Differential Diagnosis Between Streptococcus Agalactiae and Listeria Monocytogenes in the Clinical Laboratory

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Differential Diagnosis between Streptococcus Agalactiae and Listeria Monocytogenes in the Clinical Laboratory

CHRISTINE KONTNICK, M.T., ALEXANDER von GRAEVENITZ, M.D., and VINCENT PISCITELLI, M.T.

Clinical Microbiology Laboratories, Yale-New Haven Hospital, and Department of Laboratory Medicine, Yale University School of Medicine, New Haven, CT 06504

ABSTRACT Streptococci of the group B (S. agalactiae) and Listeria monocytogenes resemble each other in many morphological and biochemical characteris tics. Ten beta-hemolytic strains of each species were subjected to 26 tests commonly and easily performed in the clinical laboratory. Macroscopic and microscopic morphology on solid media showed differences only in the size of the colonies and in the length of the individual organisms. Among many other tests, hippurate hydrolysis and the CAMP reaction were pos itive in both species. In the presence of these two reactions, a negative catalase test and chaining in broth would make a presumptive diagnosis of S. agalactiae, while motility at 25 C, the presence of the Henry effect, and resistance to furadantin would be indicative of L. monocytogenes.

Introduction in Gram-stained smears; (3) a negative bacitracin test and (4) a positive test for The high incidence of Streptococcus either (a) hippurate hydrolysis, (b) the agalactiae (group B) in human speci CAMP reaction or (c) the formation of an mens, which has been recognized only in orange-red pigment. However, if one of the past decade, calls for a rapid pre these conditions for the diagnosis is not sumptive diagnosis of the species. Many met, differential diagnostic problems clinical laboratories, at least the small may arise with other Gram-positive or ones, prefer not to perform serological ganisms. This is illustrated by the follow tests (CIE, coagglutination) for the iden ing facts: tification of streptococci because of the expenses in equipment, time and money (1). A minority of S. agalactiae strains involved. For them, the presumptive is nonhemolytic on sheep or rabbit blood diagnosis of S. agalactiae usually rests agar even under anaerobic condi with the combination of: (1) beta- tions.18,45 On the other hand, colonies of hemolytic colonies on sheep or rabbit beta-hemolytic Gram-positive organisms blood agar; (2) appearance of streptococci like Corynebacterium haemolyticum, 2 7 0 K ON TN ICK, v o n GRAEVENITZ AND PISCITELLI

Listeria monocytogenes and other beta- served in S. agalactiae but this charac hemolytic streptococci may closely re teristic is highly dependent on semble those of S. agalactiae. L. anaerobiosis and the medium used; Col monocytogenes colonies41 tend to be umbia agar being the optimal one.16,34 It smaller in size than those of S. agalac may be absent in 3 percent of the beta- tiae,6 but the hemolytic zone of both is hemolytic34 and in some of the nonhemo usually small. lytic strains.16 Some L. monocytogenes (2). Chain formation of streptococci is strains are also said to produce a yel seen best in Gram-stained smears from lowish pigment after several months of liquid media1 and may be poor in Gram incubation.42 C. poinsettiae, a motile stains from solid media. Thus, coccoid Corynebacterium, also forms an orange Gram-positive organisms on solid media pigment.10 may be streptococci, but may also be It is thus obvious that one Gram- young listeriae10 (which may even form positive rod, L. monocytogenes, shares short chains).41 In broth, however, S. many features with S. agalactiae. The agalactiae usually forms long chains.17 possibility of a misdiagnosis is further (3). The bacitracin test is positive in 4 enhanced by the fact that both species to 8 percent of S. agalactiae, streptococci tend to be isolated mainly from the cer of groups other than A, B, and D, S. bovis, vix, from blood, and from the spinal fluid S. viridans and in about 48 percent of S. (although group B wound infections are pneumoniae; however, it is negative in not uncommon). In fact, L. monocyto about 0.5 percent of S. pyogenes.15 genes tends to be most often misdiag (4a). The hippurate test is reported nosed as a streptococcus species.10,20,41 positive among Gram-positive bacteria The smear of a liquid sample or culture regularly in S. bovis and S. acidomin- would immediately differentiate the two, imusK in L. monocytogenes (but not in but if a Gram stain is done either from a other Listeria species),14,15* in certain specimen that contains other streptococ species of Lactobacillus39 and Coryne- ci from a plate, or if no Gram stain is done bacterium7 and in about 6.9 percent of at ail (as is often the case on cervical enterococcus strains.15 smears), a misdiagnosis may ensue. (4b). The CAMP test36 is positive in S. The present investigation was promp pyogenes (group A) under anaerobic con ted by a routine search for S. agalactiae ditions,11 but may yield equivocal results in cervical specimens in the course of under aerobic conditions.32 It has also which a strain of L. monocytogenes was been reported positive (aerobically) in 12 discovered. It was restricted to 26 com to 20 percent of group G, K, and L strep monly and easily performed tests, not in tococci,24 in certain Corynebacterium cluding macroscopic and microscopic species,19 and in virulent strains of L. morphology. Other tests did not appear monocytogenes.3,8,19,23 It may be nega feasible for routine purposes, as will be tive in S. agalactiae if the sheep blood pointed out in the discussion. used contains staphylococcal beta anti toxin and has not been washed,36 and may Materials and Methods also be negative in nonhemolytic strains Ten strains each of S. agalactiae and L. of group B.12,35 monocytogenes were examined. The S. (4c). Production of an orange-red pig agalactiae strains had been isolated from ment in streptococci has only been ob cervical specimens in this laboratory and were grouped by the Connecticut State * In the abstract of this paper,28 five of five L. monocytogenes strains were erroneously described Department of Health. Seven L. as “failing to hydrolyze hippurate.” monocytogenes strains had been isolated STREPTOCOCCUS AGALACTIAE VS. LISTERIA MONOCYTOGENES 2 7 1

