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Staphylococcus saprophyticus found to be a common contaminant of food
Journal of Infection (t99o) 2I, I I - I 9
Staphylococcus saprophyticus contaminant
found to be a common of food
P. H e d m a n , * O. Ringertz* B. E r i k s s o n , t P. Kvarnfors,$ M. Andersson,* L. B e n g t s s o n * a n d K. Olsson*
* Department of Clinical Bacteriology, S;~dersjukhuset, The Karolinska Institute, S-IoO 64 Stockholm, ~f Goman Food Factory, Stockholm and ~ The Health Care Council, Stockholm, Sweden Accepted for publication 7 February I99o Summary The mode of transmission of Staphylococcus saprophyticus, a urinary tract pathogen, was investigated in three related studies. The presence of this organism was sought, during a period of I year, in i331 specimens of various foods, in 920 beef and pork carcasses and on Io7 cultures which had been inoculated directly from abattoir workers' protective gloves. Staphylococcus saprophyticus was found to contaminate I6"4 ~o of the various food samples with a high prevalence of 34 % in raw beef and pork. It was common in both domestic and imported raw meat products. There was no seasonal variation in the presence of S. saprophyticus in the samples obtained from carcasses. The bacterium was found in 69 % of all cultures from the workers' protective gloves. We conclude that S. saprophyticus, originating from slaughtered animals, contaminates food and eventually colonises the human intestinal tract.
Introduction
Staphylococcus saprophyticus is a well recognised c o m m o n cause of acute u r i n a r y tract infection ( U T I ) , typically present in y o u n g w o m e n in outpatient practice with a seasonal peak incidence in late s u m m e r , as reviewed by Hovelius and Mfirdh. 1 T h e reservoir for this organism and its m o d e of transmission are now known. T h e skin has been proposed as the h u m a n reservoir of S. saprophyticus which has been d e m o n s t r a t e d on both h u m a n and animal s k i n ) '3 Others, however, have f o u n d S. saprophyticus as part of the rectal flora in healthy carriers 4 and in patients with acute u r i n a r y tract infections caused by the same species. 5'6 We have recently confirmed rectal carriage of S. saprophyticus in patients in a venereal disease clinic. 7 F u r t h e r m o r e , we have d e m o n s t r a t e d a seasonal variation in the colonisation of the r e c t u m and the u r e t h r a similar to that for U T I s caused by S. saprophyticus I n the present study various food samples were investigated in order to elucidate f u r t h e r the m o d e of transmission of S. saprophyticus and to gain clues on its origin. First, a heterogenous collection of routine food samples from the H e a l t h Care Council laboratory in Stockholm was studied during a period of I year. Second, in order to concentrate the study on meat products and to verify the time of the processing of the meat, specimens of beef and pork were examined from a food factory in Stockholm, also during a period of I year. oi63-4453/9o/o4ooii +o9 $02.00/0
© I99o The British Society for the Study of Infection
I2
P. HEDMAN E T A L .
Additionally, samples were obtained from the protective gloves of workers in the meat cutting department in the same factory during the same year. Materials and methods
Food samples from the Health Care Council laboratory in Stockholm were collected from May I982 to May x983 in IO monthly periods. A total of z33I specimens was gathered by routine methods as part of the normal bacteriological public health control of food. T h e y were collected mainly from public restaurants, retail grocery shops and from stores of imported food all over the Stockholm metropolitan area. In one series of samples, only vegetables were collected. All food specimens were thoroughly homogenised in food processors and then cultured in order to demonstrate food pathogens. Duplicate cultures were incubated at 37 and 44 °C respectively. A standard protocol recorded for each sample the kind of food examined, where it was collected, its country of origin, any processing performed and how it was stored. One ml of each homogenised food specimen was incubated for 48 h at 37 °C in a S. saprophyticus selective broth consisting oftryptone (Oxoid, U.K.) of pH 7"2 with added I5o/zg/ml nalidixic acid (Winthrop, U.S.A.) and 2/zg/ml novobiocin (Sigma, U.S.A.). Subcultures were made on blood agar (Blood agar base No. 2, Oxoid, U.K.) containing 5 % defibrinated horse blood. All Staphylococcus species were further tested for their susceptibility to novobiocin by an agar diffusion method with discs containing 5 #g novobiocin (Biodisk, Sweden). Strains demonstrating an inhibitory zone of less than I5 m m were further classified according to a modification of Kloos and Schleifer 8 in serial tube reactions (Voges-Proskauer test, urease production, nitrate reduction, anaerobic production of acid from glucose and fermentation of arabinose, mannose, xylose, sucrose and glucose). All strains of Staphylococcus species were also tested according to API Staph-Ident (Analytab Products, Plainview, N.Y., U.S.A.). Meat specimens from a food factory were collected from the cold store room in the Konsum Stockholm Goman factory from November 1985 to October I986. All meat was delivered by cold transportation from various slaughterhouses all over Sweden and reached the cold room within 3 days after slaughtering the animals and cutting the carcasses into halves or quarter pieces. T w e n t y samples of beef and pork were gathered every fortnight except for January I 9 8 5 when only 20 samples of each were collected. All sampling was sought to include a few slaughterhouses as possible. Altogether, 460 specimens of beef were cut from the neck of the carcasses in thin slices with a diameter of 3-5 cm by means of sterile disposable scalpels and forceps. All 460 pork specimens were cut in a similar way from the shoulder and neck of the pig carcasses. T h e specimens were incubated for 48 h at 37 °C in the same selective broth as described above. Isolation of S. saprophyticus followed the same procedures. T h e workers in the cutting room, close to the cold store in the food factory, used cotton gloves strengthened by thin steel wires for protection. T h e gloves were dipped in the previously described selective broth for S. saprophyticus (held in single plastic containers) for 30 s on four occasions during the study. T h e containers were incubated and subcultured as described above.
