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Growth of Psychrotolerant Pseudomonads and Achromobacter on Chicken Skin
Growth of Psychrotolerant Pseudomonads and Achromobacter on Chicken Skin D. S. CLARK Division of Biology, National Research Council, Ottawa, Ontario, Canada (Received for publication March 6, 1968) INTRODUCTION
ant bacteria growing on exposed surfaces (skin, muscle and gut-cavity lining), yet definite information on many of the physical and chemical factors that affect rates of growth is scarce. Results of published work are often conflicting and difficult to interpret because experiments were made with birds contaminated naturally during processing or handling and thus under conditions not controlled as to type and amount of inoculum. Comparative testing with controlled inoculation is required to determine the relative growth rates of the principal spoilage organisms and the extent to which these rates are affected by processing and storage conditions. Work has been undertaken, therefore, to study factors which affect the growth of pseudomonads and achromobacter (the two principal groups of bacteria responsible for low-temperature spoilage of poultry (Ayres et al., 1950; Thornley et al., 1960: Walker and Ayres, 1956)) on the surface of chicken skin, muscle, and gut-cavity lining. This paper outlines methods developed for preparing and inoculating skin and describes the effect of several factors (birdto-bird variability, age of bird, location of skin on carcass, freezing and thawing of skin, and methods of scalding) on the growth of the two groups of organisms on the skin surface at 0°C. Skin was chosen as the initial substrate for study because published work (Lockhead and Landerkin, N. R. C. No. 10,055.
METHODS
Skin was obtained from broiler-type chickens (Ottawa Meat Control Strain) raised and processed in the laboratory. After slaughter (by cutting the jugular vein and carotid arteries), the chickens were scalded (at 59°C, unless otherwise stated), plucked by hand and rubbed free of loose pieces of the epidermal layer of the skin and parts of feathers with a sterile damp cloth. The skin was removed from the carcass before evisceration, washed for 15 seconds in a hot (59°C.) detergent solution (0.05% Sparkleen), rinsed thoroughly in cool sterile water, and cut into pieces or samples approximately 9 cm. square. Each piece was stretched, outside surface up, onto a circular stainless steel disk (diam. 7.6 cm.) with hooks around the outside edge to stretch the piece enough to smooth out all visible folds. The excess skin was trimmed off. The samples were held in sterile Petri dishes in a saturated atmosphere at 0°C. until inoculated (within 24 hrs). Preliminary tests showed that the skin samples were essentially free of psychrophiles; counts on uninoculated samples incubated at 0°C. for 10 days never exceeded 50 per sq. cm. Two inocula were used in all of the tests; one was composed of a mixture of 10
1575
Downloaded from http://ps.oxfordjournals.org/ at California Institute of Technology on June 20, 2015
of fresh refrigerated poultry S POILAGE is ordinarily caused by psychrotoler-
1935; Ziegler, 1954) has indicated that microbially-induced changes leading to offodour in dressed eviscerated poultry occur first on the surface of the skin; 0°C. was selected because a large percentage of unfrozen poultry (about 95% in Canada) is stored and shipped in ice.
1576
D. S. CLARK
location, 4 skin samples, one from each of 4 locations (back, breast, and upper and lower leg), were inoculated simultaneously in each test. The test was repeated 10 times for each type of inoculum. Studies to determine the effect of freezing and scalding on growth rate of Pseudomonas and Achromobacter were made with skin from the breast and leg areas of 10-weekold chickens. In freezing tests, growth rates on fresh skin were compared with those on skin that had been stored frozen for 1 month and then thawed at room tempertaure before being stretched and inoculated. Two methods of freezing were used; in one, skins sealed separately in cryovac bags were immersed for 5 min. in an alcohol-dry ice bath (-72°C.) and stored at - 1 0 0 ° C ; in the other, skin samples similarly packaged were frozen and stored in a room held at — 10°C. The growth rate tests were repeated 6 times for both types of bacteria using skin scalded at 59°C; in each test 2 fresh samples and 2 samples each of fast-frozen ( — 72°C.) and slow-frozen skin ( —10°C.) were inoculated simultaneously. To test the effect of scalding, the growth rate on unscalded skin was compared with the rates on skin scalded for 2 min. in water at 54°C. and for 1 min. in water at 59°C. The 54°C. scald (used commercially by some small producers) permitted removal during rub-down of only a portion of the yellowish epidermal layer of skin, particularly that on the tops of the mounds around the feather follicles, while TABLE 1.—Effect of scalding on growth of Pseudomonas and Achromobacter Counts per sq. cm. of skinXlO 6 after 6 days at 0°C*
Pseudomonas Achromobacter
TVT .
