ENUMERATION OF TOTAL AIRBORNE BACTERIA, YEAST AND

141 Pages·2004·0.5 MB·English

by  Gabriel H. Cosenza

ENUMERATION OF TOTAL AIRBORNE BACTERIA, YEAST AND MOLD
CONTAMINANTS AND IDENTIFICATION OF Escherichia coli O157:H7, Listeria
Spp., Salmonella Spp., AND Staphylococcus Spp. IN A BEEF AND PORK
SLAUGHTER FACILITY

By

GABRIEL HUMBERTO COSENZA SUTTON

A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL
OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT
OF THE REQUIREMENTS FOR THE DEGREE OF
DOCTOR OF PHILOSOPHY

UNIVERSITY OF FLORIDA

2004
ACKNOWLEDGMENTS
The author is sincerely grateful to Dr. S. K. Williams, associate professor and
supervisory committee chairperson, for her superb guidance and supervision in
conducting this study and manuscript preparation. He also extends his gratitude to the
other committee members, Dr. Dwain Johnson, Dr. Ronald H. Schmidt, Dr. David P.
Chynoweth and Dr. Murat O. Balaban, for their collaboration and useful
recommendations during this study. Special thanks are extended to the Institute of Food
and Agricultural Sciences of the University of Florida for sponsoring the author
throughout the program. Great appreciation and love are expressed to Adrienne and
Humberto Cosenza, the author’s parents, and to his girlfriend Candice L. Lloyd for their
support throughout the graduate program. The author wishes to thank Larry Eubanks,
Byron Davis, Tommy Estevez, Doris Sartain, Frank Robbins, Noufoh Djeri and fellow
graduate students for their friendly support and assistance. Overall the author would like
to thank God for His inspiration, love and caring.

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TABLE OF CONTENTS
page

ACKNOWLEDGMENTS..................................................................................................ii
TABLE OF CONTENTS...................................................................................................iii
LIST OF TABLES.............................................................................................................vi
ABSTRACT.....................................................................................................................viii
CHAPTER

1 INTRODUCTION........................................................................................................1
2 LITERATURE REVIEW.............................................................................................5
Bioaerosols...................................................................................................................5
Bioaerosol Sampling Methods......................................................................................7
Sedimentation........................................................................................................8
Impaction...............................................................................................................9
Impingement........................................................................................................10
Filtration..............................................................................................................11
Centrifugation......................................................................................................12
Electrostatic Precipitation....................................................................................12
Thermal Precipitation..........................................................................................13
Beef and Pork Slaughter Process................................................................................13
Stunning or Immobilization.................................................................................14
Bleeding...............................................................................................................14
Scalding/Dehairing..............................................................................................14
Evisceration.........................................................................................................15
Splitting and Chilling..........................................................................................15
Bioaerosols in the Meat Industry................................................................................16
Bioaerosols in the Dairy Industry...............................................................................20
Microbiological Analysis of Bioaerosols...................................................................23
Environmental and Beef and Pork Associated Bacteria.............................................27
Gram Negative Bacteria.............................................................................................29
Enterobacteriaceae.....................................................................................................29
Escherichia coli...................................................................................................30
Salmonella spp.....................................................................................................34
Shigella spp.........................................................................................................37
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Klebsiella spp......................................................................................................38
Serratia spp.........................................................................................................38
Enterobacter spp.................................................................................................39
Citrobacter spp....................................................................................................39
Pantoea spp.........................................................................................................40
Morganella spp....................................................................................................40
Kluyvera spp........................................................................................................41
Cedecea spp.........................................................................................................41
Leclercia spp.......................................................................................................41
Non Enterobacteriaceae.............................................................................................42
Acinetobacter spp................................................................................................42
Moraxella spp......................................................................................................42
Chryseomonas spp...............................................................................................42
Flavimonas spp....................................................................................................43
Pseudomonas spp................................................................................................43
Stenotrophomonas spp.........................................................................................43
Chryseobacterium indologenes...........................................................................44
Gram Positive Bacteria...............................................................................................44
Listeria spp..........................................................................................................44
Staphylococcus spp..............................................................................................48
Bacillus spp.........................................................................................................51
Brevibacterium spp..............................................................................................52
Cellulomonas spp................................................................................................52
Brochothrix spp...................................................................................................52
Microbacterium spp.............................................................................................53
Micrococcus spp..................................................................................................53

