APPLIED MICROBIOLOGY, Aug. 1970, p. 208-214Copyright © 1970 American Society for Microbiology

Vol. 20, No. 2Printed in U.S.A.

Almond Harvesting, Processing, and Microbial FloraA. DOUGLAS KING, JR., MARY JO MILLER, AND LINDA C. ELDRIDGE

Western Regional Research Laboratory and Biometrical Services Staff, Agricultural Research Service,Albany, California 94710

Received for publication 23 March 1970

This survey was set up on a statistical sampling plan to determine the microbialquality of almonds as they are received at the processing plant. The total aerobicbacterial count and yeast and mold count distribution were skewed by a few highcounts compared with the majority of relatively low counts. Hard shell varieties ofalmonds had lower counts than did soft shell, and almonds with complete shells hadlower counts than shelled almonds. Almonds harvested onto canvas had lowercounts than those harvested by knocking onto the ground. Nuts with the leastamounts of foreign material mixed into the sample had the lowest counts, as didnuts with the least amount of insect damage. Coliforms, Escherichia coli, and Strep-tococcus were isolated from the nuts, and their presence was correlated with soilcontamination. When almonds are stored, the total plate count, the Streptococcuscount, and the E. coli count after an initial drop remain nearly constant for more

than 3 months. In addition to the indicator organisms, several genera of bacteriawere isolated including Bacillus, Xanthomonas, Achromobacter, Pseudomonas,Micrococcus or Staphylococcus, and Brevibacterium.

Edible sweet almonds (Prunus amygdalus)have three distinct parts: the inner kernel ormeat, the middle shell portion, and an outerhull. Almond varieties vary in shell texture;therefore, they are termed hard or soft shelled.The harvesting procedure starts when the almondsare partly dried on the trees. They are shakendown to collecting sheets or onto the ground andare mechanically picked up after further drying.A hulling operation then removes the outerhulls. During this operation, some nutmeats areinadvertently removed from the shell. Thehulled nuts are then sent to the processing plantwhere the nuts are fumigated to destroy insectsand eggs before shelling. After the shell has beenremoved, the nutmeats are processed into gradedmeats or almond products.

This research was conducted at the request ofthe almond industry to determine the microbialquality of almonds as received at the processingplant. A preliminary report of this research hasbeen published (5). There has been a continuinginterest in coliforms and Escherichia coli in re-lation to almond contamination (7, 9, 13).

MATERIALS AND METHODSSampling plans. A preliminary survey showed that

10% of the almond samples were contaminated withcoliforms. Therefore, our statistical sampling plan in1966 was set up to detect coliform contamination onnutmeats in 10% of the lots with 95% confidence. A0.25- to 0.5-lb sample (113 to 227 g) was collected fromevery 200th lot of nuts received at the California Al-

mond Growers Exchange plant in Sacramento for atotal of 172 samples. It was shown in a separate pre-liminary unreported study that samples so collectedwould represent receipts at all almond processingplants because samples from all growing areas wouldbe included.

Samples used were subsamples of those taken byautomatic sampling devices used to grade each lot.The samples were carefully packaged in sterile plasticbags to prevent additional contamination. They weretaken immediately upon receipt at the plant and re-frigerated until transported to the laboratory wherethey were stored at 2 C in metal containers to preventmoisture pickup.

During the 1967 season, the sampling plan waschanged to include samples that contained suchforeign material as soil and sticks, as well as thosesamples representative of harvesting methods, varie-ties, and growing areas. A total of 99 samples wasselected.

Microbial analyses. Nutmeats were removed fromthe sample bag or from the shell in an aseptic mannerand placed in a sterile bottle. Hard-shelled varietieswere cracked with a small hammer before the meatwas removed with forceps. An equal weight of waterwas added to the approximately 10 g of nutmeats inthe bottle, and the sample was shaken to wash thesurface of the nutmeat. After 5 min, the sample wasagain shaken, and portions were removed for dilutionin 0.1% peptone water (12) or inoculation of plates formicrobial counts.

Bacteria counts were made with plate count agarcontaining 100 ,ug of cycloheximide (Upjohn) per mladded to suppress mold growth.