TABLE I

Tests and Methods

Test Medium Method Positive Control Negative Control

Arginine dihydrolase Decarboxylase Falkow1 Arizona hinshawii E. coli medium base* Bile-esculin (growth & Bile-esculin Swan1 S. faecalis E. coli esculin hydrolysis) agar* D a rl i ng 12CAMP reaction Tryptic soy agar Darling12CAMP S. agalactiae E. coli (aerobic) with 5% sheep blood* Catalase Tryptic soy broth* Broth1 E. coli 5. faecalis Citrate utilization Koser citrate Koser29 Klebsiella pneumoniae E. coli medium* Deoxyribonuclease DNase test agar von Graevenitz Serratia marcescens E. coli with methyl green* (24 & 48 hrs) Gelatin hydrolysis Photographic Lagodsky & S. marcescens E. coli X-ray strip J a n d a r d ^ Gluconate oxidation Tabletst Haynes1 Pseudomonas aeruginosa P. maltophilia Beta hemolysis Tryptic soy agar ~ S. pyogenes S. faecalis with 5% sheep blood* Henry effect Mueller-Hinton agar* Gray L. monocytogenes E. coli Hippurate hydrolysis - Hwang & Ederer S. agalactiae E. coli Hydrogen sulfide PathoTec strip§ Standard4 A. hinshawii E. coli Indole Trypticase*0 Ehrlich-Böhme1 E. coli Enterobacter aerogenes Lipase Spirit blue agar Hugo & S. marcescens E. coli with tributyrin* Beveridge2^ Lysine decarboxylase PathoTec strip§ Standard4 E. aerogenes Citrobacter diversus Malonate utilization Malonate broth* Leifson1 C. diversus E. coli Methyl red reaction MR-VP medium* Clark & Lubs1 E. coli K. pneumoniae Motility 25C Tryptic soy broth* Hanging drop1 E. coli K. pneumoniae Nitrate reducation Trypticase nitrate Standard1 E. coli S. faecalis brothw ONPG Tabletst Standard1 E. coli Salmonella sp. Ornithine decarboxylase PathoTec stripi Standard4 E. aerogenes Proteus vulgaris

Oxidase - Ko va c s 1 P. aeruginosa E. coli Phenylalanine Urea-phenyalanine Ederer et al13 P. vulgaris E. coli deaminase medium* Mueller-Hinton agarw Kirby-Bauer1Antimicrobial E. coli ATCC 25922 - susceptibility Urea-phenyalanine Ederer et a l 1^Urease P. vulgaris E. coli medium* MR-VP medium* Coblentz1Voges-Proskauer K. pneumoniae E. coli reaction (37°C, 48 hrs) Voges-Proskauer PathoTec strip§ Standard4 K. pneumoniae E. coli