Staphylococcus saprophyticus in food
13
Table I The isolation rates (%) of Staphylococcus saprophyticus in food
samples collected by the Health Care Council in Stockholm from May I982 to May I983 Sample
I 2 3 4 5 6 7 8 9 Io Total
Date
I982 May June July August October December 1983 January March April May
Number of specimens
I84 I53 I 15 43* I7o 97
S. saprophyticus (%)
z5 14 II 5 24 25
2oi
16
136
2I
II6 I I6 I331
IO 13 I6-4
* Only vegetables collected.
All statistical analyses were done by means of X2 m e t h o d s unless otherwise stated. RESULTS Food collected by the Health Care Council of Stockholm
A total of I33I samples was examined. T h e zo sampling periods and the seasonal presence of S. saprophyticus are shown in T a b l e I. Altogether, z i 8 strains of S. saprophyticus were isolated and identified by means of the tube tests described above, so giving an overall prevalence rate of I6"4 % . T h e A P I S t a p h - I d e n t system identified 2 z I (96"8 %) of these strains as S. saprophyticus. T h e I33Z collected food specimens were heterogeneous. T h e y included raw food items such as freshly cut meat and fresh vegetables from cold storehouses, r e a d y - m a d e foods such as sandwiches sold in retail shops and cooked food from restaurant kitchens. T h e types of food specimens are grouped and the presence of S. saprophyticus in each type of food is presented in Table II. M o s t meat products consisted of pork and beef. A m o n g these 6oi items of meat, there were 27o samples of raw beef or pork. As m a n y as 34 % of these specimens were contaminated with S. saprophyticus. T h e r e were 12 samples in this study of raw meat collected from a retail shop within a slaughterhouse area. T e n of these samples contained S. saprophyticus. T h e r e were 38 specimens of ice-cream a m o n g the 68 dairy food samples but none of these contained S. saprophyticus. M o s t items of confectionery were r e a d y - m a d e pastries from retail bakeries and hence were composed of various food materials. M o s t items of poultry were freshly grilled chicken collected in retail shops. T h e r e were only four specimens of egg. One sample of boiled egg contained S. saprophyticus. T h e group of mixed groceries consisted of readymade complete courses of mixed food, sandwiches, salads and sauces. All food samples were collected from various locations g r o u p e d and specified in T a b l e
P. H E D M A N E T A L .
I4
Table II The frequency of Staphylococcus saprophyticus in various types of
food Specimens from which
S. saprophyticus was isolated Type of food Raw meat Processed meat Poultry and eggs Fish Dairy products Vegetables Bread Mixed groceries Total
N u m b e r of specimens
(N)
(%)
270 331 34 6I 68 205 63 299
92 70 8 8 5 I2 3 20
34 2i 24 13 7 6 5 7
i33i
218
I6'4
This distribution was not due to chance (X2 = 98"I6, P < o-ooI, six degrees of freedom).
III. Sources of food available to the public included public restaurants of various sizes as well as canteens for schoolchildren, retired people and staffs of various companies. There were 71 specimens of imported meat in this study. Most of these were of raw beef and came from few countries as specified in Table IV. There were 48 (36 %) specimens of raw beef contaminated with S. saprophyticus out of a total of I36 of Swedish origin. Imported beef was not significantly more contaminated (P = o'I3; Fisher's exact test). Information on the storage of the food items was available in I o7o (8o %) specimen records. Details are given in Table V. Specimens of heated food consisted of food served hot in restaurants. Only one of these 4o samples contained S. saprophyticus. No other significant human pathogen was isolated from the x331 food specimens. There were, however, 435 specimens incubated at 37 °C which yielded significant growth of Escherichia coli. Staphylococcus saprophyticus was isolated from Io4 of these and hence was found to be a significant (X2 = 25"9; P < o'oI) concomitant contaminator. As many as 53% of all specimens with growth of both species were samples of raw meat. On the other hand, there was no tendency (X~= o'8; P > o'o5) to find S. saprophyticus together with E. coli in samples cultured at 44 °C. Of 95 specimens incubated at 44 °C and found to be contaminated with E. coli, only I2 had concomitant growth of S. saprophyticus. No particular food was represented among these 95 specimens incubated at 44 °C and contaminated by E. coli. T h e specimens concomitantly contaminated with S. saprophyticus, however, were more often products of raw meat (P < o.oi, Fisher's exact test).
Staphylococcus saprophyticus
in f r e s h c a r c a s s e s f r o m t h e f o o d f a c t o r y
Among the 460 specimens of fresh beef, z24 (27 %) were contaminated with S. saprophyticus and r37 (30 %) of the 460 pork samples were found to contain the same bacterium.
Staphylococcus saprophyticus Table III
in food
I5
The origin of the
I 3 3 I food samples and the frequency Staphylococcus saprophyticus
isolation of
Staphylococcus saprophyticus Sample origin Restaurants and canteens Retail grocery shops Cold storehouses Wholesale stores No information Total Table IV
Number of specimens
(N)
(%)
575
57
IO
423
98
23
95 54 184 1331
37 4 22 218
39 7 12 16'4
Country of origin of 72 samples of imported beef and the frequency of isolation of S t a p h y l o c o c c u s s a p r o p h y t i c u s Staphylococcus saprophyticus Country Poland Australia Jugoslavia U.S.A. Total
Table V
Number of specimens
(N)
(%)
27 z5 18 1 71
7 8 16 I 32
26 32 89 45
Storage of food and the frequency of isolation of
Staphylococcus
saprophyticus
Staphylococcus saprophyticus Type of storage Cold Room temperature Frozen Heated No information Total
Number of specimens
(N)
(%)
755 I6I I I4 40 261 i331
I23 19 35 I 40 218
I6 I2 3i 15 I6'4
T h e r e l a t i v e d i s t r i b u t i o n s o f t h e g r o w t h o f S. saprophyticus in f r e s h b e e f a n d p o r k f o r e a c h m o n t h o f t h e y e a r are d e p i c t e d in F i g . I. T h e r e w e r e n o o b v i o u s seasonal trends.