3 MATERIALS AND METHODS...............................................................................55
Air Sampling System..................................................................................................55
Air Sample and Carcass Swab Collection..................................................................57
Preliminary Studies.............................................................................................57
Carcass Sample Collection..................................................................................58
Air and Carcass Microbiological Sample Analysis....................................................59
Bacterial Identification of Air and Carcass Samples..................................................62
Microbiological Identification....................................................................................63
Statistical Analyses.....................................................................................................68

4 RESULTS AND DISCUSSION.................................................................................69
Preliminary Work: Air Sampling Time Determination..............................................69
Microbiology of Bioaerosols Collected During Pork Slaughters...............................70
Total Airborne Bacteria.......................................................................................70
Total Airborne Yeast and Mold...........................................................................72
Isolation of Staphylococcus species....................................................................75
Isolation of Listeria species.................................................................................77
Isolation of Escherichia coli................................................................................80
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Isolation of Salmonella species...........................................................................83
Identification of Airborne Bacteria Collected from Pork Slaughters..................84
Carcass Swabs Collected During Pork Slaughters..............................................87
Microbiology of Bioaerosols Collected During Beef Slaughters...............................89
Total Airborne Bacteria.......................................................................................89
Total Airborne Yeast and Mold...........................................................................91
Isolation of Staphylococcus species....................................................................93
Isolation of Listeria species.................................................................................95
Isolation of Escherichia coli................................................................................97
Isolation of Salmonella Species...........................................................................99
Identification of Airborne Bacteria Collected from Beef Slaughters................101
Carcass Swabs Collected During Beef Slaughters............................................103
Comparison of Bioaerosols Collected from Pork and Beef Slaughters....................105

5 SUMMARY AND CONCLUSIONS.......................................................................108
APPENDIX ORGANISMS IDENTIFIED IN AIR AND CARCASS SAMPLES......112
LIST OF REFERENCES.................................................................................................127
BIOGRAPHICAL SKETCH...........................................................................................132
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LIST OF TABLES
Table page

1 Air sampling areas for beef and pork slaughtering processes..................................55
2 Sampling time determination for air sample collection during a beef slaughter......69
3 Mean total airborne bacteria collected on tryptic soy agar plates in eight areas
during three pork slaughters.....................................................................................72
4 Mean total airborne yeast and mold collected on potato dextrose agar plates in
eight areas during three pork slaughters...................................................................74
5 Mean airborne bacteria collected on mannitol salt agar plates in eight areas
during three pork slaughters.....................................................................................76
6 Airborne bacteria isolated from mannitol salt agar plates collected from three
pork slaughters.........................................................................................................77
7 Mean airborne bacteria collected on modified oxford agar plates in eight areas
during three pork slaughters.....................................................................................78
8 Airborne bacteria isolated from modified oxford agar plates collected from three
pork slaughters.........................................................................................................79
9 Mean airborne bacteria collected on violet red bile agar plates in eight areas
during three pork slaughters.....................................................................................81
10 Airborne bacteria isolated from violet red bile agar plates collected from three
pork slaughters.........................................................................................................82
11 Mean airborne bacteria collected on xylose-lysine-tergitol 4 agar plates in eight
areas during three pork slaughters............................................................................83
12 Airborne bacteria isolated from xylose-lysine-tergitol 4 agar plates collected
from three pork slaughters........................................................................................84
13 Airborne bacteria isolated before and during three pork slaughters........................86
14 Bacteria isolated from pork carcasses held at 00C for 24 hours...............................88
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15 Mean total airborne bacteria collected on tryptic soy agar plates in eight areas
during three beef slaughters.....................................................................................90
16 Mean total airborne yeast and mold collected on potato dextrose agar plates in
eight areas during three beef slaughters...................................................................92
17 Mean airborne bacteria collected on mannitol salt agar plates in eight areas
during three beef slaughters.....................................................................................94
18 Airborne bacteria isolated from mannitol salt agar plates collected from three
beef slaughters..........................................................................................................95
19 Mean airborne bacteria collected on modified oxford agar plates in eight areas
during three beef slaughters.....................................................................................96
20 Airborne bacteria isolated from modified oxford agar plates collected from three
beef slaughters..........................................................................................................97
21 Mean airborne bacteria collected on violet red bile agar plates in eight areas
during three beef slaughters.....................................................................................97
22 Airborne bacteria isolated from violet red bile agar plates collected from three
beef slaughters..........................................................................................................98
23 Mean airborne bacteria collected on xylose-lysine-tergitol 4 agar plates in eight
areas during three beef slaughters..........................................................................100
24 Airborne bacteria isolated from xylose-lysine-tergitol 4 agar plates collected
from beef slaughters...............................................................................................100
25 Airborne bacteria isolated before and during three beef slaughters.......................101
26 Bacteria isolated from beef carcasses held at 00C for 24 hours.............................104

