Lipopolysaccharides as Determinants of Serological Variability in Pseudomonas corrugata

The variation in biochemical and serological features of 128 isolates of Pseudomonas corrugata has been studied with 56 isolates from Spain and 72 isolates from other countries. Isolates were analyzed with common diagnostic tests and with the AP150CHE system. Variability among isolates for some standard tests usually listed as positive or negative for this species, such as arginine dihydrolase and gelatin hydrolysis, lipase and lecithinase activities, pigment production, and wrinkled colony morphology, was observed. Three antisera were raised against the type strain and two Spanish isolates from tomato and pepper plants. Serological reactions were studied by indirect immunofluorescence and indirect enzyme-linked immunosorbent assay. Eighty-three isolates reacted with a single antiserum, 6 reacted with two antisera, and none reacted with three antisera. Thirty-nine isolates did not react with any of the three antisera. These results suggest that serology will not be a useful method for routine diagnosis of P. corrugata unless common antigens can be identified. Electrophoresis and immunoelectrotransfer were used to study the antigens involved. Each antiserum reacted with whole-cell lysates, giving two common bands for P. corrugata isolates and other Pseudomonas species and a ladder-like pattern characteristic of lipopolysaccharides (LPS). Common bands were not observed after proteinase K treatment. More than 10 LPS patterns were distinguished in 98 isolates after silver staining of polyacrylamide gels. There was no correlation between the geographical origin or host of the isolates and the LPS patterns. A correlation between LPS groups and serological reaction was observed.     

Identification of Yeast Strains Isolated from Agricultural Soils for Releasing Potassium-bearing Minerals

Thirty-five yeast strains were isolated from soil samples that were collected from different locations in Upper Egypt. The purified isolates were screened for the release potassium from mica on Aleksandrov agar medium. Two yeast isolates (KSY-29 and KSY-33) showed an ability to solubilize potassium by inducing clear zones around their colonies. They were identified as Pichia anomala and Rhodotorula glutinis, respectively, based on PCR analysis of the ITSI-26S region that was amplified by NL1/NL4 species-specific primers. The amount of K released from muscovite mica in the broth culture of the yeast isolates was measured after 5, 10, 15 and 20 days of the incubation at 25°C. Both yeast isolates were very effective in releasing K of muscovite in broth culture, recording 8.11–13.21 μg/ml that were released from muscovite mica after 20 days of incubation. The inoculation of maize (Zea maize) plants with these yeast isolates under different K levels (25, 50 and 100% of recommended dose of potassium, RDK) as potassium sulfate was tested on growth and K uptake by these plants in the greenhouse. Significant increases (p < 0.05) in plant height, root and shoot dry weights as well as K uptakes by shoots and roots maize plants occurred through the inoculation with KSY-29 or KSY-33 isolates.     

Serological and biological characterisation of isolates of barley yellow dwarf virus in Sweden in 1983

In 1983, cereal plants showing symptoms of barley yellow dwarf virus (BYDV), collected from 15 localities in Sweden, were tested for BYDV using enzyme-linked immunosorbent assay (ELISA). Antisera against two Swedish isolates of BYDV were used, a mild isolate (27/77) transmitted specifically by Sitobion avenae and a severe one (39/78) transmitted mainly by Rhopalosiphum padi. No virus was detected in 57 of 607 plants of oats and barley tested. Of the 550 plants in which virus was detected, 366 were infected with viruses similar to isolate 27/77, 116 with viruses similar to 39/78 and the remaining 68 reacted strongly with both antisera. When tested, the latter isolates were shown to be mixtures. Thirty-nine selected samples were also tested with antisera against the USA isolates RPV, RMV, MAV and PAV, and for transmission by S. avenae and R. padi. Twenty-six of these samples were transmitted specifically by S. avenae, one was transmitted only by R. padi and the remaining 12 samples were shown to be infected with a mixture of an S. avenae-specific isolate and one transmitted mainly by R. padi. Antisera against PAV and MAV each detected all isolates tested and the results were very similar to those with the antisera to the 39/78 and 27/77 isolates, respectively. None of the field isolates reacted with antisera against RMV or RPV. It was concluded that 1983 was an epidemic year for BYDV in Sweden and that isolates specifically transmitted by S. avenae predominated. Symptoms of infection by these isolates on oat plants ranged from mild to severe.     