Staphylococcus saprophyticus
on meat cutters' protective gloves
O n f o u r o c c a s i o n s , in N o v e m b e r I 9 8 5 , in F e b r u a r y I 9 8 6 , in M a y I 9 8 6 a n d in A u g u s t I 9 8 6 , a t o t a l o f IO7 s p e c i m e n s w a s c o l l e c t e d f r o m p r o t e c t i v e gloves. As
P. HEDMAN ET AL.
I6 75
50-r-n
I I -
i
25
FI
-i
o
d
NOV DEC
JAN
FEB
MAR APR MAY
dUN
JUL
AUG SEP
OCT
1985-1986 F i g . [. T h e i s o l a t i o n o f Staphylococcus saprophyticus i n b e e f a n d p o r k c o m p a r e d w i t h t h e relative distribution of urinary tract infectionsdue to the same bacterium during a period of I2 months as diagnosed at the Laboratoryof Bacteriology, S6dersjukhuset, Stockholm. (-), UTI; (I~), beef; ([]), pork.
m a n y as 74 (69 %) were positive for S. saprophyticus. T h e r e was no obvious seasonal distribution of these isolates. Discussion
T h e correct identification of S. saprophyticus is crucial for our study. O u r procedure is based on the classification o f Staphylococcus species presented by Kloos and Schleifer in z9758 and u p d a t e d by M ~ r d h and Schleifer in I986. 9 T h e serial tube reaction was designed to separate S. saprophyticus specifically f r o m the two closely related novobiocin-resistant species o f S. cohnH and S. xylosus. All the strains of S. saprophyticus isolated f r o m food samples were f u r t h e r tested by A P I S t a p h - I d e n t , an accepted 1° commercial test system. T h e two m e t h o d s agreed well. A m o n g the seven strains of Staphylococcus species that did not agree, the A P I S t a p h - I d e n t identified three strains o f novobiocinresistant S. sirnulans ,°.nd one strain o f novobiocin-resistant S. warneri. A P I S t a p h - I d e n t failed to identify three strains. T w o presumptive ideas derive from our study on the presence of S. saprophyticus in food. First, we assume that the h u m a n reservoir of this specific u r i n a r y tract p a t h o g e n is the patient's faecal flora and, second, we assume that colonisation of the r e c t u m originates from contaminated food. We f o u n d in an earlier report 7 a mean rectal carriage rate of S. saprophyticus of 2"6 % a m o n g 495 m e n and of 2"5 % a m o n g 433 women, all attending a venereal disease clinic d u r i n g the period of I year. In that study, there was a clear t e n d e n c y for a seasonal variation of the rectal carriage. T h e m a x i m u m prevalence rate of S. saprophyticus in the r e c t u m of 6"7 % preceded the m a x i m u m prevalence rate in
Staphylococcus saprophyticus in food
17
the urethra of 9-2 % by I month. Other studies have reported rectal carriage of S. saprophyticus. Pead and Maskell 4 f o u n d S. saprophyticus in the faecal flora among 6"4 % of I56 healthy y o u n g w o m e n attending a family-planning clinic. Marrie et al., ~ f o u n d the same b a c t e r i u m in the faecal flora o f two of nine college w o m e n with an acute U T I due to the same species. Finally, L a t h a m et al., 6 found S. saprophyticus in the r e c t u m of three of 27 w o m e n attending a student health centre. T h e r e c t u m is considered to be the h u m a n reservoir for U T I s caused b y S. saprophyticus in two papers. 11'6 In a series of studies, 12 Cooke et al., d e m o n s t r a t e d the route of transmission of the urinary pathogen E. coli from various hospital foods via the intestinal tract of the hospital patients. W e propose a similar m o d e of transmission for S. saprophyticus. T h e r e are reports of S. saprophyticus being isolated from various sites on the h u m a n skin 2' 13 and the skin has been p r o p o s e d as the h u m a n reservoir of S. saprophyticus infections. ~ W e assume that colonisation of h u m a n skin is due to faecal contamination in a way similar to that for various G r a m - n e g a t i v e Enterobacteriaceae which are often isolated from the h u m a n skin.14 W e agree that S. saprophyticus should be classified as a transient coloniser of the h u m a n skin.3. ,5 T h e overall prevalence rate of S. saprophyticus of 16"4 % in the heterogenous material of 1331 food specimens is high. Cooke et al., 12 f o u n d a prevalence rate of contaminating E. coli in hospital food to be less than IO %. In s u b g r o u p i n g all food samples there was a marked p r e p o n d e r a n c e of S. saprophyticus in animal p r o d u c t s and especially in raw b e e f and pork where the prevalence rate of S. saprophyticus was as high as 34 %. T h e presence of S. saprophyticus in meat has been reported before. 2' 16 T h e high prevalence of S. saprophyticus in chicken meat suggests a future s t u d y of chickens' faecal flora. It is worth noting that S. saprophyticus was not isolated from ice-cream which is a typical food during the high seasonal prevalence of U T I s caused b y the bacterium. Staphylococcus saprophyticus has been f o u n d to be capable of transfer from hand to hand and from hand to fomites in experimental studies, iv,is Staphylococcus saprophyticus transferred more readily than Streptococcus pyogenes and E. coli and other Enterobacteriaceae. W e consider that the high prevalence of S. saprophyticus in one c o m m o n food such as raw meat, together with its ability to transfer easily on handling, explains w h y the b a c t e r i u m is f o u n d in other food products as well and is p r o b a b l y a c o m m o n b a c t e r i u m in the h u m a n environment. O u r finding in this study of a mean colonisation rate for S. saprophyticus of 69 % on the protective gloves of meat cutters t h r o u g h o u t the year supports our idea of S. saprophyticus as a ready contaminator of the food processing environment. T h e meat cutters changed protective gloves at least once every hour of work and hence there was little chance for S. saprophyticus to multiply on the gloves. Escherichia coli incubated at 37 °C is traditionally assumed to be an indicator of faecal contamination as such and the concomitant isolations of S. saprophyticus in various food and especially in raw meat samples suggest that S. saprophyticus originates from the slaughtered animal's faecal flora which readily contaminate the meat during its preparation in the slaughterhouse. Escherichia coli incubated at 44 °C is assumed to be an indicator of recent faecal