Yeast and mold counts were made with potatodextrose agar acidified to pH 3.5 with tartaric acid

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BAClTERIA IN ALMONDS

TABLE 1. Statistical data on microbial counts per gram ofalmond samples by harvest yeara

Bacteria Yeast and moldDetermination

1966 1967 1966 1967

Range ..................... 8-30,000 89-50,000 0-21,600 2,200-530,000Mean 2,828 6,955 1,494 47,159Median .................... 1,800 3,600 440 27,000Ninth decile ............... 6,000 18,800 4,600 103,000Standard deviation ........ 3,731 8,985 3,314 66,138No. of samples tested 172 99 172 99

a Sampling plans used during 1966 and 1967 require different interpretations of the data. The 1966samples were selected to represent receipts at all processing plants. Most samples from 1967 were se-lected because they contained contaminating material at the receiving inspection.

Coliform counts were carried out by using VioletRed Bile Agar with an agar overlay (2). Positive cul-tures were transferred to Lauryl Tryptose Broth. Fromtubes showing gas, cultures were transferred to BoricAcid Lactose Broth (BALB), and a Gram stain wasmade. Cultures positive on BALB were streaked ontoLevine's Eosin Methylene Blue plates which, whenpositive, were confirmed as E. coli by procedures sug-gested by the American Public Health Association(1, 2).

Streptococcus counts were made on KF Strepto-coccus agar (8) containing 0.01% 2,3,5-triphenyl-tetrazolium chloride. During the 1966 season, allpositive cultures were transferred to Azide DextroseBroth. Those positive on this medium were transferredto Ethyl Violet Azide (EVA) Broth. Of 22 samples(13% of all samples) positive on KF Streptococcusagar, all except one were positive on EVA Broth. Be-cause of the selectivity of the KF Streptococcus agar,the Azide Dextrose and EVA Broth tests were notused in 1967.

Identification of bacteria. Routine methods wereused to plate four samples of almond meats, two fromalmonds at the processing plant and two from palletsof bagged almonds ready for shipment after proc-essing.From each plate, 25 colonies were picked at random

and transferred to BBL Trypticase Soy Broth. Thetubes were incubated at 35 C for 72 hr, and the charac-teristics of the colonies were noted. A loopful of brothwas then streaked on BBL Trypticase Soy Agar (TSA)plates to isolate any mixed colonies (incubated 72 hr at35 C). Colony characters were noted, and the isolatedorganisms were transferred to TSA slants (incubated24 hr at 35 C).

Appropriate descriptive tests were run on the iso-lates by use of standard methods (11). After thesetests were completed, the organisms were grouped bygenera (10).

RESULTSMicrobial counts. The total bacterial plate

counts on the almonds were quite low, on theaverage, when compared with those of fooditems that are not dry processed. The highest

value of bacterial counts is within the suggestedlimit of counts for many foods (4). During bothseasons, the yeast and mold count plates con-tained mostly mold and very little yeast.The data shown in Table 1 illustrate the wide

range and distribution of microbial counts. Themedian is considerably lower than the mean,indicating that the majority of counts are low,with a relatively few high-count samples thatskew the data. Ninety per cent of the valuesfound are below the value listed as the ninthdecile in Table 1.To compare microbial contamination of the

unprocessed nuts with that of processed nuts, aseries of counts were made on processed almondmeats. For 11 samples from 1966, the averagecount for bacteria was 580 per gram and foryeast and mold, 170 per gram. Comparablevalues from 1967 (the average of 12 samples)were 1,200 and 2,700 per gram, respectively.These values are somewhat lower than the valueslisted in Table 1 for receiving supplies, indicatinga net drop in microbial count during the proc-essing.

Indicator organisms. We examined the un-processed nuts for coliform and streptococcuscontent as pollution-indicator organisms inalmond processing. Both types were found onthe nutmeats when delivered to the processingplant. Table 2 lists the counts obtained from thesamples containing these organisms. From the1966 season, all 22 streptococcus-positive sam-ples were isolated from soft-shell varieties thathad been shelled or had split shells. Coliformswere also present in 11 of these samples, butonly two had E. coli. Of the 1967 samples, 51(52%) had Streptococcus organisms present,only 17 having come from whole-shell nuts.In the 1967 tests, coliforms were also found in 33of the 51 streptococcus-positive samples, but only2 of the 33 had E. coli.Of the 172 samples tested in 1966, the 42