Improved Method for Detection of Vibrio parahaemolyticus in Seafood

We have developed a new, effective procedure for detecting Vibrio parahaemolyticus in seafoods using enrichment and plating onto a chromogenic agar medium. Samples were cultured in salt Trypticase soy broth, which is a nonselective medium, and then a portion of the culture was cultured with salt polymyxin broth, which is a selective medium for V. parahaemolyticus. This two-step enrichment was more effective than the one-step enrichment in salt polymyxin broth alone. The enrichment cultures were then plated onto a new chromogenic agar containing substrates for beta-galactosidase. The V. parahaemolyticus colonies developed a purple color on this growth medium that distinguished them from other related bacterial strains. V. parahaemolyticus was isolated more frequently from naturally contaminated seafood samples using the chromogenic agar than thiosulfate citrate bile salts sucrose agar medium, which is currently used for the isolation of V. parahaemolyticus. Our findings suggest that this new enrichment and isolation scheme is more sensitive and accurate for identifying V. parahaemolyticus in seafood samples than previously used methods.     

Serological identification of group D streptococci using commercial antisera

Three commercial group D streptococcal antisera were tested for the serological identification of 100 group D enterococci; 20 Streptococcus bovis; 5 isolated from each of the following streptococcal groups: A, B, C and G; and 3 isolates from serological group F. Antisera from Difco Laboratories, BBL, and Wellcome Reagents Limited were used in the classic capillary tube precipitin test on extracts prepared using the Rantz and Randall procedure. No false positive precipitin reactions were observed. Of the enterococcal isolates, all 100 reacted with the Wellcome, 99 reacted with the BBL, and 96 reacted with the Difco group D antisera. However, of the 20 S. bovis isolates, only 2 reacted with the BBL, and 1 reacted with both the Difco and the Wellcome antisera. Each antiserum was then used to prepare staphylococcal coagglutination (CoA) reagents and each isolate was subsequently tested. A simple extraction procedure was performed by suspending colonies of an isolate in a loopful of salin on a microscope slide and gently heating the slide directly in the flame of a Bunsen burner. All 100 enterococci and all 20 S. bovis gave positive results with the BBL and the Wellcome CoA reagents. Using the Difco reagent, 2 S. bovis isolates failed to produce postitive results. No false positive results were observed with the non-Group D isolates. Our results indicate that the CoA technique using commercial group D antisera may provide faster and sometimes more sensitive serological identification than the classic capillary tube precipitin test.     

The effect of nutrient medium and age ofErwinia amylovora andErwinia herbicola cultures on their reactivity in agglutination test

The influence of the type of nutrient medium (meat-peptone nutrient agar, KING B medium, YDC medium) and the age of bacterial cultures ofErwinia amylovora andE. herbicola on reactivity of their antigens with homologous antisera were examined. In the case ofE. amylovora, the best agglutination reactions were observed with isolates cultivated on the YDC nutrient medium. After 192 h of cultivation, seven outof ten isolates reacted positively. The reactivity ofE. herbicola antigen decreased in dependence on culture age more rapidly than withE. amylovora. The highest rates of positive agglutination reactions were observed withE. herbicola cultivated on the YDC and KING B nutrient media. After 144 h of cultivation on both these nutrient media eight out of ten isolates reacted positively.     

Comparative quantitative proteomics of prochlorococcus ecotypes to a decrease in environmental phosphate concentrations

Background

Goa is a coastal state in India and salt making is being practiced for many years. This investigation aimed in determining the culturable haloarchaeal diversity during two different phases of salt production in a natural solar saltern of Ribandar, Goa. Water and sediment samples were collected from the saltern during pre-salt harvesting phase and salt harvesting phase. Salinity and pH of the sampling site was determined. Isolates were obtained by plating of the samples on complex and synthetic haloarchaeal media. Morphology of the isolates was determined using Gram staining and electron microscopy. Response of cells to distilled water was studied spectrophotometrically at 600nm. Molecular identification of the isolates was performed by sequencing the 16S rRNA.

Results

Salinity of salt pans varied from 3-4% (non-salt production phase) to 30% (salt production phase) and pH varied from 7.0-8.0. Seven haloarchaeal strains were isolated from water and sediment samples during non-salt production phase and seventeen haloarchaeal strains were isolated during the salt production phase. All the strains stained uniformly Gram negative. The orange-red acetone extract of the pigments showed similar spectrophotometric profile with absorption maxima at 393, 474, 501 and 535 nm. All isolates obtained from the salt dilute phase were grouped within the genus Halococcus. This was validated using both total lipid profiling and 16S rRNA data sequencing. The isolates obtained from pre-salt harvesting phase were resistant to lysis. 16S rRNA data showed that organisms belonging to Halorubrum, Haloarcula, Haloferax and Halococcus genera were obtained during the salt concentrated phase. The isolates obtained from salt harvesting phase showed varied lysis on suspension in distilled water and /or 3.5% NaCl.