I8
P. H E D M A N E T A L .
contamination. In this study, it was d e m o n s t r a t e d only in raw meat. T h i s again suggests that S. saprophyticus originates f r o m slaughtered animals and not f r o m r e c e n t h u m a n handling. W e were interested to see w h e t h e r there was any seasonal variation in the p r e s e n c e o f S. saprophyticus in food since a seasonal variation in rectal colonisation has been described. 7 In o r d e r to c o n c e n t r a t e the food sampling on a specific seasonal food, only vegetables were collected in A u g u s t I982. T h e r e was no way o f obtaining i n f o r m a t i o n about the origin and age o f the various food specimens collected by the H e a l t h Care Council. W e t h e r e f o r e c o n d u c t e d a separate study o f S. saprophyticus c o n t a m i n a t i o n o f recently slaughtered b e e f and p o r k carcasses over I year in the food factory. As shown in Fig. I, no obvious seasonal variation in the c o n t a m i n a t i o n o f these meat specimens with S. saprophyticus was found. T h e reason w h y t h e r e is a higher degree o f h u m a n rectal colonisation o f S. saprophyticus d u r i n g late s u m m e r in S w e d e n is not clear. In a r e c e n t r e p o r t f r o m the National F o o d Council (Statens L i v s m e d e l v e r k ) , it is stated, h o w e v e r , that t h e r e is a t e n d e n c y in S w e d e n to c o n s u m e m o r e u n c o o k e d meat in various forms such as raw m i n c e d meat, raw spiced, m a r i n a t e d , lightly salted or lightly smoked steaks and lightly grilled m e a t d u r i n g the s u m m e r m o n t h s . It m a y be significant that S. saprophyticus is m o s t c o m m o n l y f o u n d in food f r o m cold storehouses and in food stored in cold and frozen conditions w h e r e raw meats are the m o s t c o m m o n foods to be stored. It is notable, also, that S. saprophyticus has been f o u n d in m e a t f r o m various countries. T h e organism is a c o s m o p o l i t a n p a t h o g e n and U T I s caused by the b a c t e r i u m have been r e p o r t e d f r o m all parts o f the world including Saudi Arabia. 19 W e p r o p o s e that S. saprophyticus should be a d d e d to the list o f c o m m o n h u m a n pathogens with a p r o b a b l e animal reservoir and for which food is a vehicle for its transmission into the h u m a n e n v i r o n m e n t .
References
L Hovelius B, Mfirdh PA. Staphylococcus saprophyticus as a common cause of urinary tract infections. Rev Infect Dis I984; 6: 328-337. 2. M~rdh PA, Hovelius B, Hovelius K, Nilsson PO. Coagulase negative novobiocin-resistant Staphylococci on the skin of animals and man, on meat and in milk. Aeta Vet Scand I978; I9 : 243-253. 3- Sommerville-Millar DA, Noble WC. Resident and transient bacteria of the skin. J Cutan Pathol I974; I: 26o-264. 4. Pead L, Maskell R. 'Micrococci' and urinary infection. Lancet 1977; ii: 565. 5- Marrie TJ, Kwan C, Noble MA, West A, Duffield L. Staphylococcus saprophyticus as a cause of urinary tract infections. J Clin Microbiol I982 ; 16:427-43 I. 6. Latham RH, Running K, Stamm WE. Urinary tract infections in young adult women caused by Staphylococcus saprophyticus. JAMA I983; 25o: 3063-3066. 7. Ringertz O, Torssander J. Prevalence of Staphylococcus saprophyticus in patients in a venereal disease clinic. Eur J Clin Microbiol I987; 5: 358-36I. 8. Kloos WE, Schleifer KH. Simplified scheme for routine identification of human Staphylococcus species. J Clin Microbiol I975 ; I: 82-88. 9. M~rdh P-A, Schleifer KH. Coagulase-negative Staphylococci. Stockholm: Almqvist and Wiksell, i986 : I 1-26. io. Kloos WE, Wolfshol JF. Identification of Staphylococcus species with the API STAPHIDENT system. J Clin Microbiol I982; I6: 5o9-516.
Staphylococcus saprophyticus in food
19
1I. Pead L, Crump J, Maskell R. Staphylococci as urinary pathogens. J Clin Pathol 1977; 30: 427-43 I. I2. Cooke EM, Shooter RA, Kumar PJ, Rousseau SA, Foulkes AJ. Hospital food as a possible source of Escherichia coli in patients. Lancet I97O; i: 436-437 . 13. Namavar F, de Graaff J, de With C, MacLaren DM. Novobiocin resistance and virulence of strains of Staphylococcus saprophyticus isolated from urine and skin. J Med microbiol 1977; H : 243-248. 14. Noble WC, Somerville DA. Microbiology of the human skin. London: WB Saunders, I974: 172-184 . 15. Kloos WE, Musselwhite MS. Distribution and persistence of Staphylococcus and Micrococcus species and other aerobic bacteria on human skin. Appl Microbiol 1975; 3o: 381-395. 16. Donzo M, Gogov I. Micro-organisms of the family Micrococceae in the production and storing of calf meat. Vet Med Nauki 1985 ; 22 : 37-4217. Marples RR, Towers AG. A laboratory model for the investigation of contact transfer of microorganisms. J Hyg 1979; 82 : 237-248. 18. Macintosh CA, Hoffman PN. An extended model for transfer of microorganisms via the hands: differences between organisms and the effect of alcohol disinfection. J Hyg 1984; 92 : 345-35519. Suraiya SK, Hisham AS, Ching L. Significance of Staph. saprophyticus as a uropathogen in adult Saudi females. Trop Doct 1987; 17: 7-8.