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KING, MILLER, AND ELDRIDGE

TABLE 2. Numbers of inidicator organisms per gramof nutmeats oit positive samples for 1966

and 1967

Organisms 1966 1967

StreptococcusNo. positive samples/

total samples 22/172 51/99Mean counts per gram 26 14Range 1-345 1-124

ColiformNo. positive samples/

total samples 42/172 40/99Mean count per gram 8 11Range................... 1-156 1-158

which had coliforms (24%) were the soft-shelledNonpareil variety. Correlations shown in Table 3for the 1966 data indicate that coliforms weremainly present in almonds shelled before ship-

ment to the processing plant and those with splitshells. E. coli was present in seven (4.1 %) of thesamples, a value somewhat lower than thatfound by Kokal and Thorpe (7). A total of 40of the 99 samples from the 1967 sampling hadcoliform organisms present. Of these, only ninecontained E. coli. None of the correlationscalculated for numbers of coliform organismswas significant. E. coli was isolated only fromnuts that had the shell removed or had splitshells.

Correlations of microbial counts and test vari-ables. To give a more normal distribution to thedata shown in Table 1, the microbial countswere expressed as logs from which geometricmeans and confidence intervals were estimated(3). The confidence intervals were based onStudent's t distribution at the 95% probabilitylevel. Table 3 shows the correlations of microbialcounts and some of the 1966 and 1967 test vari-ables. Variables reported in per cent were trans-

TABLE 3. Correlationi coefficients of microbial data with individual sample test criteria

Determination

Per cent nuts in shellPer cent shelled meatsPer cent shellsShell conditionType of shellPer cent sticktight hullsPer cent foreign mate-

rialPer cent reject (ined-

ible)Aerobic plate countYeast and mold countStreptococcus countColiform count

Per cent nuts in shellPer cent shelled meatsShell conditionType of shellPer cent sticktight hullsPer cent foreign mate-

rialPer cent reject (ined-

ible)Harvest methodInsect damageAerobic plate countYeast and mold countStreptococcus countColiform count

Aerobic platecount

-0.175*0.398**

-0.513**0.576**

-0.637**-0.221**0.273**

0.303**

-0.358**0.254*0.308**

-0.357**-0.357**0.418**

0.391 **

0.296**0.476**

a No. = 150.b No. = 169.cNo. = 99.* Significant at 5% probability.** Significant at 1%; probability.

Year

1966b

1967c

Yeast andmold count

-0.1200.285**

-0.438**0.479**

-0.529**-0.135

0. 152*

0. 184*

0.506**

-0.1310.0390.083

-0.194-0.0770.192

0.320**

-0.1450.238*0.201 *

Streptococcutscount

-0.251**0.260**

-0.295**0.266**

-0. 198**-0.211**0.126

0. 181 *

0.299**0.179*

-0.484**0.373**0.349**

-0.048-0.413**0.162

0.224*

0.1560.421**0.479**0.150

Coliformlacount

-0.302**0.340**

-0.336**0.333**

-0.149-0.214**0.033

0.298**

0.211**0. 198*0. 197*

-0.394**0.371**0.376**

-0.171-0.336**-0.041

0.252*

0.0520.366**0.439**0.1290.475**

E. coli'

-0.442**0.494**

-0.377**0.364**

-0.120-0.206*-0.173*

0.239**

0.1500.0890.177*0.461**

-0.228*0.1560.1520.017

-0.201*0.189

0.063

0.0550.0590. 196*0.1200.364**0.332**

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BACTERIA IN ALMONDS

formed by arc sine square root and count databy logs. No influence of growing areas upon themicrobial population was found.

Variety. There were differences in microbialcontent among individual varieties that appearto be a reflection of the shell type or condition,rather than a difference in chemical compositionamong the varieties.

Shell type and shell condition. Almond shellsvary in texture from fragile to hard, dependingon the variety. The varieties we studied wereclassified as hard or soft shell. Soft-shelled vari-eties were IXL, Jordonalo, Neplus, and Non-pareil; hard-shell varieties were Davey, Drake,Llewellyn, Mission, and Peerless. The complete-ness of shell is related to its hardness (Fig. 1).The influence of shell type and condition

upon the microbial counts of almond meats isshown in Table 4. In general, the hard-shelledalmonds had lower counts than the soft-shelledvarieties. The more complete the shell, the lower

FIG. 1. In-shell almonds after removal of hulls.Notice the difference in amount of shell breakage andexposure of meats between the hard-shell (left) andsoft-shell (right) varieties as they appear when deliveredto the processing plant.

the microbial content of the meat. These dataillustrate the importance of a complete shell forpreventing contamination. The fact that the shellsfrequently crack and meats are exposed duringdrying on the tree, particularly with soft-shelledvarieties, indicates the need to maintain rela-tively clean conditions during all handling stepsin order to minimize contamination of the nut-meat.Method of harvest. During the 1967 season,

we obtained samples that had been harvestedby knocking the nuts either onto some type ofground cover (such as canvas) or onto the soil,which usually is carefully rolled and preparedin advance. Nuts without contamination, har-vested by either method, were statisticallycompared (Table 5). Those harvested on groundcloths had significantly lower bacterial and yeastand mold counts than those picked from theground.

