628

Journal of Food Protection, Vol. 83, No. 4, 2020, Pages 628–636

https://doi.org/10.4315/0362-028X.JFP-19-359

Copyright , International Association for Food Protection

Research Paper

Confounding Role of Salmonella Serotype Dublin Testing Results

of Boneless and Ground Beef Purchased for the National School

Lunch Program, October 2013 to July 2017

SCOTT L. VIAL,1 DARIN R. DOERSCHER,2 CARL M. SCHROEDER,2 ALI J. STRICKLAND,1 AND CRAIG W. HEDBERG1*

1Environmental Health Sciences, School of Public Health, University of Minnesota, 420 East Delaware Street, Minneapolis, Minnesota 55455

(ORCID: https://orcid.org/000-0003-3354-6811 [S.L.V.]); and 2Livestock and Poultry Program, Agricultural Marketing Service, U.S. Department of

Agriculture, 1400 Independence Avenue, Washington, D.C. 20250, USA

MS 19-359: Received 1 August 2019/Accepted 3 December 2019/Published Online 18 March 2020

ABSTRACT

The Agricultural Marketing Service procures boneless and ground beef for federal nutrition assistance programs. It tests

procured beef for concentrations of standard plate counts (SPCs), coliforms, and Escherichia coli and for the presence of

Salmonella and Shiga toxin–producing E. coli. Any lot exceeding predefined critical limits (100,000 CFU g 1 for SPCs, 1,000

CFU g 1 for coliforms, and 500 CFU g 1 for E. coli) or positive for Salmonella or Shiga toxin–producing E. coli is rejected for

purchase. Between 1 October 2013 and 31 July 2017, 166,796 boneless beef lots (each approximately 900 kg) and 25,051

ground beef sublots (each approximately 4,500 kg) were produced. Salmonella was detected in 1,955 (1.17%) boneless beef lots

and 219 (0.87%) ground beef sublots. Salmonella sample size increased from an individual 25-g sample to a co-enriched 325-g

sample on 1 March 2015. Salmonella presence was associated with season (lowest in spring), larger sample size, and increased

log SPC in boneless and ground beef. Increased log E. coli was associated with Salmonella presence in boneless beef, but not

ground beef. Salmonella Dublin was the most common serotype in boneless beef (743 of 1,407, 52.8%) and ground beef (35 of

171, 20.5%). Salmonella Dublin was generally associated with lower indicator microorganism concentrations compared with

other Salmonella serotypes as a group. Relative to other Salmonella, Salmonella Dublin was associated with season (more

common in spring) and smaller sample size in boneless and ground beef. Decreased log SPCs and log coliforms were associated

with Salmonella Dublin presence in boneless beef, but not in ground beef. Differential associations between Salmonella Dublin

and other serotypes with indicator microorganisms were strong enough to cause confounding and suggest that the presence of

Salmonella Dublin needs to be accounted for when evaluating indicator performance to assess Salmonella risk in boneless and

ground beef.

HIGHLIGHTS

Salmonella was associated with indicator microorganism detection and concentration.

Salmonella Dublin represented 52.8% of Salmonella in boneless and 20.5% in ground beef.

Salmonella Dublin was associated with lower indicator levels compared with other Salmonella.

Key words: Beef; National School Lunch Program; Salmonella serotype Dublin

Salmonella causes an estimated 1,000,000 cases,

19,000 hospitalizations, and 350 deaths annually in the

United States (33). Salmonella infection has a high

economic burden, costing an estimated $3.7 billion annually

in the United States (15). There are more than 2,600

Salmonella serotypes and about 360 serotypes that cause

human illness. Of these, 20 serotypes account for 69% of all

human Salmonella illnesses (4, 30).

Approximately 7% of human Salmonella infections are

attributed to contaminated beef consumption (18). Cattle

may become infected in farm or lairage environments

* Author for correspondence. Tel: 612-626-4757; Fax: 612-626-4837;

E-mail: hedbe005@umn.edu.

before or during transit to abattoirs (22, 32). Beef intended

for human consumption can become contaminated during

the production process if it comes into contact with

Salmonella on cattle hides, colonized lymph nodes, or

feces or through cross-contamination of production surfaces

and tools (2, 20, 40). In recent years, producers and

regulators instituted large-scale ground beef recalls because

of foodborne outbreaks associated with Salmonella contamination (11). In 2018, for example, a large outbreak of

Salmonella Newport involving 403 illnesses and 117

hospitalizations across 30 states was linked to ground beef

consumption and resulted in a recall of 12 million lb of

ground beef (5).
J. Food Prot., Vol. 83, No. 4 SALMONELLA DUBLIN IN BEEF PRODUCTS 629

Salmonella Dublin is a cattle-adapted serotype and is

associated with dairy cattle (16). Salmonella Dublin was the

most common bovine clinical isolate submitted to the

National Veterinary Service Laboratory in both 2016 and

2017 (25, 26). It was the second most common in 2015 (21).