Staphylococcus saprophyticus contaminant
found to be a common of food
P. H e d m a n , * O. Ringertz* B. E r i k s s o n , t P. Kvarnfors,$ M. Andersson,* L. B e n g t s s o n * a n d K. Olsson*
* Department of Clinical Bacteriology, S;~dersjukhuset, The Karolinska Institute, S-IoO 64 Stockholm, ~f Goman Food Factory, Stockholm and ~ The Health Care Council, Stockholm, Sweden Accepted for publication 7 February I99o Summary The mode of transmission of Staphylococcus saprophyticus, a urinary tract pathogen, was investigated in three related studies. The presence of this organism was sought, during a period of I year, in i331 specimens of various foods, in 920 beef and pork carcasses and on Io7 cultures which had been inoculated directly from abattoir workers' protective gloves. Staphylococcus saprophyticus was found to contaminate I6"4 ~o of the various food samples with a high prevalence of 34 % in raw beef and pork. It was common in both domestic and imported raw meat products. There was no seasonal variation in the presence of S. saprophyticus in the samples obtained from carcasses. The bacterium was found in 69 % of all cultures from the workers' protective gloves. We conclude that S. saprophyticus, originating from slaughtered animals, contaminates food and eventually colonises the human intestinal tract.
Introduction
Staphylococcus saprophyticus is a well recognised c o m m o n cause of acute u r i n a r y tract infection ( U T I ) , typically present in y o u n g w o m e n in outpatient practice with a seasonal peak incidence in late s u m m e r , as reviewed by Hovelius and Mfirdh. 1 T h e reservoir for this organism and its m o d e of transmission are now known. T h e skin has been proposed as the h u m a n reservoir of S. saprophyticus which has been d e m o n s t r a t e d on both h u m a n and animal s k i n ) '3 Others, however, have f o u n d S. saprophyticus as part of the rectal flora in healthy carriers 4 and in patients with acute u r i n a r y tract infections caused by the same species. 5'6 We have recently confirmed rectal carriage of S. saprophyticus in patients in a venereal disease clinic. 7 F u r t h e r m o r e , we have d e m o n s t r a t e d a seasonal variation in the colonisation of the r e c t u m and the u r e t h r a similar to that for U T I s caused by S. saprophyticus I n the present study various food samples were investigated in order to elucidate f u r t h e r the m o d e of transmission of S. saprophyticus and to gain clues on its origin. First, a heterogenous collection of routine food samples from the H e a l t h Care Council laboratory in Stockholm was studied during a period of I year. Second, in order to concentrate the study on meat products and to verify the time of the processing of the meat, specimens of beef and pork were examined from a food factory in Stockholm, also during a period of I year. oi63-4453/9o/o4ooii +o9 $02.00/0
© I99o The British Society for the Study of Infection
I2
P. HEDMAN E T A L .
Additionally, samples were obtained from the protective gloves of workers in the meat cutting department in the same factory during the same year. Materials and methods
Food samples from the Health Care Council laboratory in Stockholm were collected from May I982 to May x983 in IO monthly periods. A total of z33I specimens was gathered by routine methods as part of the normal bacteriological public health control of food. T h e y were collected mainly from public restaurants, retail grocery shops and from stores of imported food all over the Stockholm metropolitan area. In one series of samples, only vegetables were collected. All food specimens were thoroughly homogenised in food processors and then cultured in order to demonstrate food pathogens. Duplicate cultures were incubated at 37 and 44 °C respectively. A standard protocol recorded for each sample the kind of food examined, where it was collected, its country of origin, any processing performed and how it was stored. One ml of each homogenised food specimen was incubated for 48 h at 37 °C in a S. saprophyticus selective broth consisting oftryptone (Oxoid, U.K.) of pH 7"2 with added I5o/zg/ml nalidixic acid (Winthrop, U.S.A.) and 2/zg/ml novobiocin (Sigma, U.S.A.). Subcultures were made on blood agar (Blood agar base No. 2, Oxoid, U.K.) containing 5 % defibrinated horse blood. All Staphylococcus species were further tested for their susceptibility to novobiocin by an agar diffusion method with discs containing 5 #g novobiocin (Biodisk, Sweden). Strains demonstrating an inhibitory zone of less than I5 m m were further classified according to a modification of Kloos and Schleifer 8 in serial tube reactions (Voges-Proskauer test, urease production, nitrate reduction, anaerobic production of acid from glucose and fermentation of arabinose, mannose, xylose, sucrose and glucose). All strains of Staphylococcus species were also tested according to API Staph-Ident (Analytab Products, Plainview, N.Y., U.S.A.). Meat specimens from a food factory were collected from the cold store room in the Konsum Stockholm Goman factory from November 1985 to October I986. All meat was delivered by cold transportation from various slaughterhouses all over Sweden and reached the cold room within 3 days after slaughtering the animals and cutting the carcasses into halves or quarter pieces. T w e n t y samples of beef and pork were gathered every fortnight except for January I 9 8 5 when only 20 samples of each were collected. All sampling was sought to include a few slaughterhouses as possible. Altogether, 460 specimens of beef were cut from the neck of the carcasses in thin slices with a diameter of 3-5 cm by means of sterile disposable scalpels and forceps. All 460 pork specimens were cut in a similar way from the shoulder and neck of the pig carcasses. T h e specimens were incubated for 48 h at 37 °C in the same selective broth as described above. Isolation of S. saprophyticus followed the same procedures. T h e workers in the cutting room, close to the cold store in the food factory, used cotton gloves strengthened by thin steel wires for protection. T h e gloves were dipped in the previously described selective broth for S. saprophyticus (held in single plastic containers) for 30 s on four occasions during the study. T h e containers were incubated and subcultured as described above.