Contamination materials. During the 1967season, some samples were selected from incom-ing shipments of almonds that contained notice-able dirt. These samples were classified andanalyzed by the kind of foreign material present.The classes were: no noticeable contamination,mud balls and pieces of dried manure, mudballs only, and other foreign material (lint,ground in soil on meats, worms, rubber, rocks,etc.). The analysis shown in Table 6 illustratesthe influence of the various contaminants uponthe microbial counts.

These data indicate that bacterial counts arerelated to the amount of soil mixed with thesample. The three contaminated categories(Table 6) had significantly higher bacterial countsthan did samples with no foreign material.Although not shown in the table, the highestcounts that we noted were obtained from samples

TABLE 4. Geometric means and their confidence intervals for microbial counts by shell type and conditiontfor 1966 and 1967

Bacteria Yeast and mold

Shellcondition

1966WholeSplitRemoved

1967WholeSplitRemoved

Soft shell Hard shell Soft shell Hard shell

No.

if8'2(

2',

Geo-. metric

mean

6 1, 5107 2,0306 3,770

8 1,6103 4,4509 6,260

Confidenceinterval(95%)

640-3,5501,690-2,4302,540-5,610

420-6,1903,000-6, 5903,630-10,800

a Insufficient data.

No.

4021

81811

Geo-metricmean

1611,4309,700

1,0801,3902,280

Confidenceinterval(95%)

97-267-a

-a

292-4,010716-2,690881-5,910

No.

if822f

Geo-metricmean

6 3747 6206 1,030

8 22,1003 33,8009 29,900

Confidenceinterval(95%)

152-924460-837636-1,650

5,850-83,800

No.

4021

8

Geo-metricmean

39257

7,900

18,80023,700-48, 300 18 19, 10019,600-45,600 11121,500

Confidenceinterval(95%)

18-85-a

-a

6,430-54,80010,100-36,20010,000-46,100

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KING, MILLER, AND ELDRIDGE

TABLE 5. Geometric means and their confidenice intervals for microbial countsbased on harvest methods for 1967

Bacteria Yeast and mold

Harvest methodNo. Geometric Confidence intervalNo.

mean (95(7) Geometric mean Confidence interval (95%)

Sheets ............. 13 700 360-1,370 40,900 22,200-75,200Ground ............ 39 1,970 1,330-2,910 18,500 12,500-27,400

TABLE 6. Geometric meanis anzd their contfidentce intervals for microbial coIIuIts by type of contaminationt in1967 nlut delivery

Bacteria Yeast and moldContamination

No. Geometric mean Confidence interval (95%) Geometric mean Confidence interval (95%)

None ........... 52 1,500 1,070-2,160 22,600 16,100-31,600Manure and mud

balls........... 12 4,100 1,670-10,100 24,100 14,300-40,600Mud balls..... ... 26 6,200 3,830-10,100 36,500 23,300-57,000Other........... 9 13,900 7,830-24,800 23,600 10,500-53,100

with soil ground into the nutmeats and not easilyremoved. The yeast and mold count did not fol-low this pattern and appeared to be unrelatedto such contamination.

Insect damage. The 1967 crop had an un-usually large amount of insect damage, primarilydue to the navel orange moth. Samples werevisually graded with respect to the amount ofinsect damage on the nutmeat and were labeledfrom 1 to 5, to indicate no damage throughincreasingly high damage. The data (Fig. 2) showa positive relationship between the amount ofinsect damage and aerobic plate count (note acorrelation of 0.476 from Table 3). Three sampleswith least insect damage had significantly lowerbacterial counts than the two with the mostdamage. For yeast and mold content, samplecategories 1 and 3 were significantly lower than 5.Types of bacteria isolated from almond meats.