*Difco Laboratories, Detroit, MI tKey Scientific Products, Los Angeles, CA §General Diagnostics Div., Warner-Lambert Company, Morris Plains, NJ UBBL, BioQuest, Cockeysville, MD from blood cultures here, and three were both Tryptic Soy Brothf and from blood received from the Bacteriology Labora agar after 24 hours of growth. In table I tory at the Massachusetts General Hospi are listed tests, methods and control tal, Boston. strains used. Incubation was at 37 for 24 Colonial morphology was observed on hours unless indicated otherwise, e.g., by blood agar (Tryptic Soy Agarf with 5 per the manufacturer of test strips or tablets. cent sheep blood) after incubation for 24 to 48 hours at 37° in 7.5 percent C02. Results Gram-stained smears were prepared from The colonial morphology of the strains t Difco Laboratories, Detroit, MI. was similar except for the larger diame- 2 7 2 K ON TN ICK, v o n GRAEVENITZ AND PISCITELLI

TABLE I I

Results (Positive Strains per Total Strains Tested)

Results References S. agalactiae L. monocytogenes S. agalactiae L. monocytogenes

17 Arginine dihydrolase 10/10 0/10 3 ,44 15 ,17 ,27 27 Bile-esculin (growth & hydrolysis) 0/10 10/10 10 ,24 ,35 ,45 3,8,19 ,23 CAMP reaction 10/10 10/10 1,10 2327 Catalase 0/10 10/10 3, 5, 9,10,41 ,^2,^ 3.44 Citrate utilization (24 hrs) 0/10 0/10 Deoxyribonuclease (24 hrs) 0/10 0/10 3 Deoxyribonuclease (48 hrs) 3/10 0/10 3,5,40,41 ,44 Gelatin hydrolysis 0/10 0/10 3 Gluconate oxidation 0/10 0/10 21.41.42 Henry effect 0/10 10/10 Hippurate hydrolysis 10/10 10/10 1/6,12/15,34,35,45 14,15,28 3.5.41.42 Hydrogen sulfide 3/10 0/10 3.5.41 Indole 0/10 0/10 3,41,42,43 Lipase 10/10 10/10 3,5 Lysine decarboxylase 0/10 0/10 3 ,44 Malonate utilization 0/10 0/10 3.44 Methyl red reaction 10/10 10/10 17 1.3.31.42.44 Motility 25° C 0/10 10/10 1 ,17,35 3.5.41.42.44 Nitrate reduction 0/10 0/10 1,38 3.44 ONPG 0/10 0/10 3.5.44 Ornithine decarboxylase 0/10 0/10 3/5,41,44 Oxidase 0/10 0/10 3 ,44 Phenylalanine deaminase 0/10 0/10 Susceptibility: 9.31.41.42 Colistin 0/10 0/10 33 Furadantin 10/10 0/10 2 Nalidixic acid 0/10 0/10 5.9.41.44 Aminoglycosides 0/10 10/10 17, 35 3.5.41.44 Urease 0/10 0/10 17 3.5.41 42 44 VPR - Coblentz 10/10 10/10 - PathoTec strip 10/10 10/10 ters of the S. agalactiae colonies which In table II are listed the results of tests were about twice the size of the L. for identification and references. monocytogenes colonies of equal age. Colonies of both species were grayish- Discussion white and showed a small zone of hazy beta hemolysis. In broth, the growth of S. On the basis of our and other results, agalactiae tended to concentrate on the the following tests would lend them bottom of the tube in the form of a pellet selves for a differentiation between S. whereas L. monocytogenes caused fairly agalactiae and L. monocytogenes: micro uniform turbidity. scopic morphology from broth, arginine Microscopically, chains of 10 to 20 or dihydrolase, bile-esculin, catalase, the ganisms in broth and short chains on Henry effect, motility at 25°, and suscep blood agar were seen in S. agalactiae, but tibility to aminoglycosides and nitrofu no or only occasional short chaining was rantoin. This presupposes that other bac observed in L. monocytogenes which ap teria have already been ruled out by peared in the form of coccoid (24 hours) Gram-stained smears, colony morphology to medium size (48 hours) rods with occa (both species may be nonhemoly sional palisading and formation of Vs. tic),3,18,20,23,44,45 hippurate, CAMP, or These findings correspond to those re pigmentation tests, and that Gram- ported in the literature. 41,42 positive bacteria which may give similar STREPTOCOCCUS AGALACTIAE VS. LISTERIA MONOCYTOGENES 2 7 3