Conclusion

Salterns in Goa are transiently operated during post monsoon season from January to May. During the pre-salt harvesting phase, all the isolates obtained belonged to Halococcus sp. During the salt harvesting phase, isolates belonging to Halorubrum, Haloarcula, Haloferax and Halococcus genera were obtained. This study clearly indicates that Halococcus sp. dominates during the low salinity conditions.     

Microbial Ecology of Activated Sludge: I. Dominant Bacteria

Over 300 bacterial strains were isolated from seven samples of activated sludge by plating on sewage agar. Gram-negative bacteria of the genera Zoogloea and Comamonas predominated. Many isolates (51%) showed sudanophilic inclusions of poly-β-hydroxybutyric acid, whereas 34% accumulated iodophilic material on media containing starch. A large number required either vitamins or amino acids, or both, for growth. None of the isolates tested for their ability to bring about changes in autoclaved sewage produced an effluent comparable in quality to the activated sludge control, although the Zoogloea did produce activated sludgelike flocs. A study of 150 bacterial strains isolated from raw sewage revealed that they differed from the sludge isolates in several respects. Coliforms, which constitute nearly a quarter of the sewage isolates, were rarely encountered in sludge.     

The serological specificity of S alleles of homozygous incompatible lines of the marrow-stem kale (Brassica oleracea var.acephala DC.)

The preparation of antisera against S alleles of homozygous incompatible lines of the marrow-stem kale (Brassica oleracea var.acephala DC.) is described. The antisera obtained reacted specifically with homologous antigens on using the serological method of double diffusion into agar. The results confirmed the specificity of S alleles of incompatible lines of the marrow-stem kale.     

Bovine Feces from Animals with Gastrointestinal Infections Are a Source of Serologically Diverse Atypical Enteropathogenic Escherichia coli and Shiga Toxin-Producing E. coli Strains That Commonly Possess Intimin

Shiga toxin-producing Escherichia coli (STEC) and enteropathogenic E. coli (EPEC) cells were isolated from 191 fecal samples from cattle with gastrointestinal infections (diagnostic samples) collected in New South Wales, Australia. By using a multiplex PCR, E. coli cells possessing combinations of stx₁, stx₂, eae, and ehxA were detected by a combination of direct culture and enrichment in E. coli (EC) (modified) broth followed by plating on vancomycin-cefixime-cefsulodin blood (BVCC) agar for the presence of enterohemolytic colonies and on sorbitol MacConkey agar for the presence of non-sorbitol-fermenting colonies. The high prevalence of the intimin gene eae was a feature of the STEC (35 [29.2%] of 120 isolates) and contrasted with the low prevalence (9 [0.5%] of 1,692 fecal samples possessed STEC with eae) of this gene among STEC recovered during extensive sampling of feces from healthy slaughter-age cattle in Australia (M. Hornitzky, B. A. Vanselow, K. Walker, K. A. Bettelheim, B. Corney, P. Gill, G. Bailey, and S. P. Djordjevic, Appl. Environ. Microbiol. 68:6439-6445, 2002). Forty-seven STEC serotypes were identified, including O5:H−, O8:H19, O26:H−, O26:H11, O113:H21, O157:H7, O157:H− and Ont:H− which are known to cause severe disease in humans and 23 previously unreported STEC serotypes. Serotypes Ont:H− and O113:H21 represented the two most frequently isolated STEC isolates and were cultured from nine (4.7%) and seven (3.7%) animals, respectively. Fifteen eae-positive E. coli serotypes, considered to represent atypical EPEC, were identified, with O111:H− representing the most prevalent. Using both techniques, STEC cells were cultured from 69 (36.1%) samples and EPEC cells were cultured from 30 (15.7%) samples, including 9 (4.7%) samples which yielded both STEC and EPEC. Culture on BVCC agar following enrichment in EC (modified) broth was the most successful method for the isolation of STEC (24.1% of samples), and direct culture on BVCC agar was the most successful method for the isolation of EPEC (14.1% samples). These studies show that diarrheagenic calves and cattle represent important reservoirs of eae-positive E. coli.     