Salmonella Dublin was the most prevalent Salmonella

serotype in a large study of U.S. dairy cattle, mostly from

Wisconsin, with clinical symptoms, accounting for 23% of

isolates (38). Salmonella Dublin is also commonly detected

in nonclinical bovine samples. The National Veterinary

Service Laboratory reported Salmonella Dublin was the

fourth most common nonclinical Salmonella serotype in

2015 and 2017 (21). It may also be found in lymph nodes at

harvest (40). An additional study of bulk milk tanks in New

York found Salmonella Dublin in 4% of all samples and in

60% of all Salmonella-positive samples (6).

Salmonella transmission between cattle typically occurs via the fecal-oral route but also rarely occurs via

respiratory or conjunctiva routes (13, 14, 27). Salmonella

infections in pregnant cows can cross the placenta, leading

to calf abortions (13, 27). Clinical signs of Salmonella

infections in cows include diarrhea, pneumonia, abortion,

and death (14, 38). Clinical severity varies with age and is

most severe in cattle younger than 3 months, with neonatal

diarrhea being a particular challenge (27, 37). Salmonella

Dublin carrier status is a well-documented problem in cattle

(6, 14). Carrier animals may maintain Salmonella Dublin in

their lymph nodes or internal organs (3, 13, 14). These

animals may continue to shed Salmonella Dublin constantly

or intermittently, or they may become latent carriers. The

presence of carrier cattle with subclinical Salmonella

Dublin infections complicates its controls on farms and

ranches (13, 14, 16, 22).

Salmonella Dublin has not been frequently associated

with human illness and was not listed among the top 20

culture-confirmed human serotypes reported to the Centers

for Disease Control and Prevention (CDC) in 2016 (4).

However, Salmonella Dublin infection incidence rates in

humans have increased faster than rates of other Salmonella

serotypes in recent years. A study determined Salmonella

Dublin infections in humans were significantly more severe

than Salmonella infections caused by other serotypes:

infection with Salmonella Dublin was more likely to result

in sepsis, more likely to require hospitalization, and more

likely to cause death (12). Furthermore, Salmonella Dublin

incidence was higher than that of other Salmonella

serotypes in adults older than 18 years, but a lower

proportion of children aged 5 to 17 years were affected by

Salmonella Dublin compared with other Salmonella serotypes in the United States.

Salmonella Dublin frequently displays antimicrobial

resistance (AMR), with up to 79% of isolates being AMR

(12, 16, 38). AMR complicates treatment and leads to more

serious illnesses. Furthermore, Salmonella Dublin AMR

increased significantly over the last 20 years. Given the

seriousness of Salmonella Dublin infection, its increasing

incidence, and AMR prevalence, it is a notable human

public health concern.

An outbreak of Salmonella Dublin in August 2019

resulted in 10 reported cases in the United States (8). Of the

10 cases, 8 were hospitalized, and 1 died. The Centers for

Disease Control and Prevention estimates that typical (non–

Dublin-specific) Salmonella infections only result in

hospitalization in approximately 20% of all cases. Further

evidence that this outbreak led to more severe health

outcomes than general Salmonella outbreaks was the

Salmonella Dublin presence in blood samples for five

cases. Cases presented in six states and occurred predominately among older men. These severe human health

outcomes call for a more in-depth analysis of the

Salmonella Dublin ecology.

Indicator microorganisms are used during beef and

other meat production and processing to monitor food

quality, hygiene, and process control (31). Monitoring and

controlling processes with these indicator organisms is

intended to reduce the likelihood of pathogen presence (1).

However, multiple studies have shown a sporadic to weak

correlation between presence of indicator microorganisms

and Salmonella (7, 9, 19). One study of the association

between indicator microorganisms and Salmonella serotypes in lymph nodes found highly variable indicator

concentrations by lymph node location and serotype,

including Salmonella Dublin (3).

Data and analysis of the U.S. Department of Agriculture Agricultural Marketing Service (AMS) boneless and

ground beef purchasing programs from 2011 to 2018

determined Salmonella Dublin accounted for 51 and

17.2% of Salmonella isolates recovered from boneless and

ground beef, respectively (7, 39). These studies examined

associations between contract-established critical limit

exceedance and pathogen presence. Weak associations

between critical limit exceedance and Salmonella presence

were found. Potential associations between Salmonella

presence and indicator microorganism concentrations on

broader, continuous scales and potential variation associated

with specific Salmonella serotypes have not been explored.

Here we examine AMS data for October 2013 through

July 2017. Our goals are twofold. First, we describe

associations between indicator microorganism concentration and pathogen presence, along with season and AMS

purchase requirements. Second, we investigate Salmonella

Dublin’s unique relationship with these same factors

relative to Salmonella-negative lots and to lots positive

for other Salmonella serotypes.