Staphylococcus saprophyticus in food
13
Table I The isolation rates (%) of Staphylococcus saprophyticus in food
samples collected by the Health Care Council in Stockholm from May I982 to May I983 Sample
I 2 3 4 5 6 7 8 9 Io Total
Date
I982 May June July August October December 1983 January March April May
Number of specimens
I84 I53 I 15 43* I7o 97
S. saprophyticus (%)
z5 14 II 5 24 25
2oi
16
136
2I
II6 I I6 I331
IO 13 I6-4
* Only vegetables collected.
All statistical analyses were done by means of X2 m e t h o d s unless otherwise stated. RESULTS Food collected by the Health Care Council of Stockholm
A total of I33I samples was examined. T h e zo sampling periods and the seasonal presence of S. saprophyticus are shown in T a b l e I. Altogether, z i 8 strains of S. saprophyticus were isolated and identified by means of the tube tests described above, so giving an overall prevalence rate of I6"4 % . T h e A P I S t a p h - I d e n t system identified 2 z I (96"8 %) of these strains as S. saprophyticus. T h e I33Z collected food specimens were heterogeneous. T h e y included raw food items such as freshly cut meat and fresh vegetables from cold storehouses, r e a d y - m a d e foods such as sandwiches sold in retail shops and cooked food from restaurant kitchens. T h e types of food specimens are grouped and the presence of S. saprophyticus in each type of food is presented in Table II. M o s t meat products consisted of pork and beef. A m o n g these 6oi items of meat, there were 27o samples of raw beef or pork. As m a n y as 34 % of these specimens were contaminated with S. saprophyticus. T h e r e were 12 samples in this study of raw meat collected from a retail shop within a slaughterhouse area. T e n of these samples contained S. saprophyticus. T h e r e were 38 specimens of ice-cream a m o n g the 68 dairy food samples but none of these contained S. saprophyticus. M o s t items of confectionery were r e a d y - m a d e pastries from retail bakeries and hence were composed of various food materials. M o s t items of poultry were freshly grilled chicken collected in retail shops. T h e r e were only four specimens of egg. One sample of boiled egg contained S. saprophyticus. T h e group of mixed groceries consisted of readymade complete courses of mixed food, sandwiches, salads and sauces. All food samples were collected from various locations g r o u p e d and specified in T a b l e
P. H E D M A N E T A L .
I4
Table II The frequency of Staphylococcus saprophyticus in various types of
food Specimens from which
S. saprophyticus was isolated Type of food Raw meat Processed meat Poultry and eggs Fish Dairy products Vegetables Bread Mixed groceries Total
N u m b e r of specimens
(N)
(%)
270 331 34 6I 68 205 63 299
92 70 8 8 5 I2 3 20
34 2i 24 13 7 6 5 7
i33i
218
I6'4
This distribution was not due to chance (X2 = 98"I6, P < o-ooI, six degrees of freedom).
III. Sources of food available to the public included public restaurants of various sizes as well as canteens for schoolchildren, retired people and staffs of various companies. There were 71 specimens of imported meat in this study. Most of these were of raw beef and came from few countries as specified in Table IV. There were 48 (36 %) specimens of raw beef contaminated with S. saprophyticus out of a total of I36 of Swedish origin. Imported beef was not significantly more contaminated (P = o'I3; Fisher's exact test). Information on the storage of the food items was available in I o7o (8o %) specimen records. Details are given in Table V. Specimens of heated food consisted of food served hot in restaurants. Only one of these 4o samples contained S. saprophyticus. No other significant human pathogen was isolated from the x331 food specimens. There were, however, 435 specimens incubated at 37 °C which yielded significant growth of Escherichia coli. Staphylococcus saprophyticus was isolated from Io4 of these and hence was found to be a significant (X2 = 25"9; P < o'oI) concomitant contaminator. As many as 53% of all specimens with growth of both species were samples of raw meat. On the other hand, there was no tendency (X~= o'8; P > o'o5) to find S. saprophyticus together with E. coli in samples cultured at 44 °C. Of 95 specimens incubated at 44 °C and found to be contaminated with E. coli, only I2 had concomitant growth of S. saprophyticus. No particular food was represented among these 95 specimens incubated at 44 °C and contaminated by E. coli. T h e specimens concomitantly contaminated with S. saprophyticus, however, were more often products of raw meat (P < o.oi, Fisher's exact test).
Staphylococcus saprophyticus
in f r e s h c a r c a s s e s f r o m t h e f o o d f a c t o r y
Among the 460 specimens of fresh beef, z24 (27 %) were contaminated with S. saprophyticus and r37 (30 %) of the 460 pork samples were found to contain the same bacterium.
Staphylococcus saprophyticus Table III
in food
I5
The origin of the
I 3 3 I food samples and the frequency Staphylococcus saprophyticus
isolation of
Staphylococcus saprophyticus Sample origin Restaurants and canteens Retail grocery shops Cold storehouses Wholesale stores No information Total Table IV
Number of specimens
(N)
(%)
575
57
IO
423
98
23
95 54 184 1331
37 4 22 218
39 7 12 16'4
Country of origin of 72 samples of imported beef and the frequency of isolation of S t a p h y l o c o c c u s s a p r o p h y t i c u s Staphylococcus saprophyticus Country Poland Australia Jugoslavia U.S.A. Total
Table V
Number of specimens
(N)
(%)
27 z5 18 1 71
7 8 16 I 32
26 32 89 45
Storage of food and the frequency of isolation of
Staphylococcus
saprophyticus
Staphylococcus saprophyticus Type of storage Cold Room temperature Frozen Heated No information Total
Number of specimens
(N)
(%)
755 I6I I I4 40 261 i331
I23 19 35 I 40 218
I6 I2 3i 15 I6'4
T h e r e l a t i v e d i s t r i b u t i o n s o f t h e g r o w t h o f S. saprophyticus in f r e s h b e e f a n d p o r k f o r e a c h m o n t h o f t h e y e a r are d e p i c t e d in F i g . I. T h e r e w e r e n o o b v i o u s seasonal trends.