To determine the types of aerobic bacteria presenton almond meats, a total of 25 colonies wereisolated from each of two processed and twounprocessed samples. The colonies were classifiedas to genera by the method of Skerman (10).Most (60) of the cultures were Bacillus species,gram-positive, and catalase-positive with endo-spores.

Numerically, the second most important group(16 isolates) was the gram-negative, catalase-positive, motile, and rod-shaped bacteria. Ofthese, two seemed to fit the classification of thegenus Xanthomonas and two Achromobacter.Except for two nonmotile cultures, the remainingbacteria had polar flagella but could not be easily

placed in a genus, although they were related toPseudomonas as were two nonmotile cultures.The third group, containing 15 isolates, con-

sisted of gram-positive cocci that fit into theclassification of Micrococcus or Staphylococcus.The last bacterial group of seven isolates canbe described as gram-positive, catalase-positive,having nonmotile rods that do not form endo-spores, and loosely classified in the genus Brevi-bacterium.

Survival studies. Figure 3 shows the survival oftotal bacteria, E. coli, and Streptococcus specieson almond meats stored in plastic bags at 2 or24 C. The total bacterial counts were made onnormal samples of almonds; for the counts of E.coli and Streptococcus species, the organisms were

1,,000 80,000

7,0.000-

o ~ > __ __O Fi- tJ 50,000\il'>0'.0\0,I<L,000 _ 40,000_ ,',

V _ 7 _ m -~~~~~~~~30.000- 777ii'7:;""

2 3 4 2 3 4 5

SEVERITY OF INSECT DAMAGE

FIG. 2. Geometric means and their 95% confidenceintervals for severity of insect damage to almond meatsin storage. The amount of insect damage increases fromnone (1) to severe (5).

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200 1000 _ 5000

180_m

160 m 800 4000z

1140 o~~~~~~~~~~~~0120 6 600 3000_0100 0

80 LU400 -2000 '

60 @ *,>_ STREP. (2°C) 0| 40X l 200 AEROBIC PLATE COUNT (2'C)

20 -----------ECOLI (2C) 500STREP.(24 C)

0 -0 00 20 40 60 80 100 120 140 160 180 200 220 240

DAYS OF STORAGE

FIG. 3. Survival of total aerobic plate count bacteria,E. coli, anid Streptococcus on almond meats stored inplastic bags at 2 or 24 C.

inoculated on almonds before storage. Figure 3indicates that after an initial drop, the totalaerobic count is nearly constant with time andshows no marked decline in 225 days at 2 C.Also, there is an initial decrease in the E. colicount which precedes a gradual leveling off.After 225 days of storage, E. coli was still present.Almonds inoculated with Streptococcus speciesand stored at both 2 and 24 C show curvessimilar in shape to those of total bacteria and E.coli survival in cold storage. The data show aninitial steep drop in the count in the first severaldays and then a leveling off. At room tempera-ture, the organism dies off more rapidly than atcold temperatures.

Salmonella typhimurium also was inoculatedon whole almond meats that were stored at 2 C.After 190 days, the organism could still be re-covered, indicating a long survival time onalmond meats. Thus, if the nutmeats are con-taminated, the bacteria can persist for a long time.Kokal (6) reported that the tannins of walnut

skins kill E. coli. In experiments with groundalmond skins added to the growth medium, wewere unable to demonstrate a similar influence;in fact, the unwashed skins had a stimulatoryinfluence on E. coli, further indicating the non-antagonistic aspect of the almond to bacteria.

DISCUSSIONThe data reported in this paper reflect the

microbial population on the surface of almondmeats. A portion of the bacterial contamina-tion stems from soil and dust contact with nut-meats. This is shown by the lower counts forhard-shelled varieties which have a more com-plete shell and less chance for soil contamination.The effect of soil contact is also reflected in thelower counts for nuts harvested on cloths asopposed to those collected from the ground.