results in the latter three tests (see Intro like, but more semicircular to rec duction) were ruled out as well. tangular toward the staphylococcus A review of the reliability of the tests streak.8,23 Nonvirulent L. monocy mentioned for the differential diagnosis togenes strains are CAMP nega between S. agalactiae and L. monocyto tive.8,23 genes reveals the following. The catalase 2. The deoxyribonuclease test on the and the motility tests have been most fre classical test plate1 has been re quently recommended.5,10 Both require ported positive in 58 of 137 (42.5 special precautions (no transfer of blood percent) S. agalactiae strains17 and and incubation at 25° and 37°, respec in 15 of 15 L. monocytogenes tively), and have been reported negative strains.3 The tubed medium, using in subcultures of L. monocytogenes.20 methyl green and no hydrochloric The Henry effect, i.e., the appearance of acid43 failed to detect depolymeriza a blue-green iridescence ofL. monocyto tion in L. monocytogenes even after genes colonies when observed by ob 48 hours incubation. lique trans-illumination, requires a 3. The ONPG test was found negative blood-free agar.21 One group has ob by us and others3 for L, monocyto served Henry-negative L. monocyto genes, whereas 7 of 7 L. monocyto genes stains among those kept for refer genes strains were found ONPG ence purposes and has seen Henry- positive by other authors.44 These positive streptococci.20 The Bile-Esculin differences are unexplainable by us. Agarf seems excellent as long as en- (TSI was the growth medium.) terococci have been ruled out.27 Sensi 4. The lipase test using tributyrin was tivity to aminoglycosides would rule out positive for both species. Others S. agalactiae,17,34 but L. monocytogenes have found the same3 and opposite44 strains resistant to streptomycin31 and results for L. monocytogenes. A kanamycin5,9 have been observed. Sus tween esterase has been found lack ceptibility to nalidixic acid and to nitro ing in S. agalactiae.17 furantoin has, to our knowledge, never In table III are listed tests that could been systematically investigated in also be used for differential diagnostic either species but the use of nalidixic purposes but that are impractical in either acid for the isolation of both2 and the use using too much time or employing media of nitrofurantoin for the isolation of L. not immediately available in a routine monocytogenes33 imply resistance. laboratory. To them one could possibly Our results are in agreement with those add the indoxylbutyrate reaction which is of the authors quoted in table II with the positive in L. monocytogenes and nega following exceptions: tive in streptococci38 although S. agalac 1. The CAMP test is generally positive tiae is not specifically mentioned in this in S. agalactiae with the rare excep report. In table IV it is shown that other tions mentioned previously (two tests give either consistently positive or nonhemolytic strains recently iso consistently negative results for both lated in our laboratory were CAMP species, or that they are variable in both. positive). In our strains of L. A few are variable in one species and monocytogenes, however, the area constant in the other one. They should be of indicative hemolysis was smaller used for differential purposes only when than in S. agalactiae3 and not arrow- a result is observed that is known to occur in one of the two species exclusively. f Difco Laboratories, Detroit, MI. Some sugar fermentations in L. mono- 2 7 4 KO N TN ICK , v o n GRAEVENITZ AND PISCITELLI

TABLE I I I

A dditional Possible Tests fo r the D iffe re n tia tio n between S. agalactiae and L. monocytogenes

Results References One Day Growth in Media with: S. agalactiae L. monocytogenes S. agalactiae L. monocytogenes

17 0.05 percent - 1 percent KCN 5,44 35 5,22 0.04 percent K 2 Te 0 3 17 22 0.03 percent NaN 3 17 41 40 percent NaCl 17 41 Growth at 10° C cytogenes may take several days and are colonial morphology (including hemol difficult to reproduce.3 ysis or lack thereof), Gram stain, hip- In the clinical laboratory, the most purate, CAMP, or pigment tests can be specific and most easily available tests used. The latter is probably most specific should be used for the diagnosis of S. if positive. The others, if positive, still agalactiae or L. monocytogenes. For a leave the differential diagnosis between preliminary exclusion of most other or S. agalactiae, L. monocytogenes, some ganisms following initial plate growth, corynebacteria and lactobacilli, and a few