Staphylococcus saprophyticus
on meat cutters' protective gloves
O n f o u r o c c a s i o n s , in N o v e m b e r I 9 8 5 , in F e b r u a r y I 9 8 6 , in M a y I 9 8 6 a n d in A u g u s t I 9 8 6 , a t o t a l o f IO7 s p e c i m e n s w a s c o l l e c t e d f r o m p r o t e c t i v e gloves. As
P. HEDMAN ET AL.
I6 75
50-r-n
I I -
i
25
FI
-i
o
d
NOV DEC
JAN
FEB
MAR APR MAY
dUN
JUL
AUG SEP
OCT
1985-1986 F i g . [. T h e i s o l a t i o n o f Staphylococcus saprophyticus i n b e e f a n d p o r k c o m p a r e d w i t h t h e relative distribution of urinary tract infectionsdue to the same bacterium during a period of I2 months as diagnosed at the Laboratoryof Bacteriology, S6dersjukhuset, Stockholm. (-), UTI; (I~), beef; ([]), pork.
m a n y as 74 (69 %) were positive for S. saprophyticus. T h e r e was no obvious seasonal distribution of these isolates. Discussion
T h e correct identification of S. saprophyticus is crucial for our study. O u r procedure is based on the classification o f Staphylococcus species presented by Kloos and Schleifer in z9758 and u p d a t e d by M ~ r d h and Schleifer in I986. 9 T h e serial tube reaction was designed to separate S. saprophyticus specifically f r o m the two closely related novobiocin-resistant species o f S. cohnH and S. xylosus. All the strains of S. saprophyticus isolated f r o m food samples were f u r t h e r tested by A P I S t a p h - I d e n t , an accepted 1° commercial test system. T h e two m e t h o d s agreed well. A m o n g the seven strains of Staphylococcus species that did not agree, the A P I S t a p h - I d e n t identified three strains o f novobiocinresistant S. sirnulans ,°.nd one strain o f novobiocin-resistant S. warneri. A P I S t a p h - I d e n t failed to identify three strains. T w o presumptive ideas derive from our study on the presence of S. saprophyticus in food. First, we assume that the h u m a n reservoir of this specific u r i n a r y tract p a t h o g e n is the patient's faecal flora and, second, we assume that colonisation of the r e c t u m originates from contaminated food. We f o u n d in an earlier report 7 a mean rectal carriage rate of S. saprophyticus of 2"6 % a m o n g 495 m e n and of 2"5 % a m o n g 433 women, all attending a venereal disease clinic d u r i n g the period of I year. In that study, there was a clear t e n d e n c y for a seasonal variation of the rectal carriage. T h e m a x i m u m prevalence rate of S. saprophyticus in the r e c t u m of 6"7 % preceded the m a x i m u m prevalence rate in
Staphylococcus saprophyticus in food
17
the urethra of 9-2 % by I month. Other studies have reported rectal carriage of S. saprophyticus. Pead and Maskell 4 f o u n d S. saprophyticus in the faecal flora among 6"4 % of I56 healthy y o u n g w o m e n attending a family-planning clinic. Marrie et al., ~ f o u n d the same b a c t e r i u m in the faecal flora o f two of nine college w o m e n with an acute U T I due to the same species. Finally, L a t h a m et al., 6 found S. saprophyticus in the r e c t u m of three of 27 w o m e n attending a student health centre. T h e r e c t u m is considered to be the h u m a n reservoir for U T I s caused b y S. saprophyticus in two papers. 11'6 In a series of studies, 12 Cooke et al., d e m o n s t r a t e d the route of transmission of the urinary pathogen E. coli from various hospital foods via the intestinal tract of the hospital patients. W e propose a similar m o d e of transmission for S. saprophyticus. T h e r e are reports of S. saprophyticus being isolated from various sites on the h u m a n skin 2' 13 and the skin has been p r o p o s e d as the h u m a n reservoir of S. saprophyticus infections. ~ W e assume that colonisation of h u m a n skin is due to faecal contamination in a way similar to that for various G r a m - n e g a t i v e Enterobacteriaceae which are often isolated from the h u m a n skin.14 W e agree that S. saprophyticus should be classified as a transient coloniser of the h u m a n skin.3. ,5 T h e overall prevalence rate of S. saprophyticus of 16"4 % in the heterogenous material of 1331 food specimens is high. Cooke et al., 12 f o u n d a prevalence rate of contaminating E. coli in hospital food to be less than IO %. In s u b g r o u p i n g all food samples there was a marked p r e p o n d e r a n c e of S. saprophyticus in animal p r o d u c t s and especially in raw b e e f and pork where the prevalence rate of S. saprophyticus was as high as 34 %. T h e presence of S. saprophyticus in meat has been reported before. 2' 16 T h e high prevalence of S. saprophyticus in chicken meat suggests a future s t u d y of chickens' faecal flora. It is worth noting that S. saprophyticus was not isolated from ice-cream which is a typical food during the high seasonal prevalence of U T I s caused b y the bacterium. Staphylococcus saprophyticus has been f o u n d to be capable of transfer from hand to hand and from hand to fomites in experimental studies, iv,is Staphylococcus saprophyticus transferred more readily than Streptococcus pyogenes and E. coli and other Enterobacteriaceae. W e consider that the high prevalence of S. saprophyticus in one c o m m o n food such as raw meat, together with its ability to transfer easily on handling, explains w h y the b a c t e r i u m is f o u n d in other food products as well and is p r o b a b l y a c o m m o n b a c t e r i u m in the h u m a n environment. O u r finding in this study of a mean colonisation rate for S. saprophyticus of 69 % on the protective gloves of meat cutters t h r o u g h o u t the year supports our idea of S. saprophyticus as a ready contaminator of the food processing environment. T h e meat cutters changed protective gloves at least once every hour of work and hence there was little chance for S. saprophyticus to multiply on the gloves. Escherichia coli incubated at 37 °C is traditionally assumed to be an indicator of faecal contamination as such and the concomitant isolations of S. saprophyticus in various food and especially in raw meat samples suggest that S. saprophyticus originates from the slaughtered animal's faecal flora which readily contaminate the meat during its preparation in the slaughterhouse. Escherichia coli incubated at 44 °C is assumed to be an indicator of recent faecal