Scalded
Scalded
1.3 2.5
2.0 1.2
2.5 0.9
* Initial inoculation, 2.5X10 4 per sq. cm.
Downloaded from http://ps.oxfordjournals.org/ at California Institute of Technology on June 20, 2015
strains of Pseudomonas and the other of 10 strains of Achromobacter. The organisms were isolated from the surface of processed chickens in poultry plants and classified according to Thornley et al. (1960). All grew well at 0°C. Cells of each strain were harvested from 24-hr-old SM agar (Difco) cultures, suspended in a 0.1% aqueous peptone solution (Straka and Stokes, 1957) having an ionic strength of 0.5 (Torloni, 1962a), adjusted photometrically to 50% T at 490 m(/.., mixed in equal volumes in a nebulizer (Torloni, 1962b) and sprayed uniformly (about 25,000 cells per sq. cm. in all tests) over the surface of skin samples in an inoculation chamber (Clark, 1963). After inoculation the skin samples were incubated in a saturated atmosphere at 0°C. and sampled after 6 days. In one test, to determine the growth curves of the two types of organisms on scalded skin, counts on inoculated breast samples were assessed every two or three days during a 13 day incubation period. In sampling, three 2 sq. cm. areas on each skin sample were washed by the spray-gun technique (Clark, 1965a, b) and duplicate counts for each washing were determined by surface plating (Clark, 1967) on SM agar. The agar plates were incubated at 20°C. Analyses of variance were made on the counts. The effects of bird-to-bird variability and skin age and location on bacterial growth were studied with skin (scalded at 59°C.) from 8- and 16-week-old chickens. In bird-to-bird variability tests, each inoculation was made with six samples of breast skin, 1 each from six different 8-week-old birds. Six such tests were made for each group of bacteria, using skin samples from a different set of birds every second time. Tests for determination of effect of age were also repeated 6 times for each inocula, using 3 skin samples from 8-week-old birds and 3 skin samples from 16-week-old birds in each inoculation. To study the effect of
1577
PSEUDOMONADS AND ACHROMOBACTER ON SKIN
the 59°C. scald (widely used commercially) allowed complete removal of the outer layer of skin during rub-down. Unscalded skin, obtained from birds plucked "dry" was washed for 15 sec. in detergent solution at 25°C. and skin scalded at 54°C. was washed in detergent solution at 54°C. The growth tests were repeated 10 times; in each test 2 skin samples for each of the three conditions were inoculated simultaneously.
