiol, 2009; 75:1892–1900.

Capita

, Alvarez-Astorga

, Allonsa-Calleja

, Moreno

, Del Camino Garcia Fernandez

. Occurrence of Salmonellae in retail chicken carcasses and their products in Spain. Int J Food Microbiol, 2003; 81:169–173.

Carli

, Kahraman

, Sen

, Sonmez

. Septicemia and blindness by Salmonella typhimurium, Salmonella enteritidis, and Salmonella essen in adult chicken. Veterinarium, 1996; 7:22–26.

Cook

, Reid-Smith

, Irwin

, McEwen

, Valdivieso-Garcia

, Ribble

. Antimicrobial resistance in Campylobacter, Salmonella, and Escherichia coli isolated from retail turkey meat from Southern Ontario, Canada. J Food Prot, 2009; 72:473–481.

[DEFRA] Department for Environment, Food and Rural Affairs. Code of Practice for the Prevention and Control of Salmonella in Chickens Reared for Meat on Farm. London: DEFRA, 2002.

10.

Dione

, Ieven

, Garin

, Marcotty

, Geerts

. Prevalence and antimicrobial resistance of Salmonella isolated from broiler farms, chicken carcasses, and street-vended restaurants in Casamance, Senegal. J Food Prot, 2009; 72:2423–2427.

11.

Eblen

, Barlow

, Naugle

. U.S. Food Safety and Inspection Service testing for Salmonella in selected raw meat and poultry products in the United States, 1998 through 2003: an establishment-level analysis. J Food Prot, 2006; 69:2600–2606.

12.

Eyigor

, Carli

, Unal

. Implementation of real-time PCR to tetrathionate broth enrichment step of Salmonella detection in poultry. Lett Appl Microbiol, 2002; 34:37–41.

13.

Eyigor

, Temelli

, Carli

. Salmonella serogroup profile in retail poultry and red meat in Turkey. 2^nd Mediterranean Summit of WPSA, Book of Proceedings, October 4–7 Antalya, Turkey, 2009; 59–61.

14.

[FDA] Food and Drug Administration. Chapter 5: Salmonella. Bacteriological Analytical Manual, 8th Andrews

, Hammack

. 2007. www.fda.gov/Food/ScienceResearch/LaboratoryMethods/BacteriologicalAnalyticalManualBAM/ucm070149.htm. 2010 February . (Online.).

15.

Fedorka-Cray

, Cox

, Richardson

, Ladely

, Buhr

. Effect of colony numbers selected from plating media on Salmonella serogroup detection from naturally contaminated chicken carcasses. International Association for Food Protection, IAFP 2009, 96^th Annual meeting, July 12–15. Grapevine, Texas, 2009. Technical Poster Abstract T1-12, 27.

16.

Feldsine

, Lienau

, Leung

, Mui

, Humbert

, Bohnert

, Mooijman

, Schulten

, in't Veld

, Rollier

, Leuschner

, Capps

. Detection of Salmonella in fresh cheese, poultry products, and dried egg products by the ISO 6579 Salmonella culture procedure and the AOAC official method: collaborative study. J AOAC Int, 2003; 86:275–295.

17.

Foley

, Lynne

, Nayak

. Salmonella challenges: prevalence in swine and poultry and potential pathogenicity of such isolates. J Anim Sci, 2008; 86:149–162.

18.

Grimont

PAD

, Weill

. Antigenic formulae of the Salmonella serovars: White-Kauffmann-Le Minor Scheme. 9 ^th WHO Collaborating Centre for Reference and Research on Salmonella, 2007. www.pasteur.fr/sante/clre/cadrecnr/salmoms-index.html. 2010 February . (Online.).

19.

Hadjinicolaou

, Demetriou

, Emmanuel

, Kakoyiannis

, Kostrikis

. Molecular beacon-based real-time PCR detection of primary isolates of Salmonella Typhimurium and Salmonella Enteritidis in environmental and clinical samples. BMC Microbiol, 2009; 9:97.

20.

Hammack

, Amaguana

, Andrews

. Rappaport-Vassiliadis medium for recovery of Salmonella spp. from low microbial load foods: collaborative study. J AOAC Int, 2001; 84:65–83.

21.

Hernandez

, Sierra

, Rodriguez–Alvarez

, Torres

, Arevalo

, Calvo

, Arias

. Salmonella enterica serotypes isolated from imported frozen chicken meat in the Canary Islands. J Food Prot, 2005; 68:2702–2706.

22.

Hoorfar

, Cook

, Malorny

, Wagner

, De Medici

, Abdulmawjood

, Fach

. Making internal amplification control mandatory for diagnostic PCR. J Clin Microbiol, 2003; 41:5835.

23.