MATERIALS AND METHODS

Sampling and analysis. Samples of boneless and ground

beef scheduled for raw, uncooked delivery at school food service

and similar facilities were collected and processed as previously

described (7, 39). Trained vendor personnel at each establishment

collected samples using a scalpel excision or drill-sampling device

for boneless beef and by grab sampling for boneless beef. Samples

were packaged, placed on dry ice, and shipped overnight to AMSdesignated laboratories for analysis in accordance with AMS

purchase specifications (34). AMS-designated laboratories performed all microbiological analyses according to the Food Safety

and Inspection Service (FSIS) Microbiological Laboratory

Guidebook (36). Two FSIS and AMS testing protocol changes

occurred in early 2015. First, the boneless beef Salmonella sample

size was increased from 25 to 325 g (35). Second, the Salmonella
 



630 VIAL ET AL. J. Food Prot., Vol. 83, No. 4

analysis method was changed from the individual 25-g sample to a

325-g co-enriched sample also used for E. coli O157:H7 testing

(35). AMS required its vendors to switch sampling methods in

March 2015. Salmonella isolates recovered by AMS-designated

laboratories were sent to the FSIS Eastern Laboratory for

serotyping (24). AMS maintains all results electronically.

More boneless beef than ground beef lots were sampled

between 2013 and 2017, because AMS purchase specifications

require testing of every 2,000 lb of boneless beef compared with

every 10,000 lb of ground beef. In addition, boneless beef sublots

found positive are discarded and new replacement sublots are

tested before grinding. Lastly, some boneless beef is used without

grinding.

Data analysis. Indicator microorganism and Salmonella data

for 166,796 AMS boneless beef lots produced by 15 vendors and

25,051 resultant ground beef sublots produced by 10 vendors

between 1 October 2013 and 31 July 2017 were analyzed. Critical

limits were defined as 100,000 CFU g 1 for standard plate counts

(SPCs), 1,000 CFU g 1 for coliforms, and 500 CFU g 1 for E. coli.

All data analyses were conducted using STATA 14.2 (34). Initially,

univariable associations between indicator microorganism detection, season, and pathogen sample size with Salmonella presence

were determined. Seasons were defined as spring (March to May),

summer (June to August), fall (September to November), and

winter (December to February). Spring was used as the seasonal

referent category. Seasonal associations were determined using

Pearson chi-square tests. The Salmonella sample size was defined

as 25 g for lots produced before 1 March 2015 and as 325 g for

lots produced on or after 1 March 2015. Boneless beef lots and

ground beef sublots without detectable SPCs, total coliforms, or E.

coli were considered to have concentrations of 0 log CFU g 1

.

The boneless beef and ground beef analyses considered three

outcomes. The primary outcome of interest was Salmonella

presence. Salmonella-positive samples were further stratified into

two outcome groups. One outcome group consisted of all

Salmonella Dublin isolates. The other outcome group consisted

of Salmonella serotypes other than Salmonella Dublin. Associations between indicator microorganism detection (SPCs, total

coliforms, and E. coli) and outcome (all Salmonella serotypes,

Salmonella Dublin, or a serotype other than Salmonella Dublin)

were determined using 2 by 2 tables to calculate odds ratios (ORs)

and 95% confidence intervals (95% CI). Univariable associations

among log SPCs, log total coliforms, and log E. coli with each

outcome were determined using logistic regression to determine

ORs and 95% CI. Multivariable associations between seasons,

pathogen sample sizes, log SPCs, log total coliforms, and log E.

coli with each outcome were determined using mixed-effects

general linear modeling with nesting by the vendor. General linear

modeling was completed using stepwise deletion of nonsignificant

variables. Final models reported ORs and 95% CI and included

only those variables significantly associated with outcome.

RESULTS AND DISCUSSION

Salmonella presence in boneless and ground beef.

Salmonella was present in 1,955 (1.17%) boneless beef lots

(Table 1). Salmonella prevalence differed significantly in

univariable analysis by season (χ

2

3 1⁄4 125.33, P , 0.001)

pathogen sample size (OR 1⁄4 2.16, 95% CI: 1.96 to 2.39),

and vendor (χ2

14 1⁄4 576.8, P , 0.001). Salmonella

prevalence was highest in the fall, followed by winter,

summer, and spring. Of the 15 vendors, 13 produced at least

one Salmonella-positive lot. Salmonella prevalence ranged

TABLE 1. AMS vendor Salmonella performance

Boneless beef Ground beef

Salmonella positives Salmonella positives

No. of No. of

lots n % sublots n %

Overall 166,796 1,955 1.17 25,051 219 0.87

Seasona

Spring 23,647 156 0.66 3,500 18 0.51

Summer 31,873 325 1.02 4,514 59 1.31

Fall 58,085 887 1.53 8,831 73 0.83

Winter 53,191 587 1.10 8,206 69 0.84

Sample sizeb

25 g 74,350 533 0.72 14,055 95 0.68

325 g 92,446 1,422 1.54 10,996 124 1.13

a Boneless beef: χ2

3 1⁄4 125.3, P , 0.001. Ground beef: χ23 1⁄4 15.3,

P 1⁄4 0.002. b 325-g co-enriched sampling began 1 March 2015. Boneless beef:

OR 1⁄4 2.16, 95% CI: 1.96 to 2.39. Ground beef: OR 1⁄4 1.68, 95%

CI: 1.27 to 2.22.

by vendor from 3.41 to 0.00%. Salmonella was significantly

more likely to be recovered using the larger sample size,

with 1.54% of lots tested from March 2015 onward found

positive for Salmonella compared with 0.72% lots produced

before March 2015.