in
1
8
^ *,• ^^*^
y^\^^'
-
PSEUDOMONAS -—^
S 7 fe 6
,/Z-
»^^^ ACHROMOBACTER
^ ^ ^ ^ ^ ^
I
2
3
i _1 1 1 1 1 1 1_ 4 5 6 7 8 9 10 II 12 13 INCUBATION TIME, DAYS
Table 1 shows that scalding had a sig- FIG. 1. Growth of Pseudomonas and Achromobacter nificant effect on the growth rate of both on skin scalded at 59° C. types of organisms. Pseudomonas grew best on scalded skin, particularly skin scalded calculated from analysis of variance of the at 59°C, whereas Achromobacter grew results, were not significant for all factors best on unscalded skin; skin scalded at the and their interactions. The corresponding lower temperature (54°C.) gave intermedi- values for Achromobacter were: average ate growth rates in both cases. These count, about 900,000 cells/sq. cm.; and differences appear to be the result of rela- standard deviation between 370-540. tive preferences for the outermost and un- Again, the F values were not significant. derlying layers of skin as substrates, since Figure 1 shows the relative growth rates of the intermediate growth rates were ob- the two groups of organisms on skin tained when part of the epidermal layer scalded at 59°C. The "mean generation was removed (54°C. scald). When pseudo- time" (Elliott and Michener, 1965) for the monads are the predominant spoilage orga- Pseudomonas was about 12 hr while for the nisms on fresh commercially-processed Achromobacter it was about 14 hr. These poultry, as they frequently are (Ayres et generation times are approximately the al, 1950; Thornley et al., 1960; Nagel et same as those reported for psychrotolerant al., 1960), the shelf-life for birds scalded at bacteria on the skin of commercially pro59°C. should be significantly shorter than cessed chickens contaminated naturally that for birds scalded at 54°C. This would during processing (Dawson et al., 1963; seem to explain the longer shelf-life re- Hall and Spencer, 1963). ported for poultry scalded at the lower The study showed that any location on temperature (Ziegler and Stadelman, the skin of the bird is suitable for sampling 1955). in tests to determine the bacteriological Unlike scalding, the other factors exa- quality of whole refrigerated poultry, since mined in comparative tests (bird-to-bird psychrotolerant spoilage bacteria grow variability, age, location and freezing and equally well anywhere on the skin surface. thawing) had no significant effect on the The study also showed that in comparative growth rate of either type of organism. The studies of factors affecting bacterial growth average count after 6 days in all tests with on chicken skin, skin from different birds Pseudomonas was about 2,000,000 cells/sq. and from birds of different ages (at least cm.; the standard deviation in each case between 8 and 16 weeks old) can be used was between 270 and 340 and the F values, indiscriminately. In addition the skin can
Downloaded from http://ps.oxfordjournals.org/ at California Institute of Technology on June 20, 2015
RESULTS AND DISCUSSION
E dIO \ 9
1578
D. S. CLARK
also be frozen for convenience in storing a supply for continued or periodic work. ACKNOWLEDGMENTS
The author wishes to thank N. U. Cholette for skilled technical assistance. REFERENCES
NEWS AND NOTES (continued from page 1574) Forsythe, Henningsen Foods, Inc., Springfield, ELANCO NOTES Missouri. The U.S. Participation Committee is a During the annual meeting of the Poultry and Egg National Board in Chicago, Elanco Products function of the U.S.A. Branch of the World's Poultry Science Association. Co., Indianapolis, Indiana, a division of Eli Lilly The following Committee Chairmen were Company, was elected to membership in the Good Egg Club "for outstanding cooperation and general named: Science—Dr. Hans Lineweaver, Western Regional Research Laboratory, U.S. Department excellence in promotion of eggs." of Agriculture, Albany, California; Papers SelecCONGRESS COMMITTEES tion—Dr. R. N. Shoffner, University of MinneThe U.S. Participation Committee for the 14th sota, St. Paul, Minnesota; Travel Grants—Dr. World's Poultry Congress, which is scheduled for W. J. Stadelman, Purdue University, Lafayette, September, 1970, in Spain, was activated at a Indiana; Commercial Participation—Richard Ammeeting in Chicago, Illinois, on August 23rd, under nion, N.E.P.P. Co., Trenton, New Jersey; Memthe direction of its Chairman, Dr. Richard H. bership and Promotion—Gene Shephard, Institute (continued on page 1611)
Downloaded from http://ps.oxfordjournals.org/ at California Institute of Technology on June 20, 2015
Ayres, J. C , W. S. Ogilvy and G. F. Stewart, 1950. Post mortem changes in stored meats. I. Microorganisms associated with development of slime on eviscerated cut-up poultry. Food Technol. 4 : 199-205. Clark, D. S., 1963. Uniform inoculation of nutrient surfaces. Biotech. Bioeng. 5 : 123-129. Clark, D. S., 1965a. Method of estimating the bacterial population on surfaces. Can. J. Microbiol. 11:407-413. Clark, D. S., 1965b. Improvement of spray gun method of estimating bacterial populations on surfaces. Can. J. Microbiol. 11: 1021-1022. Clark, D. S., 1967. Comparison of pour and surface plate methods for determination of bacterial counts. Can. J. Microbiol. 13: 1409-1412. Dawson, L. E., W. L. Mallmann, D. G. Bigbee, R. Walker and M. E. Zabik, 1963. Influence of surface pasteurization and chlortetracycline on bacterial incidence on fryers. Food Technol. 17: 100-103. Elliott, R. P., and H. D. Michener, 1965. Factors affecting the growth of psychrophilic microorganisms in foods. Technical Bulletin No. 1320, Agricultural Research Service, U.S. Dept. Agriculture: p. 2.