Huong

, Fries

, Padungtod

, Hanh

, Kyule

, Baumann

MPO

, Zessin

. Prevalence of Salmonella in retail chicken meat in Hanoi, Vietnam. Ann NY Acad Sci, 2006; 1081:257–261.

24.

[ISO] International Organization for Standardization. Microbiology of Food and Animal Feeding Stuffs-Horizontal Method for the Detection of Salmonella spp. ISO 6579:2002(E) Geneva, Switzerland: International Organization for Standardization, 2002.

25.

Lee

, Jung

, Rayamahji

, Lee

, Jeon

, Choi

, Kweon

, Yoo

. A multiplex real-time PCR for differential detection and quantification of Salmonella spp., Salmonella enterica serovar Typhimurium and Enteritidis in meats. J Vet Sci, 2009; 10:43–51.

26.

Little

, Richardson

, Owen

, de Pinna

, Threlfall

. Prevalence, characterisation and antimicrobial resistance of Campylobacter and Salmonella in raw poultry meat in the UK, 2003–2005. Int J Environ, 2008; 18:403–414.

27.

Lofstrom

, Knutsson

, Axelsson

, Radstrom

. Rapid and specific detection of Salmonella spp. in animal feed samples by PCR after culture enrichment. Appl Environ Microbiol, 2004; 70:69–75.

28.

Maharjan

, Joshi

, Manandhar

. Prevalence of Salmonella species in various raw meat samples of a local market in Kathmandu. Ann NY Acad Sci, 2006; 1081:249–256.

29.

Malorny

, Bunge

, Helmuth

. A real-time PCR for the detection of Salmonella Enteritidis in poultry meat and consumption eggs. J Microbiol Methods, 2007; 70:245–251.

30.

Miranda

, Mondragon

, Martinez

, Guarddon

, Rodriguez

. Prevalence and antimicrobial resistance patterns of Salmonella from different raw foods in Mexico. J Food Prot, 2009; 72:966–971.

31.

Nayak

, Stewart-King

. Molecular epidemiological analysis and microbial source tracking of Salmonella enterica serovars in a preharvest turkey production environment. Foodborne Pathog Dis, 2008; 5:115–126.

32.

Oloya

, Doetkott

, Khaitsa

. Antimicrobial drug resistance and molecular characterization of Salmonella isolated from domestic animals, humans, and meat products. Foodborne Pathog Dis, 2009; 6:273–284.

33.

O'Regan

, McCabe

, Burgess

, McGuinness

, Barry

, Duffy

, Whyte

, Fanning

. Development of a real-time multiplex PCR assay for the detection of multiple Salmonella serotypes in chicken samples. BMC Microbiol, 2008; 8:156.

34.

Poppe

. Epidemiology of Salmonella enterica serovar Enteritidis. Salmonella Enterica Serovar Enteritidis in Humans and Animals: Epidemiology, Pathogenesis, and Control. Saeed

, Gast

, Wall

. Ames, Iowa: Iowa State University Press, 1999; 3–18.

35.

Schönenbrücher

, Mallinson

, Bülte

. A comparison of standard cultural methods for the detection of foodborne Salmonella species including three new chromogenic plating media. Int J Food Microbiol, 2008; 123:61–66.

36.

Schrank

, Mores

, Costa

, Frazzon

, Soncini

, Schrank

, Vainstein

, Silva

. Influence of enrichment media and application of a PCR based method to detect Salmonella in poultry industry products and clinical samples. Vet Microbiol, 2001; 82:45–53.

37.

[USDA] United States Department of Agriculture Food Safety and Inspection Service, Office of Public Health Science. Isolation and Identification of Salmonella from Meat, Poultry and Eggs Products. Laboratory QA/QC Division: 950 College Station Road Athens, GA, 2008.

38.

Waltman

. Epidemiology of Salmonella enterica serovar Enteritidis. Salmonella Enterica Serovar Enteritidis in Humans and Animals: Epidemiology, Pathogenesis, and Control. Saeed

, Gast

, Wall

. Ames, Iowa: Iowa State University Press, 1999; 419–432.

39.

Woods

, Reen

, Gilroy

, Buckley

, Frye

, Boyd

. Rapid multiplex PCR and real-time TaqMan PCR assays for detection of Salmonella enterica and the highly virulent serovars Choleraesuis and Paratyphi C. J Clin Microbiol, 2008; 46:4018–4022.

40.

Zaidi

, Calva

, Estrada-Garcia

, Leon

, Vazquez

, Figueroa

, Lopez

, Contreras

, Abbott

, Zhao

, McDermott

, Tollefson

. Integrated food chain surveillance system for Salmonella spp. in Mexico. Emerg Infect Dis, 2008; 14:429–435.