Cold,pH and salt tolerant Penicillium spp. inhabit the high altitude soils in Himalaya,India

Twenty five fungal cultures (Penicillium spp.), isolated from soil samples from the high altitudes in the Indian Himalayan region, have been characterized following polyphasic approach. Colony morphology performed on five different media gave varying results; potato dextrose agar being the best for the vegetative growth and sporulation as well. Microscopic observations revealed 18 isolates to be biverticillate and 7 monoverticillate. Based on the phenotypic characters (colony morphology and microscopy), all the isolates were designated to the genus Penicillium. Exposure to low temperature resulted in enhanced sporulation in 23 isolates, while it ceased in case of two. The fungal isolates produced watery exudates in varying amount that in many cases increased at low temperature. All the isolates could grow between 4 and 37 °C, (optimum 24 °C), hence considered psychrotolerant. While all the isolates could tolerate pH from 2 to 14 (optimum 5–9), 7 isolates tolerated pH 1.5 as well. While all the fungal isolates tolerated salt concentration above 10 %; 10 isolates showed tolerance above 20 %. Based on ITS region (ITS1-5.8S-ITS2) analysis the fungal isolates belonged to 25 different species of Penicillium (showing similarity between 95 and 100 %). Characters like tolerance for low temperature, wide range of pH, and high salt concentration, and enhancement in sporulation and production of secondary metabolites such as watery exudates at low temperature can be attributed to the ecological resilience possessed by these fungi for survival under low temperature environment of mountain ecosystem.     

Antimicrobial and antifungal activity of soil actinomycetes isolated from coal mine sites

In the current study, twenty-eight soil samples were collected from coalmine sites of Telangana, India. The isolates were purified and identified based on their culture characterization on oatmeal agar, glycerol asparagine agar, yeast extract-malt extract agar, inorganic salt starch agar, and starch casein agar medium. Further, the supernatant of all the isolates were tested for antimicrobial and antifungal activities. The biochemical and microscopic studies of isolated strains results indicates the potential isolate strains belongs to Streptomyces genus. Among all the strains the biological activity of BHPL-KSKU5 showed higher anti-bacterial and anti-funagal activity. The molecular characterization of BHPL-KSKU5 16s rDNA gene sequence and phylogenetic tree showed that is mostly related to the Streptomysis felleus (S. felleus) strain. This isolate was submitted to gene bank NCBI with accession number MH553077. In addition, physiological studies such as utilization of carbon, nitrogen, amino acid sources of potential isolated were studied. Further, optimization, purification and characterization of the novel compound producing strain may be helpful for discovering the new therapeutic microbial agent.     

Enterotoxigenic Bacteria in Food and Water from an Ethiopian Community

Food and water samples from an Ethiopian community were screened for the presence of enterotoxin-producing bacteria. Using the Chinese hamster ovary cell assay, 40 of 213 isolates (18.8%) produced heat-labile (LT) enterotoxin. These LT-producing isolates comprised 33 of 177 (18.6%) strains from 24 of 68 food samples (35.3%) and 7 of 36 (19.4%) isolates of 4 of 17 water samples (23.5%). One LT-producing strain each of Salmonella emek and of Shigella dysenteriae was found. Three pseudomonads, all LT producers, produced heat-stable enterotoxin as gauged by the suckling mouse test. Two strains of LT-enterotoxigenic Escherichia coli O68 were found in water samples. No enterotoxigenic E. coli were isolated from food samples, but 13 of the LT-producing strains were Enterobacter, Klebsiella, Serratia, and Proteus species, and 7 food samples yielded more than one species of enterotoxigenic bacterium. Of the enterotoxigenic isolates from food, 15 were oxidase-positive strains of the genera Aeromonas, Pseudomonas, Achromobacter, Flavobacterium, and Vibrio. LT-enterotoxigenic Enterobacter, Acinetobacter, Klebsiella, Proteus, Providencia, and Serratia species represented 20 of the food and water isolates. Culture supernatant fluids of representative strains of oxidase-positive and oxidase-negative species giving positive reactions in Chinese hamster ovary cell tests induced fluid accumulation in rabbit ileal loops. Eight of the food samples and two of the water samples contained more than one isolate or species of enterotoxigenic bacterium. The stability of the LT production by oxidase-positive bacteria and non-E. coli strains was assessed by the rabbit skin and adrenal cell tests after 9 months and 1 year of storage, respectively, in Trypticase soy broth with glycerol at −70°C. Only 33% of the oxidase-positive strains were still LT enterotoxigenic. Of the oxidase-negative strains, 50 and 33% were LT producing at 9 months and 1 year, respectively. None of the E. coli isolates, both enterotoxigenic and nonenterotoxigenic, possessed K88, K99, or colonization factor antigen. The survey demonstrates the presence in food and water of enterotoxigenic bacteria of the same species as those isolated from cases of infantile diarrhea in the same community, although a correlation between these sources and infantile diarrhea remains to be established.     