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Abstract of Dissertation Presented to the Graduate School
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosophy

ENUMERATION OF TOTAL AIRBORNE BACTERIA, YEAST AND MOLD
CONTAMINANTS AND IDENTIFICATION OF Escherichia coli O157:H7, Listeria
Spp., Salmonella Spp., AND Staphylococcus Spp. IN A BEEF AND PORK
SLAUGHTER FACILITY
By
Gabriel Humberto Cosenza Sutton

August 2004

Chair: Sally K. Williams
Major Department: Animal Sciences
Environmental air monitoring programs can be employed to reduce unsanitary
conditions in animal slaughterhouses due to suspended bacterial particles in the air. The
main objectives of this study were to enumerate total airborne bacteria and yeast and
mold contaminants and determine the presence of Escherichia coli O157:H7, Listeria
spp., Salmonella spp., and Staphylococcus spp. in bioaerosols generated in a slaughter
facility and on pork and beef carcasses. Air samples were taken before and during three
separate pork and beef slaughter processes at the bleeding area, hide removal or dehairing
area, back splitting area and holding cooler using an Andersen N6 single stage impactor.
Pork and beef carcass surface bacterial swabs were collected from five different carcass
sides which had been held in the holding cooler at 00C for 12 hours.
Total airborne bacterial (TAB) counts (log CFU/m3 of air) were generally higher
during slaughtering than before slaughtering. TAB counts were greater than three logs
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during slaughtering and less than three logs before slaughtering. The holding cooler had
TAB counts less than or equal to two logs. Similar recovery rates for Staphylococcus,
Escherichia and Salmonella species were obtained through direct air and enriched air
microbiological sample analysis methods. Most of the Gram negative airborne bacteria
isolated during slaughtering were from the Enterobacteriaceae and Pseudomonadaceae
family. The predominant Gram positive airborne bacteria isolated during slaughtering
were Staphylococcus, Microbacterium, Bacillus and Micrococcus species. Potentially
pathogenic Staphylococcus aureus, Escherichia coli and Salmonella spp. were isolated
from bioaerosols generated during slaughtering and from pork and beef carcasses.
Neither Listeria spp. nor Escherichia coli O157:H7 were insolated from air samples or
pork and beef carcasses.
The isolation of various microorganisms, including Staphylococcus, Escherichia,
and Salmonella spp., from air samples and carcass swabs support the theory that
bioaerosols transport bacteria and contribute to contamination of pork and beef carcasses.
The determination of the levels and types of airborne bacterial contaminants present in a
small scale slaughter facility has various implications. The effectiveness of a plant’s
sanitation program can be evaluated and the sources of airborne contamination can be
determined allowing for increased food safety.
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CHAPTER 1
INTRODUCTION
In the past it was thought that food products were contaminated when they came in
contact with contaminated surfaces, but now it is known that additional product
contamination occurs from contact with airborne bacteria. Unsanitary environmental
conditions in food processing plants can occur due to suspended bacterial particles in the
air. These biological particles are microscopic, with a diameter of 0.5 to 50 µm and are
suspended in the air as an aerosol. Airborne contaminants are also known as bioaerosols
and include bacteria, fungi, viruses and pollen. These may be present in the air as solid
(dust) or as liquid (condensation and water). An aerosol is a two-phase system of
gaseous phase (air) and particulate matter (dust, pathogens), thus making an important
bacterial vehicle. Pathogenic bacteria attach to dust particles and condensation, and
travel around the processing facility. This contaminated air comes in contact with food
products, containers, equipment and other food contact surfaces during processing.
According to the Food and Drug Administration (FDA), the food industry must reduce
product contamination by reducing airborne microorganisms (11, 18).
Airborne contaminants cause human illness due to ingestion of contaminated foods
and also reduce product shelf life resulting in an economic loss (18). Swabbing of
equipment is typically used to determine the sanitation level of food processing plants.
This method does not always provide an effective enumeration of airborne contaminants.
Air sampling is more effective because it collects aerosols settled on equipment and food
contact surfaces. Through air sampling, food processing facilities can identify airborne
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