Hall, K. N., and J. V. Spencer, 1963. The effect of ethanol on shelf-life and flavor of chicken meat. Poultry Sci. 4 3 : 573-576. Lochhead, A. G., and G. B. Landerkin, 1935. Bacteriological studies of dressed poultry. I. Preliminary investigations of bacterial action at chill temperatures. Sci. Agri. 15: 765-770. Nagel, C. W„ K. L. Simpson, H. Ng, R. H. Vaughn and G. F. Stewart, 1960. Microorganisms associated with spoilage of refrigerated poultry. Food Technol. 14: 21-23. Straka, R. P., and J. L. Stokes, 1957. Rapid destruction of bacteria in commonly used diluents and its elimination. Appl. Microbiol. 5 : 21-25. Thronley, M. J., M. Ingram and E. M. Barnes, 1960. The effects of antibiotics and irradiation on Pseudomonas-Achromobacter flora of chilled poultry. J. Appl. Bact. 23: 487-498. Torloni, M., 1962a. Influence of the ionic concentration of nebulized suspensions on stability of microbial aerosols. Biotechnol. Bioeng. 4 : 341-344. Torloni, M., 1962b. A simple and versatile nebulizer. Appl. Microbiol. 10: 6-8. Walker, H. W., and J. C. Ayres, 1956. Incidence and kind of microorganisms associated with commercially dressed poultry. Appl. Microbiol. 4:345-350. Ziegler, F., J. V. Spencer and W. J. Stadleman, 1954. A rapid method for determining spoilage in fresh poultry meat. Poultry Sci. 3 3 : 12531255. Ziegler, F., and W. J. Stadelman, 1955. The effect of different scald water temperatures on shelflife of fresh non-frozen fryers. Poultry Sci. 34: 237-238.
ant bacteria growing on exposed surfaces (skin, muscle and gut-cavity lining), yet definite information on many of the physical and chemical factors that affect rates of growth is scarce. Results of published work are often conflicting and difficult to interpret because experiments were made with birds contaminated naturally during processing or handling and thus under conditions not controlled as to type and amount of inoculum. Comparative testing with controlled inoculation is required to determine the relative growth rates of the principal spoilage organisms and the extent to which these rates are affected by processing and storage conditions. Work has been undertaken, therefore, to study factors which affect the growth of pseudomonads and achromobacter (the two principal groups of bacteria responsible for low-temperature spoilage of poultry (Ayres et al., 1950; Thornley et al., 1960: Walker and Ayres, 1956)) on the surface of chicken skin, muscle, and gut-cavity lining. This paper outlines methods developed for preparing and inoculating skin and describes the effect of several factors (birdto-bird variability, age of bird, location of skin on carcass, freezing and thawing of skin, and methods of scalding) on the growth of the two groups of organisms on the skin surface at 0°C. Skin was chosen as the initial substrate for study because published work (Lockhead and Landerkin, N. R. C. No. 10,055.
