AEM Accepted Manuscript Posted Online 24 April 2015 Appl. Environ. Microbiol. doi:10.1128/AEM.04086-14 Copyright © 2015, American Society for Microbiology. All Rights Reserved.
1
Distribution and Characterization of Salmonella enterica Isolates from Irrigation Ponds
2
in the Southeastern U.S.A.
3 4
Zhiyao Luo1, Ganyu Gu2, Amber Ginn1, Mihai C. Giurcanu3, Paige Adams4, George
5
Vellidis4, Ariena H. C. van Bruggen2, Michelle D. Danyluk 1,5 and Anita C. Wright1*
6 7
1
8
32611, United States; 2 Emerging Pathogens Institute and Department of Plant Pathology,
9
University of Florida, Gainesville, FL 32611; 3 Department of Statistics, University of
10
Florida, Gainesville, FL, 32611; 4 Biological & Agricultural Engineering Department,
11
University of Georgia, Tifton, GA 31793; 5 Citrus Research and Education Center, University
12
of Florida, Lake Alfred, FL 33850
Department of Food Science and Human Nutrition, University of Florida, Gainesville, FL
13 14
* Corresponding author: Dr. Anita C. Wright, Department of Food Science and Human
15
Nutrition, Bldg 475 Newell Dr., P.O. Box 11030, University of Florida, Gainesville, FL
16
32611, United States; [email protected]; 352-392-1991 x 311
17 18 19
Running title: Distribution, diversity, and characterization of Salmonella from irrigation ponds
20 21
Key words: Salmonella, water, sediment, distribution, antibiotic resistance test, rep-PCR,
22
irrigation ponds
1
23
ABSTRACT
24
Irrigation water has been implicated as a likely source of produce contamination by
25
Salmonella enterica. Therefore, the distribution of S. enterica was surveyed monthly in
26
irrigation ponds (n=10) located within a prime agricultural region in Southern Georgia and
27
Northern Florida. All ponds and 28.2% of all samples (n=635) were positive for Salmonella
28
with an overall geometric mean concentration (0.26 MPN/L) that was relatively low
29
compared to prior reports for rivers in this region. Salmonella peaks were seasonal; levels
30
correlated with increased temperature and rainfall (p95% similarity. Genotypes did not partition by pond,
37
season, or sample type. Genetic similarity to known serotypes indicated Hadar, Montevideo,
38
and Newport as the most prevalent. All ponds achieved the current safety standards for
39
generic Escherichia coli in agricultural water, and regression modeling showed E. coli levels
40
were a significant predictor for the probability of Salmonella occurrence. However, persistent
41
populations of Salmonella were widely distributed in irrigation ponds, and associated risks
42
for produce contamination and subsequent human exposure are unknown, supporting
43
continued surveillance of this pathogen in agricultural settings.
44
2
45
INTRODUCTION
46
Salmonella enterica is the leading cause of bacterial food-borne illnesses and accounts
47
for approximately 42,000 cases of infections annually in the U.S. (1). Traditionally,
48
salmonellosis has been considered a zoonotic disease due to its frequent association with
49
poultry and eggs (2, 3); however, recent outbreaks are increasingly attributed to fresh fruits
50
and produce (4), and the number of disease cases per outbreak is sometimes greater for
51
vegetables than for other food products (5). These outbreaks confirm that environmental
52
transmission of Salmonella from produce can lead to human illness (6, 7). Recent
53
investigations have focused on irrigation water as a potential environmental source of
54
Salmonella for pre-harvest contamination of produce (8-10, 11, 12). Furthermore, the static
55
nature of some irrigation ponds may sustain persistent populations of Salmonella (13, 14).
56
Assessment of the microbial quality of agricultural water is now required under the
57
proposed Produce Safety Rule (PSR) of the Food Safety Modernization Act (FSMA) in order
58
to ensure the safety of fresh produce (15). The recommended microbial standards for the
59
quality of agricultural water that comes in direct contact with pre-harvest produce (other than
60
sprouts) requires a threshold of generic Escherichia coli to be
1
Distribution and Characterization of Salmonella enterica Isolates from Irrigation Ponds
2
in the Southeastern U.S.A.