T A B L E I V

A dditional Tests Not P ractical fo r D iffe r e n tia l Diagnosis between S. agalactiae and L. monocytogenes

Results References S. agalactiae L. monocytogenes S. agalactiae L. monocytogenes

Acids from carbohydrates * 17 3 ,44 Mouse pathogenicity +t 37 3 (l d 50i o 7) (LD50102-107) Rabbit eye pathogenicity vt 3 17,35 Susceptibility to penicillin, So far generally So far generally 5,9 ampicillin, erythromycin, sensitive sensitive lincomycin, chloramphenicol, tetracycline Growth in broth with 6.5% NaCl V + 6,15 ,17 41 Growth at 45° C. 17 41 Reduction of 0.01 % methylene - 17 41 blue milk Growth, initiation at pH 9.6 17 41 pH in glucose broth <5.0 + + 17 41 Resistance to 60° C. for 30 min _ 17 41 Reduction of TTC V V 17 41 Double zone hemolysis V _ 14 41 Dextran formation in 5% sucrose agar - 17 3 Lecithinase V + 17 3W ,43 Phosphatase V + 17 43 Bacitracin test -/(+) _ 15 ,16 41 Growth in SF broth 15 41 Casein hydrolysis V 17 41 41 Guanine-cytosine ratio (moles %) 33-42 38 §