METHODS
Skin was obtained from broiler-type chickens (Ottawa Meat Control Strain) raised and processed in the laboratory. After slaughter (by cutting the jugular vein and carotid arteries), the chickens were scalded (at 59°C, unless otherwise stated), plucked by hand and rubbed free of loose pieces of the epidermal layer of the skin and parts of feathers with a sterile damp cloth. The skin was removed from the carcass before evisceration, washed for 15 seconds in a hot (59°C.) detergent solution (0.05% Sparkleen), rinsed thoroughly in cool sterile water, and cut into pieces or samples approximately 9 cm. square. Each piece was stretched, outside surface up, onto a circular stainless steel disk (diam. 7.6 cm.) with hooks around the outside edge to stretch the piece enough to smooth out all visible folds. The excess skin was trimmed off. The samples were held in sterile Petri dishes in a saturated atmosphere at 0°C. until inoculated (within 24 hrs). Preliminary tests showed that the skin samples were essentially free of psychrophiles; counts on uninoculated samples incubated at 0°C. for 10 days never exceeded 50 per sq. cm. Two inocula were used in all of the tests; one was composed of a mixture of 10
1575
Downloaded from http://ps.oxfordjournals.org/ at California Institute of Technology on June 20, 2015
of fresh refrigerated poultry S POILAGE is ordinarily caused by psychrotoler-
1935; Ziegler, 1954) has indicated that microbially-induced changes leading to offodour in dressed eviscerated poultry occur first on the surface of the skin; 0°C. was selected because a large percentage of unfrozen poultry (about 95% in Canada) is stored and shipped in ice.
1576
D. S. CLARK
location, 4 skin samples, one from each of 4 locations (back, breast, and upper and lower leg), were inoculated simultaneously in each test. The test was repeated 10 times for each type of inoculum. Studies to determine the effect of freezing and scalding on growth rate of Pseudomonas and Achromobacter were made with skin from the breast and leg areas of 10-weekold chickens. In freezing tests, growth rates on fresh skin were compared with those on skin that had been stored frozen for 1 month and then thawed at room tempertaure before being stretched and inoculated. Two methods of freezing were used; in one, skins sealed separately in cryovac bags were immersed for 5 min. in an alcohol-dry ice bath (-72°C.) and stored at - 1 0 0 ° C ; in the other, skin samples similarly packaged were frozen and stored in a room held at — 10°C. The growth rate tests were repeated 6 times for both types of bacteria using skin scalded at 59°C; in each test 2 fresh samples and 2 samples each of fast-frozen ( — 72°C.) and slow-frozen skin ( —10°C.) were inoculated simultaneously. To test the effect of scalding, the growth rate on unscalded skin was compared with the rates on skin scalded for 2 min. in water at 54°C. and for 1 min. in water at 59°C. The 54°C. scald (used commercially by some small producers) permitted removal during rub-down of only a portion of the yellowish epidermal layer of skin, particularly that on the tops of the mounds around the feather follicles, while TABLE 1.—Effect of scalding on growth of Pseudomonas and Achromobacter Counts per sq. cm. of skinXlO 6 after 6 days at 0°C*
Pseudomonas Achromobacter
TVT .
Scalded
Scalded
1.3 2.5
2.0 1.2
2.5 0.9
* Initial inoculation, 2.5X10 4 per sq. cm.
Downloaded from http://ps.oxfordjournals.org/ at California Institute of Technology on June 20, 2015
strains of Pseudomonas and the other of 10 strains of Achromobacter. The organisms were isolated from the surface of processed chickens in poultry plants and classified according to Thornley et al. (1960). All grew well at 0°C. Cells of each strain were harvested from 24-hr-old SM agar (Difco) cultures, suspended in a 0.1% aqueous peptone solution (Straka and Stokes, 1957) having an ionic strength of 0.5 (Torloni, 1962a), adjusted photometrically to 50% T at 490 m(/.., mixed in equal volumes in a nebulizer (Torloni, 1962b) and sprayed uniformly (about 25,000 cells per sq. cm. in all tests) over the surface of skin samples in an inoculation chamber (Clark, 1963). After inoculation the skin samples were incubated in a saturated atmosphere at 0°C. and sampled after 6 days. In one test, to determine the growth curves of the two types of organisms on scalded skin, counts on inoculated breast samples were assessed every two or three days during a 13 day incubation period. In sampling, three 2 sq. cm. areas on each skin sample were washed by the spray-gun technique (Clark, 1965a, b) and duplicate counts for each washing were determined by surface plating (Clark, 1967) on SM agar. The agar plates were incubated at 20°C. Analyses of variance were made on the counts. The effects of bird-to-bird variability and skin age and location on bacterial growth were studied with skin (scalded at 59°C.) from 8- and 16-week-old chickens. In bird-to-bird variability tests, each inoculation was made with six samples of breast skin, 1 each from six different 8-week-old birds. Six such tests were made for each group of bacteria, using skin samples from a different set of birds every second time. Tests for determination of effect of age were also repeated 6 times for each inocula, using 3 skin samples from 8-week-old birds and 3 skin samples from 16-week-old birds in each inoculation. To study the effect of
1577
PSEUDOMONADS AND ACHROMOBACTER ON SKIN
the 59°C. scald (widely used commercially) allowed complete removal of the outer layer of skin during rub-down. Unscalded skin, obtained from birds plucked "dry" was washed for 15 sec. in detergent solution at 25°C. and skin scalded at 54°C. was washed in detergent solution at 54°C. The growth tests were repeated 10 times; in each test 2 skin samples for each of the three conditions were inoculated simultaneously.