I8
P. H E D M A N E T A L .
contamination. In this study, it was d e m o n s t r a t e d only in raw meat. T h i s again suggests that S. saprophyticus originates f r o m slaughtered animals and not f r o m r e c e n t h u m a n handling. W e were interested to see w h e t h e r there was any seasonal variation in the p r e s e n c e o f S. saprophyticus in food since a seasonal variation in rectal colonisation has been described. 7 In o r d e r to c o n c e n t r a t e the food sampling on a specific seasonal food, only vegetables were collected in A u g u s t I982. T h e r e was no way o f obtaining i n f o r m a t i o n about the origin and age o f the various food specimens collected by the H e a l t h Care Council. W e t h e r e f o r e c o n d u c t e d a separate study o f S. saprophyticus c o n t a m i n a t i o n o f recently slaughtered b e e f and p o r k carcasses over I year in the food factory. As shown in Fig. I, no obvious seasonal variation in the c o n t a m i n a t i o n o f these meat specimens with S. saprophyticus was found. T h e reason w h y t h e r e is a higher degree o f h u m a n rectal colonisation o f S. saprophyticus d u r i n g late s u m m e r in S w e d e n is not clear. In a r e c e n t r e p o r t f r o m the National F o o d Council (Statens L i v s m e d e l v e r k ) , it is stated, h o w e v e r , that t h e r e is a t e n d e n c y in S w e d e n to c o n s u m e m o r e u n c o o k e d meat in various forms such as raw m i n c e d meat, raw spiced, m a r i n a t e d , lightly salted or lightly smoked steaks and lightly grilled m e a t d u r i n g the s u m m e r m o n t h s . It m a y be significant that S. saprophyticus is m o s t c o m m o n l y f o u n d in food f r o m cold storehouses and in food stored in cold and frozen conditions w h e r e raw meats are the m o s t c o m m o n foods to be stored. It is notable, also, that S. saprophyticus has been f o u n d in m e a t f r o m various countries. T h e organism is a c o s m o p o l i t a n p a t h o g e n and U T I s caused by the b a c t e r i u m have been r e p o r t e d f r o m all parts o f the world including Saudi Arabia. 19 W e p r o p o s e that S. saprophyticus should be a d d e d to the list o f c o m m o n h u m a n pathogens with a p r o b a b l e animal reservoir and for which food is a vehicle for its transmission into the h u m a n e n v i r o n m e n t .
References
L Hovelius B, Mfirdh PA. Staphylococcus saprophyticus as a common cause of urinary tract infections. Rev Infect Dis I984; 6: 328-337. 2. M~rdh PA, Hovelius B, Hovelius K, Nilsson PO. Coagulase negative novobiocin-resistant Staphylococci on the skin of animals and man, on meat and in milk. Aeta Vet Scand I978; I9 : 243-253. 3- Sommerville-Millar DA, Noble WC. Resident and transient bacteria of the skin. J Cutan Pathol I974; I: 26o-264. 4. Pead L, Maskell R. 'Micrococci' and urinary infection. Lancet 1977; ii: 565. 5- Marrie TJ, Kwan C, Noble MA, West A, Duffield L. Staphylococcus saprophyticus as a cause of urinary tract infections. J Clin Microbiol I982 ; 16:427-43 I. 6. Latham RH, Running K, Stamm WE. Urinary tract infections in young adult women caused by Staphylococcus saprophyticus. JAMA I983; 25o: 3063-3066. 7. Ringertz O, Torssander J. Prevalence of Staphylococcus saprophyticus in patients in a venereal disease clinic. Eur J Clin Microbiol I987; 5: 358-36I. 8. Kloos WE, Schleifer KH. Simplified scheme for routine identification of human Staphylococcus species. J Clin Microbiol I975 ; I: 82-88. 9. M~rdh P-A, Schleifer KH. Coagulase-negative Staphylococci. Stockholm: Almqvist and Wiksell, i986 : I 1-26. io. Kloos WE, Wolfshol JF. Identification of Staphylococcus species with the API STAPHIDENT system. J Clin Microbiol I982; I6: 5o9-516.
Staphylococcus saprophyticus in food
19
1I. Pead L, Crump J, Maskell R. Staphylococci as urinary pathogens. J Clin Pathol 1977; 30: 427-43 I. I2. Cooke EM, Shooter RA, Kumar PJ, Rousseau SA, Foulkes AJ. Hospital food as a possible source of Escherichia coli in patients. Lancet I97O; i: 436-437 . 13. Namavar F, de Graaff J, de With C, MacLaren DM. Novobiocin resistance and virulence of strains of Staphylococcus saprophyticus isolated from urine and skin. J Med microbiol 1977; H : 243-248. 14. Noble WC, Somerville DA. Microbiology of the human skin. London: WB Saunders, I974: 172-184 . 15. Kloos WE, Musselwhite MS. Distribution and persistence of Staphylococcus and Micrococcus species and other aerobic bacteria on human skin. Appl Microbiol 1975; 3o: 381-395. 16. Donzo M, Gogov I. Micro-organisms of the family Micrococceae in the production and storing of calf meat. Vet Med Nauki 1985 ; 22 : 37-4217. Marples RR, Towers AG. A laboratory model for the investigation of contact transfer of microorganisms. J Hyg 1979; 82 : 237-248. 18. Macintosh CA, Hoffman PN. An extended model for transfer of microorganisms via the hands: differences between organisms and the effect of alcohol disinfection. J Hyg 1984; 92 : 345-35519. Suraiya SK, Hisham AS, Ching L. Significance of Staph. saprophyticus as a uropathogen in adult Saudi females. Trop Doct 1987; 17: 7-8.