3 4
Zhiyao Luo1, Ganyu Gu2, Amber Ginn1, Mihai C. Giurcanu3, Paige Adams4, George
5
Vellidis4, Ariena H. C. van Bruggen2, Michelle D. Danyluk 1,5 and Anita C. Wright1*
6 7
1
8
32611, United States; 2 Emerging Pathogens Institute and Department of Plant Pathology,
9
University of Florida, Gainesville, FL 32611; 3 Department of Statistics, University of
10
Florida, Gainesville, FL, 32611; 4 Biological & Agricultural Engineering Department,
11
University of Georgia, Tifton, GA 31793; 5 Citrus Research and Education Center, University
12
of Florida, Lake Alfred, FL 33850
Department of Food Science and Human Nutrition, University of Florida, Gainesville, FL
13 14
* Corresponding author: Dr. Anita C. Wright, Department of Food Science and Human
15
Nutrition, Bldg 475 Newell Dr., P.O. Box 11030, University of Florida, Gainesville, FL
16
32611, United States; [email protected]; 352-392-1991 x 311
17 18 19
Running title: Distribution, diversity, and characterization of Salmonella from irrigation ponds
20 21
Key words: Salmonella, water, sediment, distribution, antibiotic resistance test, rep-PCR,
22
irrigation ponds
1
23
ABSTRACT
24
Irrigation water has been implicated as a likely source of produce contamination by
25
Salmonella enterica. Therefore, the distribution of S. enterica was surveyed monthly in
26
irrigation ponds (n=10) located within a prime agricultural region in Southern Georgia and
27
Northern Florida. All ponds and 28.2% of all samples (n=635) were positive for Salmonella
28
with an overall geometric mean concentration (0.26 MPN/L) that was relatively low
29
compared to prior reports for rivers in this region. Salmonella peaks were seasonal; levels
30
correlated with increased temperature and rainfall (p95% similarity. Genotypes did not partition by pond,
37
season, or sample type. Genetic similarity to known serotypes indicated Hadar, Montevideo,
38
and Newport as the most prevalent. All ponds achieved the current safety standards for
39
generic Escherichia coli in agricultural water, and regression modeling showed E. coli levels
40
were a significant predictor for the probability of Salmonella occurrence. However, persistent
41
populations of Salmonella were widely distributed in irrigation ponds, and associated risks
42
for produce contamination and subsequent human exposure are unknown, supporting
43
continued surveillance of this pathogen in agricultural settings.
44
2
45
INTRODUCTION
46
Salmonella enterica is the leading cause of bacterial food-borne illnesses and accounts
47
for approximately 42,000 cases of infections annually in the U.S. (1). Traditionally,
48
salmonellosis has been considered a zoonotic disease due to its frequent association with
49
poultry and eggs (2, 3); however, recent outbreaks are increasingly attributed to fresh fruits
50
and produce (4), and the number of disease cases per outbreak is sometimes greater for
51
vegetables than for other food products (5). These outbreaks confirm that environmental
52
transmission of Salmonella from produce can lead to human illness (6, 7). Recent
53
investigations have focused on irrigation water as a potential environmental source of
54
Salmonella for pre-harvest contamination of produce (8-10, 11, 12). Furthermore, the static
55
nature of some irrigation ponds may sustain persistent populations of Salmonella (13, 14).
56
Assessment of the microbial quality of agricultural water is now required under the
57
proposed Produce Safety Rule (PSR) of the Food Safety Modernization Act (FSMA) in order
58
to ensure the safety of fresh produce (15). The recommended microbial standards for the
59
quality of agricultural water that comes in direct contact with pre-harvest produce (other than
60
sprouts) requires a threshold of generic Escherichia coli to be