in
1
8
^ *,• ^^*^
y^\^^'
-
PSEUDOMONAS -—^
S 7 fe 6
,/Z-
»^^^ ACHROMOBACTER
^ ^ ^ ^ ^ ^
I
2
3
i _1 1 1 1 1 1 1_ 4 5 6 7 8 9 10 II 12 13 INCUBATION TIME, DAYS
Table 1 shows that scalding had a sig- FIG. 1. Growth of Pseudomonas and Achromobacter nificant effect on the growth rate of both on skin scalded at 59° C. types of organisms. Pseudomonas grew best on scalded skin, particularly skin scalded calculated from analysis of variance of the at 59°C, whereas Achromobacter grew results, were not significant for all factors best on unscalded skin; skin scalded at the and their interactions. The corresponding lower temperature (54°C.) gave intermedi- values for Achromobacter were: average ate growth rates in both cases. These count, about 900,000 cells/sq. cm.; and differences appear to be the result of rela- standard deviation between 370-540. tive preferences for the outermost and un- Again, the F values were not significant. derlying layers of skin as substrates, since Figure 1 shows the relative growth rates of the intermediate growth rates were ob- the two groups of organisms on skin tained when part of the epidermal layer scalded at 59°C. The "mean generation was removed (54°C. scald). When pseudo- time" (Elliott and Michener, 1965) for the monads are the predominant spoilage orga- Pseudomonas was about 12 hr while for the nisms on fresh commercially-processed Achromobacter it was about 14 hr. These poultry, as they frequently are (Ayres et generation times are approximately the al, 1950; Thornley et al., 1960; Nagel et same as those reported for psychrotolerant al., 1960), the shelf-life for birds scalded at bacteria on the skin of commercially pro59°C. should be significantly shorter than cessed chickens contaminated naturally that for birds scalded at 54°C. This would during processing (Dawson et al., 1963; seem to explain the longer shelf-life re- Hall and Spencer, 1963). ported for poultry scalded at the lower The study showed that any location on temperature (Ziegler and Stadelman, the skin of the bird is suitable for sampling 1955). in tests to determine the bacteriological Unlike scalding, the other factors exa- quality of whole refrigerated poultry, since mined in comparative tests (bird-to-bird psychrotolerant spoilage bacteria grow variability, age, location and freezing and equally well anywhere on the skin surface. thawing) had no significant effect on the The study also showed that in comparative growth rate of either type of organism. The studies of factors affecting bacterial growth average count after 6 days in all tests with on chicken skin, skin from different birds Pseudomonas was about 2,000,000 cells/sq. and from birds of different ages (at least cm.; the standard deviation in each case between 8 and 16 weeks old) can be used was between 270 and 340 and the F values, indiscriminately. In addition the skin can
Downloaded from http://ps.oxfordjournals.org/ at California Institute of Technology on June 20, 2015
RESULTS AND DISCUSSION
E dIO \ 9
1578
D. S. CLARK
also be frozen for convenience in storing a supply for continued or periodic work. ACKNOWLEDGMENTS
The author wishes to thank N. U. Cholette for skilled technical assistance. REFERENCES