*Both are (1) positive for glucose, maltose, trehalose and glycerol; (2) negative for raffinose, ara- binose, mannitol, dulcitol, adonitol and inulin; and (3) variable for lactose and starch. L. monocytogenes is variable and S. agalactiae is negative for xylose, rhamnose and sorbitol. L. monocytogenes is positive and S. agalactiae is variable for salicin. L. monocytogenes is variable and S. agalactiae is positive for sucrose. tOnly beta hemolytic strains are positive.3 §These data are for streptococci in general. No data are available for S. agalactiae. wBeta-hemolytic strains only. The test method was identical; the time of incubation is not mentioned.3 STREPTOCOCCUS AGALACTIAE VS. LISTERIA MONOCYTOGENES 2 7 5 other streptococci (the latter three are monocytogenes infections. With a report of ten under consideration if no beta-hemolysis cases. Amer. J. Med. 45:904-921, 1968. 10. Center for Disease Control: Proficiency is encountered). A negative catalase test Testing-Bacteriology II, July 1974. Atlanta, and chaining in broth would rule out L. Center for Disease Control, 1975. monocytogenes while making the pre 11. C h r is t e n s e n , P., Ka h l m e t e r , G ., J o n s s o n , S., and K r o n v a l l , G.: New method for the sumptive diagnosis of S. agalactiae most serological grouping of streptococci with likely (only nonhemolytic strains would specific antibodies adsorbed to protein need further tests). L. monocytogenes A-containing staphylococci. Infect. Immunity 7:881-885, 1973. would be confirmed by motility at 25° 12. D a r l in g , C. L.: Standardization and evalua and absence of it at 37°, the Henry effect tion of the CAMP reaction for the prompt, pre and resistance to nitrofurantoin. The sumptive identification of Streptococcus simple and group-specific coagglutina agalactiae (Lancefield group B) in clinical material. J. Clin. Microbiol. 1:171-174, 1975. tion method11 for S. agalactiae has not 13. E d e r e r , G. M., C h u , J. H., and B l a z e v ic , D. been checked out on L. monocytogenes; J.: Rapid test for urease and phenyalanine it requires relatively costly commercial deaminase production. Appl. Microbiol. 21:545, 1971. reagents. 14. FACKLAM, R. R.: Personal communication, 1976. Acknowledgement 15. F a c k l a m , R. R., Pa d u l a , J. F., T h a c k e r , L. G., W o r t h a m , E . C., and Sc o n y e r s , B. J.: Thanks are extended to Mr. John Redys from the Presumptive identification of group A, B, and Connecticut State Department of health for group D streptococci. Appl. Microbiol. 27:107-113, ing the streptococcal strains and to Dr. Lawrence 1974. Kunz from the Bacteriology Laboratory, Mas 16. FALLON, R. J.: The rapid recognition of Lan sachusetts General Hospital for providing us with cefield group B haemolytic streptococci. J. the L. monocytogenes strains. Clin. Pathol. 27:902-905, 1974. 17. F l a n d r o is , J.-P. and F l e u r e t t e , J.: Carac References tères bactériologiques de Streptococcus agalactiae. Fréquence et conditions d’appari 1. B a i l e y , W. R. and S c o t t , E. C.: Diagnostic tion de la pigmentation. Sensibilité aux an Microbiology, 4th ed. St. Louis: C. V. Mosby tibiotiques. Ann. Biol. Clin. 33:369-377, 1975. Co., 1974. 18. F r a n k , A. and v o n G r a e v e n it z , A.: Sep 2. B e e r e n s , H., and TAHON, M. M.: Milieu a ticemia and meningitis in a newborn due to a l’acide nalidixique pour l’isolement des strep- non-hemolytic group B streptococcus. Infec tococques, D. pneumoniae, Listeria, Erysip- tion 3:60, 1975. elothrix. Ann. Inst. Pasteur 111 :90-93, 1968. 19. F r a s e r , G.: The effect on animal erythrocytes 3. B e r g e r , U.: Einige Eigenschaften der Keim- of combination of diffusible substances pro traegerstaemme von Listeria monocytogenes. duced by bacteria. J. Pathol. Bacteriol. 88:43- Med. Microbiol. Immunol. 161:215-229, 1975. 53, 1964. 4. B l a z e v ic , D. J., Schreckenberger , P. C., 20. G r a n t , J. and G u r n e y , A.: Investigation of the and M a t s e n , J. M.: Evaluation of the Henry effect. II. Isolation at species level. PathoTec “Rapid I-D System.” Appl. Mi IRCS (Research on Biomedical Technology, crobiol. 26:886-889, 1973. Microbiology, Parasitology and Infectious 5. BQJSEN-MOLLER, J.: Human listeriosis. Diag Disease, Veterinary Science) 2:1682, 1974. nostic, epidemiological and clinical studies. 21. G r a y , M. L.: A rapid method for the detection Acta Pathol. Microbiol. Scand. Suppl. B of colonies of Listeria monocytogenes. Zen- 229:1-157, 1972. tralbl. Bakteriol. Abt. I Orig. 169:373-377, 6. B r a u n s t e in , H., T u c k e r , E. B., and G ib s o n , 1957. B. C.: Identification and significance of Strep 22. G r a y , M. L., St a f s e t h , H . J., and T h o r p , F.: tococcus agalactiae (Lancefield group B). The use of potassium tellurite, sodium azide, Amer. J. Clin. Pathol. 51:207-213, 1969. and acetic acid in a selective medium for the 7. B r o o k s , R. F. an d H u c k e r , G. J.: A study o f isolation of Listeria monocytogenes. J. Bac certain members of the genus Corynebac- teriol. 59:443-444, 1950. terium . J. Bacteriol. 48:295-312, 1944. 23. G r o v e s , R. D. and W e l s h im e r , H . J.: Correla 8. B r z i n , B. and SEELIGER, H. R. R.: A brief note tion of virulence in Listeria monocytogenes on the CAMP phenomenon in Listeria. Prob with three in-vitro reactions. Abstr. Annual lems of Listeriosis. Wooduine, M. ed. Leices Meeting American Society of Microbiology ter, Leicester University Press, 1975. D3, 1976. 9. B u c h n e r , L. H., and Sc h n e i e r s o n , S. S.: 24. H e e s c h e n , W ., T o l l e , A., and Z e i d l e r , H .: Clinical and laboratory aspects of Listeria Zur Klassifizierung der Gattung Streptococcus. 2 7 6 KON TN ICK , v o n GRAEVENITZ AND PISCITELLI