Nuts with the lowest amount of contaminatingforeign material had lower counts than thosewith large amounts, particularly if pieces of soilwere ground into the meats. Portable catchingframes or ground cloths would lessen soil andmicrobial contact. The increase in microbialcounts with increased insect damage illustratesthe need for better insect control both in theorchard and after harvest.The yeast and mold surface counts correlate

with shell factors during the 1966 season but notin 1967. The correlation with rejected materialis an indication of the type of spoilage includedin this classification. The correlations do notexplain the wide range of yeast and mold countsobserved. The lack of correlations in 1967 isprobably a result of the different sampling planused that season.The indicator organisms (coliforms, E. coli,

and Streptococcus) were associated with soilcontamination of nutmeats, as were the totalbacteria. Kokal and Thorpe (7) showed that theincidence of E. coli on almonds was associatedwith processing operations after harvesting,although some E. coli cells were found on nuts be-fore harvest. The present findings indicate con-tamination of nutmeats before processing, so theirpresence on processed nuts does not necessarilyindicate poor manufacturing practice.We had hoped to determine if Streptococcus

or E. coli was the best indicator organism to usefor measuring contamination of almonds. Theyboth appear to be present in similar numbers andare associated with the same kind of contamina-tion. Neither test appears to be superior, exceptthat the Streptococcus test is easier and less timeconsuming. 0

The genera of bacteria isolated from the nutkernel, Bacillus, Xanthomonas, Achromobacter,Pseudomonas, Micrococcus, and Brevibacterium,would be expected to be associated with soil orplant material and to survive on the kernel. Thesurvival of heat resistant Bacillus species onalmonds could be of consequence if almonds areused as an ingredient for heat-processed foodproducts. Survival tests for total bacterial count,streptococci, S. typhimurium, and E. coli indicatethat the almond does not exert an inhibitoryeffect and that organisms can persist on thenuts for long periods of time. This illustrates theimportance of preventing contamination of thenutmeat, particularly during normal processingwhen there is only a dry cleaning operation; itis not common practice to wash nutmeats.The data show that the microbial quality of the

almond as it is received at the processing plantis generally good. However, the surface contami-nation of the nutmeat can be partially controlled

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214

by lessening the degree of soil or dust contactand thereby improving the quality of the productfor further processing.

ACKNOWLEDGMENTS

The authors recognize the Dried Fruit Association of Cali-fornia and the California almond industry for support andcooperation in this study; the California Almond Growers Ex-change (J. Mattei and A. Cardosa) for coordinating and obtain-ing the almond samples; L. Queen for laboratory assistance; andD. E. O'Connell and E. F. Schultz, Jr., for assistance in planningthe statistical analysis.

LITERATURE CITED

1. American Public Health Association. 1965. Standard methodsfor the examination of water, sewage, and industrial wastes,12th ed. American Public Health Association, New York.

2. American Public Health Association. 1966. Recommendedmethods for the microbiological examination of foods, p.

205, 2nd ed. American Public Health Association, NewYork.

3. Association of Food and Drug Officials of the U. S. 1966.Microbial examination of precooked frozen foods. Assoc.Food Drug Officials U.S., Quart. Bull. 30, p. 55-77 (supple-mental issue).

APPL. MICROBIOL.

4. Elliott, R. P., and H. D. Michener. 1961. Microbiologicalprocess report. Microbiological standards and handlingcodes for chilled and frozen foods. A review. Appl. Micro-biol. 9:452-468.

5. King, A. D., Jr., D. E. O'Connell, F. P. Boyle, and M. J.Miller. 1968. Find low count in almonds. Food Eng. 40:142.

6. Kokal, D. 1965. Viability of Escherichia coli on English walnutmeats (Juiglans regis). J. Food Sci. 30:325-332.

7. Kokal, D., and D. W. Thorpe. 1969. Occurrence of Escher-ichia coli in almonds of Nonpareil variety. Food Technol.23:227-232.

8. Lewis, K. H., and R. Angelotti (ed.). 1964. Examination offoods for enteropathogenic and indicator bacteria. PublicHealth Service publication no. 1142, Washington, D. C.

9. Ostrolenk, M., and A. C. Hunter. 1939. Bacteria of the colon-aerogenes group on nutmeats. Food Res. 4:453-460.

10. Skerman, V. B. D. 1967. A guide to the identification of thegenera of bacteria, 2nd ed. The Williams & Wilkins Co.,Baltimore.

11. Society of American Bacteriologists. 1957. Manual of micro-biological methods. McGraw-Hill Book Co., Inc., NewYork.

12. Straka, R. P., and J. L. Stokes. 1957. Rapid dostruction ofbacteria in commonly used diluents and its elimination.Appl. Microbiol. 5:21-25.

13. Wenzel, M. and L. A. Black. 1939. Bacterial species on

nutmeats used in ice cream mixes. Food Res. 4:191-201.

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