NEWS AND NOTES (continued from page 1574) Forsythe, Henningsen Foods, Inc., Springfield, ELANCO NOTES Missouri. The U.S. Participation Committee is a During the annual meeting of the Poultry and Egg National Board in Chicago, Elanco Products function of the U.S.A. Branch of the World's Poultry Science Association. Co., Indianapolis, Indiana, a division of Eli Lilly The following Committee Chairmen were Company, was elected to membership in the Good Egg Club "for outstanding cooperation and general named: Science—Dr. Hans Lineweaver, Western Regional Research Laboratory, U.S. Department excellence in promotion of eggs." of Agriculture, Albany, California; Papers SelecCONGRESS COMMITTEES tion—Dr. R. N. Shoffner, University of MinneThe U.S. Participation Committee for the 14th sota, St. Paul, Minnesota; Travel Grants—Dr. World's Poultry Congress, which is scheduled for W. J. Stadelman, Purdue University, Lafayette, September, 1970, in Spain, was activated at a Indiana; Commercial Participation—Richard Ammeeting in Chicago, Illinois, on August 23rd, under nion, N.E.P.P. Co., Trenton, New Jersey; Memthe direction of its Chairman, Dr. Richard H. bership and Promotion—Gene Shephard, Institute (continued on page 1611)
Downloaded from http://ps.oxfordjournals.org/ at California Institute of Technology on June 20, 2015
Ayres, J. C , W. S. Ogilvy and G. F. Stewart, 1950. Post mortem changes in stored meats. I. Microorganisms associated with development of slime on eviscerated cut-up poultry. Food Technol. 4 : 199-205. Clark, D. S., 1963. Uniform inoculation of nutrient surfaces. Biotech. Bioeng. 5 : 123-129. Clark, D. S., 1965a. Method of estimating the bacterial population on surfaces. Can. J. Microbiol. 11:407-413. Clark, D. S., 1965b. Improvement of spray gun method of estimating bacterial populations on surfaces. Can. J. Microbiol. 11: 1021-1022. Clark, D. S., 1967. Comparison of pour and surface plate methods for determination of bacterial counts. Can. J. Microbiol. 13: 1409-1412. Dawson, L. E., W. L. Mallmann, D. G. Bigbee, R. Walker and M. E. Zabik, 1963. Influence of surface pasteurization and chlortetracycline on bacterial incidence on fryers. Food Technol. 17: 100-103. Elliott, R. P., and H. D. Michener, 1965. Factors affecting the growth of psychrophilic microorganisms in foods. Technical Bulletin No. 1320, Agricultural Research Service, U.S. Dept. Agriculture: p. 2.
Hall, K. N., and J. V. Spencer, 1963. The effect of ethanol on shelf-life and flavor of chicken meat. Poultry Sci. 4 3 : 573-576. Lochhead, A. G., and G. B. Landerkin, 1935. Bacteriological studies of dressed poultry. I. Preliminary investigations of bacterial action at chill temperatures. Sci. Agri. 15: 765-770. Nagel, C. W„ K. L. Simpson, H. Ng, R. H. Vaughn and G. F. Stewart, 1960. Microorganisms associated with spoilage of refrigerated poultry. Food Technol. 14: 21-23. Straka, R. P., and J. L. Stokes, 1957. Rapid destruction of bacteria in commonly used diluents and its elimination. Appl. Microbiol. 5 : 21-25. Thronley, M. J., M. Ingram and E. M. Barnes, 1960. The effects of antibiotics and irradiation on Pseudomonas-Achromobacter flora of chilled poultry. J. Appl. Bact. 23: 487-498. Torloni, M., 1962a. Influence of the ionic concentration of nebulized suspensions on stability of microbial aerosols. Biotechnol. Bioeng. 4 : 341-344. Torloni, M., 1962b. A simple and versatile nebulizer. Appl. Microbiol. 10: 6-8. Walker, H. W., and J. C. Ayres, 1956. Incidence and kind of microorganisms associated with commercially dressed poultry. Appl. Microbiol. 4:345-350. Ziegler, F., J. V. Spencer and W. J. Stadleman, 1954. A rapid method for determining spoilage in fresh poultry meat. Poultry Sci. 3 3 : 12531255. Ziegler, F., and W. J. Stadelman, 1955. The effect of different scald water temperatures on shelflife of fresh non-frozen fryers. Poultry Sci. 34: 237-238.