During the same period, AMS vendors produced

25,051 sublots of ground beef scheduled for raw, uncooked

delivery to school food service and similar facilities (Table

1). These sublots were produced by 10 vendors using AMS

boneless beef that had met AMS indicator microorganism

and pathogen testing purchase specifications. Salmonella

was detected in 219 (0.87%) ground beef sublots.

Prevalence differed significantly by season (χ

2

3 1⁄4 15.3, P

1⁄40.002), pathogen sample size (OR1⁄41.68, 95% CI: 1.27 to

2.22), and vendor (χ2

9 1⁄4 198.39, P , 0.001). Salmonella

was most commonly found in ground beef during the

summer, followed by winter, fall, and spring. All 10 vendors

produced at least one Salmonella-positive ground beef

sublot, and prevalence ranged from 3.37 to 0.06%.

Salmonella was significantly more likely to be recovered

from sublots produced after 1 March 2015 (1.13%)

compared with those produced earlier (0.68%).

Average indicator microorganism concentrations were

low but positively skewed in both boneless and ground beef.

These concentrations remained skewed following log

transformation (Table 2). Skewness was partly because of

relatively high proportions of boneless beef lots and ground

beef sublots containing nondetectable concentrations of

indicator microorganisms. In boneless beef, 16.7% of lots

had nondetectable SPCs, 91.7% had nondetectable total

coliforms, and 98.1% had nondetectable E. coli. In ground

beef, 21.1% of sublots had nondetectable SPCs, 87.2% had

nondetectable coliforms, and 95.3% had nondetectable E.

coli.

Indicator microorganism detection was consistently

associated with Salmonella presence (Table 3). In boneless

beef, Salmonella was significantly more likely to be
J. Food Prot., Vol. 83, No. 4 SALMONELLA DUBLIN IN BEEF PRODUCTS 631

TABLE 2. AMS vendor indicator organism performance detected when SPCs (OR 1⁄4 1.62, 95% CI: 1.10 to 2.46),

1

) Concn (log CFU g coliforms (OR 1⁄4 2.79, 95% CI: 2.04 to 3.77), or E. coli (OR

1⁄4 3.12, 95% CI: 2.02 to 4.65) were detected.

Boneless beef Ground beef Increased indicator microorganism log concentrations

were consistently associated with Salmonella presence in

Total Total univariable analysis. In boneless beef, Salmonella detection Indicatora SPCs coliforms E. coli SPCs coliforms E. coli

was significantly associated with increased log SPCs (OR 1⁄4

Maximum 6.34 5.30 4.41 5.60 4.30 3.64 1.15, 95% CI: 1.10 to 1.20), log total coliforms (OR 1⁄4 1.26,

Q3 2.32 0.00 0.00 2.52 0.00 0.00 95% CI: 1.14 to 1.40), and log E. coli (OR 1⁄4 1.77, 95% CI:

Median 1.78 0.00 0.00 1.95 0.00 0.00 1.53 to 2.06). In ground beef, Salmonella was also

Q1 1.00 0.00 0.00 1.00 0.00 0.00 significantly associated with increased log SPCs (OR 1⁄4 Minimum 0.00 0.00 0.00 0.00 0.00 0.00 1.45, 95% CI: 1.27 to 1.67), log total coliforms (OR 1⁄4 1.87, a Q3, third quarter; Q1, first quarter. 95% CI: 1.56 to 2.24), and log E. coli (OR 1⁄4 2.01, 95% CI:

1.54 to 2.63). Using multivariable modeling, Salmonella

detected when SPCs (OR 1⁄4 1.50, 95% CI: 1.30 to 1.72), detection in boneless beef was significantly associated with

total coliforms (OR1⁄4 1.34, 95% CI: 1.15 to 1.55), or E. coli season, larger 325-g sample size (OR 1⁄4 2.68, 95% CI: 2.41

(OR 1⁄4 2.24, 95% CI: 1.76 to 2.81) were detected. In ground to 2.99), increased log SPCs (OR 1⁄4 1.15, 95% CI: 1.09 to

beef, Salmonella was significantly more likely to be 1.22), and increased log E. coli (OR1⁄41.65, 95% CI: 1.41 to

TABLE 3. AMS vendor boneless beef indicator detection and Salmonella presence

Boneless beef Ground beef

Outcome positive Outcome positive

No. of No. of

Outcome Indicator lots n % OR 95% CI sublots n % OR 95% CI

1 1⁄4 Salmonella positive, SPCs

0 1⁄4 Salmonella negative Detected 139,024 1,724 1.24 1.50 1.30–1.72 19,799 188 0.95 1.62 1.10–2.46