Marine Ammonia- and Nitrite-Oxidizing Bacteria: Serological Diversity Determined by Immunofluorescence in Culture and in the Environment

Immunofluorescence assays for marine ammonium- and nitrite-oxidizing bacteria were used to assess the diversity of nitrifying bacteria isolated from marine environments. The antisera show relatively broad specificity, in that each reacts with several strains of the same physiological type as the strain to which the antiserum was prepared. The antisera do not, however, react with any strains of differing physiological type. Seventy percent of the 30 unidentified ammonium-oxidizing isolates tested reacted with one or both of the antisera produced to marine ammonium-oxidizing strains, and 8 of the 9 unidentified nitrite-oxidizing strains tested reacted with 1 or more of the 3 nitrite oxidizer antisera used. Ammonium- and nitrite-oxidizing bacteria were enumerated in samples taken in a depth profile (to 750 m) in the Southern California Bight by immunofluorescence assays for two ammonium oxidizers and two nitrite oxidizers. Average abundances of the two types of nitrifiers were 3.5 × 10⁵ and 2.8 × 10⁵ cells liter⁻¹, respectively. Nitrifiers constitute 0.1 to 0.8% of the total bacterial population in these samples.     

Charactererization of novel non-toxic Bacillus thuringiensis isoloates from Korea

J.Y. ROH, H.W. PARK, B.R. JIN, H.S. KIM, Y.M. YU AND S.K. KANG. 1996. Four Bacillus thuringiensis isolates from soil samples produced parasporal inclusions which were non-toxic to insects. The isolates were named B. thuringiensis NTB-1, NTB-2, NTB-3 and NTB-4. The parasporal inclusions were shown to be ovoid by phase contrast and scanning electron microscopy. The serotypes of the four isolates were determined by agglutination using 33 antisera; NTB-1 and NTB-4 seemed to be subsp. isruelensis ,and NTB-2 seemed to be subsp. pondzcheriensis . NTB-3 did not react with the 33 antisera. However, comparison of parasporal protein and plasmid DNA patterns of the four isolates with those of 15 known non-toxic B. thuringiensis strains demonstrated that the four isolates are novel.     

Improved Isolation of Vibrio vulnificus from Seawater and Sediment with Cellobiose-Colistin Agar

An improved selective medium, cellobiose-colistin (CC) agar, gave a significantly higher (P < 0.05) isolation rate of Vibrio vulnificus from water and sediment samples than did modified cellobiose-polymyxin B-colistin (mCPC) agar. In a total of 446 alkaline peptone water preenrichments amended with polymyxin B, V. vulnificus was isolated from 154 preenrichments (35%) with mCPC agar and from 179 preenrichments (40%) with CC agar. CC agar gave a higher plating efficiency of V. vulnificus cells than did cellobiose-polymyxin B-colistin (CPC) agar, mCPC agar, or thiosulfate-citrate-bile salts-sucrose (TCBS) agar; the only significant difference was observed with TCBS agar, which gave much lower plating efficiencies than the other selective media. Determination of MICs demonstrated that the concentrations of colistin and polymyxin B in CPC agar inhibit growth of a proportion of V. vulnificus strains.     

Serological cross-reactivity of environmental isolates of Enterobacter, Escherichia, Stenotrophomonas, and Aerococcus with Shigella spp.-specific antisera

Using protocols designed for the isolation of Shigella from environmental freshwater samples from different regions of Bangladesh, 11 bacterial strains giving rise to Shigella-like colonies on selective agar plates and showing serological cross-reaction with Shigella-specific antisera were isolated. Phylogenetic analyses revealed that three of the isolates were most closely related to Escherichia coli, four to Enterobacter sp., two to Stenotrophomonas, and two isolates belonged to the Gram-positive genus Aerococcus. The isolates cross-reacted with six different serotypes of Shigella and were, in each case, highly type-specific. Two of the isolates belonging to the Enterobacter and Escherichia genera gave extremely strong cross-reactivity with Shigella dysenteriae and Shigella boydii antisera, respectively. The Aerococcus isolates gave relatively weak but significant cross-reactions with S. dysenteriae. Western blot analysis revealed that a number of antigens from the isolates cross-react with Shigella spp. The results indicate that important Shigella spp. surface antigens are shared by a number of environmental bacteria, which have implications for the use of serological methods in attempts for the detection and recovery of Shigella from aquatic environments.     