Zentralbl. Bakteriol. Abt. I Orig. 205:250-259, 35. M o a t t i, N., Su c , C ., an d L a n d r e a u d , F.: 1967. Isolement et identification rapide de Strep 25. H u g o , W. B. and B e v e r id g e , E. G.: A quan tococcus agalactiae (group B de Lan cefield) — titative and qualitative study of the lipolytic Sa fréquence dans les prélèvements humains. activity of single strains of seven bacterial Med. Mal. Infect. 4:527-535, 1974. species. J. Appl. Bacteriol. 25:72-82, 1962. 36. M u n c h -Pe t e r s e n , E., C h r i s t i e , R., a n d 26. H w a n g , M-N. and E d e r e r , G. M.: Rapid hip- Si m m o n s , R. T.: Further notes on a lytic purate hydrolysis method for presumptive phenomenon shown by group B streptococci. identification of group B streptococci, J. Clin. Austral. J. Exp. Biol. M ed. Sci. 23:193-195, Microbiol. 2:114-115, 1975. 1945. 27. I s e n b e r g , H. D., G o l d b e r g , D., and 37. PATTISON, J. H.: The pathogenicity for mice of SAMPSON, J.: Laboratory studies with a selec group B streptococci of bovine origin. J. Hyg. tive enterococcus medium. Appl. Microbiol. 47:159-165, 1949. 20:433-436, 1970. 38. R o d l e r , M. and Vesztergombi, Z.: Biochem 28. K o n t n i c k , C., and v o n G r a e v e n i t z , A.: ical properties of Listeria monocytogenes Diagnostic implications of a positive hippurate strains. Acta Microbiol. Acd. Sci. Hung. test in Listeria monocytogenes. Abstr. Annual 19:337-339, 1972. Meeting American Society of Microbiology, 39. R o g o s a , M ., W is e m a n , R. F., M i t c h e l l , J. A., C46, 1975. D is r a e l y , M . N., and B e a m a n , A. J.: Species 29. KOSER, S. A.: Utilization of the salts of organic differentiation of oral lactobacilli from man, in acids by the colon-aerogenes group. J. Bac cluding descriptions of Lactobacillus teriol. 8:493-520, 1923. salivarius nov. sp. and Lactobacillus cel- 30. LAGODSKY, H. and JANDARD, M l l e .: Sur une lobiosus nov. sp. J. Bacteriol. 65:681-699, Variante de methode rapide pour diagnose de 1953. gelatinolyse microbienne. Ann. Inst. Pasteur 40. R o m e r o , R. and W il k in s o n , H. W .: Identifica 107:128-131, 1964. tion of group B streptococci by immuno- fluorescent staining. Appl. Microbiol. 28:199- 31. L in z e n m e ie r , G. and Se e l i g e r , H.: Die in- vitro Empfindlichkeit von Listeria 204, 1974. monocytogenes (Pirie) gegen Sulfonamide und 41. Se e l i g e r , H. P. R .: Listeriosis. Basel-New Antibiotika. Zentralbl. Bakteriol. Abt. I Orig. York: S. Karger, 1961. i 60:543-558, 1953-54. 42. Se e l i g e r , H. P. R. and W e l s h im e r , H. J.: Genus Listeria. Bergey’s Manual of Determi 32. LUTTICKEN, R. and F r i t s c h e , D.: Bestim native Bacteriology, 8th ed. Buchanan, R. E. mung der B-Streptokokken (Streptococcus and Gibbons, N. E., ed. Baltimore: Williams & agalactiae) mit Hilfe des “Dreifachtests” nach Wilkins Co., 1974. Wallerström und des CAMP-Tests (Identifica 43. v o n G r a e v e n i t z , A.: Deoxyribonuclease- tion of Group B Streptococci (Streptococcus indole medium and its use in an identification agalactiae) by means of the Triple-Test of Wal- system for non-lactose fermenting Gram- lerstrom and the CAMP-Test) Zentralbl. Bakt. negative rods. Ann. Clin. Lab. Sci. 3:284-288, Hyg., I. Abt. Orig. A 226:298-304, 1974. 1973. 33. M c B r id e , M . E. and G i r a r d , K. F.: A selective 44. W e t z l e r , T. F., F r e e m a n , N. R., F r e n c h , M. method for the isolation of Listeria LV., R e n k o w s k i, L. A., E v e l a n d , W . C., and monocytogenes from mixed bacterial popula C a r v e r , O. J.: Biological characterization of tions. J. Lab. Clin. Med. 55:153-157, 1969. Listeria monocytogenes. Health Lab. Sci. 34. M e r r it t , K., T r e a d w e l l , T. L., and Ja c o b s , 5:46-62, 1968. N. J.: Rapid recognition of group B streptococci 45. W i l k i n s o n , H. W ., T h a c k e r , L. G., and by pigment production and counterim- F a c k l a m , R. R.: Nonhemolytic group B strep munoelectrophoresis. J. Clin. Microbiol. tococci of human, bovine, and ichthyic origin. 3:287-290, 1976. Inf. Immun. 7:496-498, 1973.