Not detected 27,772 231 0.83 5,272 31 0.59

Total coliforms

Detected 13,829 210 1.52 1.34 1.15–1.55 3,205 63 1.97 2.79 2.04–3.77

Not detected 152,967 1,745 1.14 21,846 156 0.71

E. coli

Detected 3,121 79 2.53 2.24 1.76–2.81 1,188 29 2.44 3.12 2.02–4.65

Not detected 163,675 1,876 1.15 23,863 190 0.80

1 1⁄4 Dublin positive, SPCs

0 1⁄4 other Salmonella Detected 1,234 621 50.32 0.42 0.29–0.60 147 30 20.41 0.97 0.32–3.61

serotype positive Not detected 173 122 70.52 24 5 20.83

Total coliforms

Detected 153 35 22.88 0.23 0.15–0.34 46 5 10.87 0.39 0.11–1.11

Not detected 1,254 708 56.46 125 30 24.00

E. coli

Detected 58 5 8.62 0.08 0.02–0.20 23 2 8.70 0.33 0.04–1.48

Not detected 1,349 738 54.71 148 33 22.30

1 1⁄4 Dublin positive, SPCs

0 1⁄4 Salmonella Detected 137,921 621 0.45 1.02 0.84–1.25 19,621 30 0.15 1.61 0.62–5.30

negative Not detected 27,663 122 0.44 5,246 5 0.10

Total coliforms

Detected 13,654 35 0.26 0.55 0.38–0.77 3,147 5 0.16 1.15 0.35–3.00

Not detected 151,930 708 0.47 21,720 30 0.14

E. coli

Detected 3,047 5 0.16 0.36 0.12–0.84 1,161 2 0.17 1.24 0.14–4.85

Not detected 162,537 738 0.45 23,706 33 0.14

1 1⁄4 Other Salmonella SPCs

serotype positive, 0 1⁄4 Detected 137,913 613 0.44 2.41 1.81–3.28 19,708 117 0.59 1.65 1.01–2.84

Salmonella negative Not detected 27,592 51 0.18 5,260 19 0.36

Total coliforms

Detected 13,737 118 0.86 2.40 1.95–2.94 3,183 41 1.29 2.97 2.01–4.35

Not detected 151,768 546 0.36 21,785 95 0.44

E. coli

Detected 3,095 53 1.71 4.61 3.41–6.13 1,180 21 1.78 3.73 2.22–6.00

Not detected 162,410 611 0.38 23,788 115 0.48
632 VIAL ET AL. J. Food Prot., Vol. 83, No. 4

TABLE 4. General linear models for AMS boneless beef and Salmonella presencea

Boneless beef Ground beef

Outcome Variable OR 95% CI OR 95% CI

1 1⁄4 Salmonella positive, 0 1⁄4 Salmonella negative Sample size 2.68 2.41–2.99 1.55 1.12–2.14

log SPCs 1.15 1.09–1.22 1.23 1.03–1.47

log E. coli 1.65 1.41–1.94

Season

Summer 1.52 1.25–1.85 2.47 1.41–4.31

Fall 2.79 2.35–3.32 1.56 0.90–2.71

Winter 2.15 1.79–2.58 1.75 1.00–3.06

1 1⁄4 Salmonella Dublin positive, 0 1⁄4 other Salmonella Sample size 0.63 0.45–0.90 0.04 0.00–0.32

serotype positive log SPCs 0.63 0.52–0.76

log total coliforms 0.46 0.31–0.70

Season

Summer 0.16 0.08–0.32 0.25 0.02–2.87

Fall 0.31 0.17–0.58 0.01 0.00–0.14

Winter 0.16 0.08–0.30 0.08 0.01–0.87

1 1⁄4 Salmonella Dublin positive, 0 1⁄4 Salmonella negative Sample size 3.95 3.32–4.71 0.16 0.05–0.58

log SPCs 1.12 1.02–1.23

log total coliforms 0.68 0.50–0.91

Season

Summer 0.97 0.71–1.33 0.97 0.32–2.97

Fall 2.62 2.04–3.37 0.04 0.01–0.22

Winter 1.81 1.39–2.36 0.41 0.14–1.18

1 1⁄4 Other Salmonella serotype positive, 0 1⁄4 Salmonella Sample size 2.22 1.84–2.66 2.12 1.41–3.19

negative log SPCs 1.29 1.17–1.43 1.27 1.01–1.60

log E. coli 1.89 1.57–2.29

Season

Summer 2.25 1.58–3.19 2.87 1.35–6.11

Fall 2.54 1.82–3.56 2.61 1.26–5.43

Winter 2.79 1.98–3.92 2.23 1.04–4.76

a Initial general linear models considered season, sample size, and concentrations of log SPCs, log total coliforms, and log E. coli with

nesting by vendor. Spring was the seasonal referent.

1.94) (Table 4). In ground beef, Salmonella detection was

associated with season, larger sample size (OR 1⁄4 1.55, 95%

CI: 1.12 to 2.14), and increased log SPC (OR 1⁄4 1.23, 95%

CI: 1.03 to 1.47).