Prevalence of Pandemic Thermostable Direct Hemolysin-Producing Vibrio parahaemolyticus O3:K6 in Seafood and the Coastal Environment in Japan

Although thermostable direct hemolysin (TDH)-producing Vibrio parahaemolyticus has caused many infections in Asian countries, the United States, and other countries, it has been difficult to detect the same pathogen in seafoods and other environmental samples. In this study, we detected and enumerated tdh gene-positive V. parahaemolyticus in Japanese seafoods with a tdh-specific PCR method, a chromogenic agar medium, and a most-probable-number method. The tdh gene was detected in 33 of 329 seafood samples (10.0%). The number of tdh-positive V. parahaemolyticus ranged from <3 to 93/10 g. The incidence of tdh-positive V. parahaemolyticus tended to be high in samples contaminated with relatively high levels of total V. parahaemolyticus. TDH-producing strains of V. parahaemolyticus were isolated from 11 of 33 tdh-positive samples (short-necked clam, hen clam, and rock oyster). TDH-producing strains of V. parahaemolyticus were also isolated from the sediments of rivers near the coast in Japan. Representative strains of the seafood and sediment isolates were examined for the O:K serovar and by the PCR method specific to the pandemic clone and arbitrarily primed PCR and pulsed-field gel electrophoresis techniques. The results indicated that most O3:K6 tdh-positive strains belonged to the pandemic O3:K6 clone and suggested that serovariation took place in the Japanese environment.     

Morphological, Cultural, Biochemical, and Serological Comparison of Japanese Strains of Vibrio parahemolyticus with Related Cultures Isolated in the United States

Morphological, cultural, biochemical, and serological characteristics of 79 strains of Vibrio parahemolyticus isolated from patients suffering from gastroenteric disease in Japan were compared with 17 suspected V. parahemolyticus cultures isolated from wound infections and 14 nonpathogenic vibrios isolated from an estuarine environment in the United States. These groups were differentiated on the basis of several key reactions which included: the range of growth temperature and salt tolerance; the production of catalase and acetoin; the hydrolysis of starch; the fermentation and utilization as single carbon source of sucrose, cellobiose, and arabinose; and the ability to swarm on 1% agar. The separation of the groups on the basis of cultural and biochemical analyses was confirmed by means of slide agglutinations with specific anti-K antisera. The results of this study strongly suggest that the wound infection isolates are V. parahemolyticus species which are easily distinguished from the nonpathogenic estuarine vibrios.     

Diagnostic properties of three conventional selective plating media for selection of <Emphasis Type="Italic">Bacillus cereus</Emphasis>, <Emphasis Type="Italic">B. thuringiensis</Emphasis> and <Emphasis Type="Italic">B. weihenstephanensis</Emphasis>

The aim of this study was to assess the diagnostic properties of the two selective plating media and a chromogenic medium for identification of Bacillus cereus. The 324 isolates were B. cereus (37%), Bacillus weihenstephanensis (45%) or Bacillus thuringiensis (18%), as identified by a new combination of techniques. All isolates were growing on mannitol–egg yolk–polymyxin agar (MYP), and they did not form acid from mannitol. However, a significant lower number of B. thuringiensis isolates did not show lecithinase activity. All isolates were also growing on polymyxin–egg yolk–mannitol–bromothymol blue agar (PEMBA); however, 11% isolates indicated that they did produce acid from mannitol, and 15% isolates did not show any lecithinase activity. Five of the isolates did not grow at all on the chromogenic agar, and 14 of the growing isolates were β-glucosidase negative. It is concluded that the two recommended selective plating media MYP and PEMBA for detection of B. cereus group bacteria both have their limitations for identification of some B. cereus, B. weihenstephanensis or B. thuringiensis. However, MYP is preferable compared to PEMBA. The chromogenic medium has its own advantages and limitations, and some of the limitations seem to be solved by incubation at 30°C instead of the recommended 37°C.