Serotyping was done for 1,407 (72.0%) of 1,955

Salmonella isolates recovered from boneless beef lots

(Table 5). Most of these isolates were Salmonella Dublin

(n 1⁄4 743, 52.8%), followed by Salmonella serotypes

Newport (n 1⁄4 147, 10.5%), Montevideo (n 1⁄4 98, 7.0%),

Anatum (n 1⁄4 47, 3.3%), Typhimurium (n 1⁄4 45, 3.2%), and

Muenchen (n 1⁄4 38, 2.7%). The proportion of serotyped

Salmonella isolates determined to be Salmonella Dublin

differed significantly in the univariable analysis by season

(χ

2

3 1⁄4 42.5, P , 0.001) and vendor (χ

2

14 1⁄4 502.3, P ,

0.001). Salmonella Dublin was the most common serotype

in all four seasons, accounting for 65.8% of serotypes in

spring, 37.0% in summer, 58.7% in fall, and 49.3% in

winter. Among vendors with at least 30 serotyped

Salmonella isolates, the proportion of Salmonella Dublin

isolates ranged from 88.7 to 1.6%. Unlike overall

Salmonella prevalence, Salmonella Dublin prevalence

among serotyped isolates did not differ significantly by

pathogen sample size (OR 1⁄4 1.06, 95% CI: 0.83 to 1.35).

In ground beef, serotypes were determined for 171

(78.4%) of 219 Salmonella-positive sublots (Table 5). The

most common Salmonella serotype was Salmonella Dublin

(n 1⁄4 35, 20.5%), followed by Salmonella serotypes

Montevideo (n 1⁄4 27, 15.8%), Newport (n 1⁄4 20, 11.7%),

Anatum (n 1⁄4 16, 9.4%), and Typhimurium (n 1⁄4 11, 6.4%).

The proportion of serotyped Salmonella isolates determined

to be Salmonella Dublin differed significantly in the

univariable analysis by season (χ

2

3 1⁄4 17.8, P , 0.001)

and vendor (χ

2

10 1⁄4 52.2, P , 0.001). Salmonella Dublin

was the most common individual serotype found in each

season except fall. Salmonella Dublin accounted for 43.8%

of serotypes in spring, 24.4% in summer, and 28.3% in

winter. It accounted for 3.5% of serotypes in fall. Among

vendors with at least 15 serotyped Salmonella isolates,

Salmonella Dublin prevalence ranged from 54.2 to 0% of

isolates.

Salmonella Dublin compared with other Salmonella

serotypes. In boneless beef, Salmonella Dublin–positive

lots (n 1⁄4 743) were inversely associated with indicator

microorganism concentrations compared with other Salmonella serotypes as a group (n 1⁄4 664). Indicator microorganisms were less likely to be detected in Salmonella
J. Food Prot., Vol. 83, No. 4 SALMONELLA DUBLIN IN BEEF PRODUCTS 633

TABLE 5. AMS boneless beef Salmonella serotype performance

Boneless beef Ground beef

No. of

lots

Salmonella Dublin

positives

n %

Other Salmonella

serotypes

n %

No. of

sublots

Salmonella Dublin

positives

n %

Other Salmonella

serotypes

n %

Overall 1,407 743 52.8 664 47.2 171 35 20.5 136 79.5

Seasona

Spring

Summer

Fall

Winter

120

235

618

434

79

87

363

214

65.8

37.0

58.7

49.3

41

148

255

220

34.2

63.0

41.3

50.7

16

45

57

53

7

11

2

15

43.8

24.4

3.5

28.3

9

34

55

38

56.3

75.6

96.5

71.7

Salmonella sample sizeb

25 g 383

325 g 1,024

198

545

51.7

53.2

185

479

48.3

46.8

72

99

21

14

29.2

14.1

51

85

70.8

85.9

a Boneless beef: Salmonella Dublin, χ2

3 1⁄4 42.5, P , 0.001. Ground beef: Salmonella Dublin, χ

2

3 1⁄4 17.8, P , 0.001. b 325-g co-enriched sampling began 1 March 2015. Boneless beef: Salmonella Dublin, OR 1⁄4 1.06, 95% CI: 0.83 to 1.35. Ground beef:

Salmonella Dublin, OR 1⁄4 0.40, 95% CI: 0.17 to 0.91.

Dublin–positive lots and, when detected, were present in

lower concentrations compared with lots positive for other

Salmonella serotypes. Salmonella Dublin–positive boneless

beef lots were significantly less likely to contain detectable

SPCs (OR 1⁄4 0.42, 95% CI: 0.29 to 0.60), total coliforms

(OR 1⁄4 0.23, 95% CI: 0.15 to 0.34), and E. coli (OR 1⁄4 0.08,

95% CI: 0.02 to 0.20) than lots that tested positive for other

Salmonella serotypes (Table 3). Furthermore, these Salmonella Dublin–positive lots were significantly lower in log

SPCs (OR1⁄4 0.63, 95% CI: 0.56 to 0.71), log total coliforms

(OR 1⁄4 0.29, 95% CI: 0.21 to 0.41), and log E. coli

concentration (OR 1⁄4 0.10, 95% CI: 0.04 to 0.25) using

univariable analysis. A multivariable model determined

Salmonella Dublin was significantly associated with season,

pathogen sample size (OR 1⁄4 0.63, 95% CI: 0.45 to 0.90),

log SPCs (OR 1⁄4 0.63, 95% CI: 0.52 to 0.76), and log total

coliform concentration (OR 1⁄4 0.46, 95% CI: 0.31 to 0.70)

compared with other Salmonella serotypes (Table 4).

In ground beef, indicator microorganism detection and

concentration rarely differed significantly between Salmonella Dublin and other Salmonella serotypes. There were no

significant differences in indicator microorganism detection

(SPCs, total coliforms, or E. coli) between Salmonella

Dublin and other Salmonella serotypes (Table 3). However,

Salmonella Dublin was associated with lower log SPC

concentration compared with other Salmonella serotypes

(OR 1⁄4 0.71, 95% CI: 0.51 to 0.99). There were no similar

associations with log coliform or log E. coli concentration.

In the ground beef multivariable model, season and

pathogen sample size were significantly associated with

Salmonella Dublin, but indicator microorganism concentrations were not (Table 4). Salmonella Dublin was significantly less likely to be detected relative to the other

Salmonella serotypes in ground beef using the larger

pathogen sample size after March 2015 (OR 1⁄4 0.04, 95%

CI: 0.00 to 0.32).

Salmonella Dublin–positive lots compared with

Salmonella-negative lots. Salmonella Dublin–positive

boneless beef lots were significantly less likely to contain

detectable total coliforms (OR1⁄40.55, 95% CI: 0.38 to 0.77)

or detectable E. coli (OR 1⁄4 0.36, 95% CI: 0.12 to 0.84)

compared with Salmonella-negative lots (Table 3). Using

univariable analysis, lots with higher concentrations of log

SPCs (OR 1⁄4 0.92, 95% CI: 0.85 to 0.99), log coliforms (OR

1⁄4 0.58, 95% CI: 0.43 to 0.77), and log E. coli (OR 1⁄4 0.92,

95% CI: 0.85 to 0.99) were less likely to contain Salmonella

Dublin than lots with lower indicator microorganism

concentrations. In the boneless beef multivariable model,

Salmonella Dublin was significantly associated with larger

sample size (OR 1⁄4 3.95, 95% CI: 3.32 to 4.71), higher log

SPCs (OR1⁄41.12, 95% CI: 1.02 to 1.23), and lower log total

coliform concentration (OR 1⁄4 0.68, 95% CI: 0.50 to 0.91)

compared with Salmonella-negative lots (Table 4).

In ground beef, however, there were no significant

associations between Salmonella Dublin presence and

indicator microorganism detection compared with the

Salmonella-negative sublots. In univariable analysis, only

log SPC concentration was significantly associated with

Salmonella Dublin, which had a protective effect (OR 1⁄4

0.71, 95% CI: 0.51 to 0.99) as concentration increased. In

the multivariable model, Salmonella Dublin was significantly associated with season and sample size, but not with

indicator organism concentrations. The multivariable model

found Salmonella Dublin was significantly more likely in

ground beef sublots sampled using the smaller, pre-March

2015 sampling protocol than in the larger, post-March 2015

samples (OR 1⁄4 0.16, 95% CI 1⁄4 0.05 to 0.58). Curiously,

sample size had strong but opposite associations with

Salmonella Dublin in boneless and ground beef. In boneless

beef, Salmonella Dublin was significantly more likely to be

present in the later and larger samples, but in ground beef, it

was less likely to be present in the later and larger samples.

Salmonella isolated from sublots produced after 1 March

2015 were less likely to be Salmonella Dublin (14.1%) than
634 VIAL ET AL. J. Food Prot., Vol. 83, No. 4

lots produced earlier (29.2%) (OR 1⁄4 0.40, 95% CI: 0.17 to

0.91).

Other Salmonella serotypes compared with Salmonella-negative lots. Indicator microorganism detection was

significantly associated with the detection of Salmonella

serotypes other than Salmonella Dublin (Table 3). In

boneless beef, non–Salmonella Dublin serotypes were

significantly associated with SPC detection (OR 1⁄4 2.41,

95% CI: 1.81 to 3.28), coliform detection (OR 1⁄4 2.40, 95%

CI: 1.95 to 2.94), and E. coli detection (OR 1⁄4 4.61, 95% CI:

3.41 to 6.13). Under univariable analysis, boneless beef lots

were significantly more likely to contain non–Salmonella

Dublin serotypes as log SPCs (OR 1⁄4 1.37, 95% CI: 1.26 to

1.49), log total coliforms (OR1⁄41.83, 95% CI: 1.61 to 2.08),

and log E. coli (OR1⁄42.61, 95% CI: 2.19 to 3.10) increased.

In the multivariable model, non–Salmonella Dublin serotypes were significantly associated with season, larger

sample size (OR 1⁄4 2.22, 95% CI: 1.84 to 2.66), higher log

SPC concentration (OR 1⁄4 1.29, 95% CI: 1.17 to 1.43), and

higher log E. coli concentration (OR 1⁄4 1.89, 95% CI: 1.57

to 2.29) (Table 4).

In ground beef, non–Salmonella Dublin serotypes were

significantly associated with SPC detection (OR 1⁄4 1.66,

95% CI: 1.01 to 2.84), total coliform detection (OR 1⁄4 2.97,

95% CI: 2.01 to 4.35), and E. coli detection (OR 1⁄4 3.73,

95% CI: 2.22 to 6.00). Univariable analysis found

significant non–Salmonella Dublin associations with increased log SPCs (OR 1⁄4 1.60, 95% CI: 1.34 to 1.91), log

total coliforms (OR 1⁄4 1.95, 95% CI: 1.56 to 2.44), and log

E. coli concentration (OR 1⁄4 2.27, 95% CI: 1.67 to 3.09). In

the multivariable model, non–Salmonella Dublin serotypes

were significantly associated with larger sample size (OR 1⁄4

2.12, 95% CI: 1.41 to 3.19), higher log SPC concentration

(OR 1⁄4 1.27, 95% CI: 1.01 to 1.60), and season.

This study found consistent, positive associations

between indicator detection and Salmonella presence and

increased odds of Salmonella presence with increased

concentration of log SPCs and log E. coli in boneless beef

and increased log SPC concentration in ground beef. These

findings support the continued use of microbial indicators as

qualitative controls for boneless and ground beef production. Good hygiene and process control indicated by lower

indicator microorganism concentrations appear to generally

reduce the risk of Salmonella presence. However, the

effectiveness of these measures was markedly reduced by

the inclusion of lots contaminated by Salmonella Dublin.

Generally, increased concentrations of log SPCs and log E.

coli were associated with Salmonella overall and other

Salmonella serotypes, but not with Salmonella Dublin

presence. The differential associations of Salmonella Dublin

presence relative to the other serotypes with microbial

indicator concentration reduces their effectiveness to assess

the risk of Salmonella presence. The results of this study

provide new information that AMS-approved beef vendors

for the National School Lunch Program can use to better

understand testing results and further refine their food safety

systems.

Compared with other serotypes, Salmonella Dublin was

more likely in lots with nondetectable indicators and with

lower log SPC concentrations. Salmonella Dublin also had a

markedly different seasonality from that of other Salmonella serotypes. Salmonella Dublin was more strongly

associated with spring compared with the other seasons.

Spring had the highest proportion of Salmonella Dublin

isolates in both boneless and ground beef. Furthermore,

during spring, Salmonella Dublin prevalence tended to be

significantly higher than during the other seasons.

Other studies have attributed higher Salmonella

prevalence during the summer and fall to other environmental variables, including temperature, moisture, and

latitude, that were not measured in this study (10, 22).

Although herd-level factors were not examined in this

study, it is possible that the differential seasonal findings

may also correlate with underlying cattle management

practices within the source population. Pregnancy and

calving-related stress in the spring may weaken a cow’s

immune system, thereby increasing the likelihood of latent

infection reactivation and increased shedding by carriers

(19). This reactivation and shedding of Salmonella Dublin

on farms may, in turn, contribute to higher Salmonella

Dublin prevalence in meat processing operations. In months

subsequent to spring, increased metabolic demand because

of milk production may also affect Salmonella prevalence in

herds and in meat processing. The observation that the

proportion of Salmonella isolates found to be Salmonella

Dublin decreased from 52.8% in boneless beef to 20.5% in

ground beef was a key finding. Decreased Salmonella

Dublin prevalence in ground beef may be because of

variations in the growth phase among Salmonella serotypes,

along with inherent variation in storage temperature and

nutrient availability within lots and sublots during production and type of meat (17, 23, 28, 29). Salmonella serotypes

other than Salmonella Dublin may be outcompeting

Salmonella Dublin during the period between boneless beef

testing and ground beef testing on the basis of these and

similar factors. These findings suggest Salmonella Dublin

may be less fit than other Salmonella serotypes in boneless

and ground beef containing higher microbial indicator

concentrations. It is also possible that ground beef is a less

hospitable environment for Salmonella Dublin for unknown

reasons.

Future research should examine the risk of Salmonella

contamination compared with various indicator microorganism concentration levels and combinations of those

levels. It may be that certain combinations of indicator

concentrations signal higher risk than other combinations.

For example, a boneless beef lot containing higher

concentrations of both log SPCs and E. coli (e.g., ,104

CFU g 1) may be more likely to contain Salmonella than a

lot with relatively fewer SPCs, E. coli, or both. This

research should be informed by the knowledge that risk may

also vary by Salmonella serotype and by serotype

prevalence in source cattle. Once identified, indicator

concentration combinations that represent increased risk

of Salmonella presence could serve as triggers for additional

preventative actions aimed at reducing indicator concentra-
J. Food Prot., Vol. 83, No. 4 635

tion and thereby reducing risk of Salmonella presence in

distributed beef products.

ACKNOWLEDGMENTS

The authors thank the boneless and ground beef vendors described in

this report, the staff at AMS-designated laboratories and at the FSIS

Eastern Laboratory for sample analysis, and Bruce Alexander, Jeff Bender,

and Scott Wells for technical support. Support for this project was

provided by the Pew Charitable Trusts.

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