DNA Macrorestriction Patterns Research Articles

Antimicrobial Resistance, Serotypes, and Virulence Factors ofStreptococcus suisIsolates from Diseased Pigs

Streptococcus suis isolates from diseased pigs were examined for susceptibility to nine antimicrobials, possession of virulence-associated factors (VFs), and distribution of serotypes. The association between antimicrobial resistance (AMR) and serotypes as well as VFs was subsequently assessed. Among the isolates investigated, serotype 2 (66.04%) was mostly prevalent, followed by serotypes 1 (23.27%), 9 (1.26%), and 7 (0.63%), whereas 14 isolates were untypable by the polymerase chain reaction typing method used. Analysis with pulsed-field gel electrophoresis revealed the isolates had diverse DNA macrorestriction patterns. The frequency of antimicrobial resistance among the S. suis isolates was higher than that reported from other countries. It is notable that multiple antimicrobial resistance (three or more antimicrobials) was observed with 98.73% of the S. suis isolates, and the dominant resistance phenotype was erythromycin-tilmicosin-clindamycin-chloramphenicol-levofloxacin-ceftiofur-kanamycin-tetracycline-penicillin (35.85%). The most prevalent VFs were those encoded by muramidase-released protein (61.64%), followed by suilysin (56.60%) and extracellular factor (46.54%). Presence of VFs and the possession of certain AMR phenotypes were significantly associated as determined by statistical analysis. Together, these findings indicate that the clinical S. suis isolates obtained from diseased pigs in China are genetically diverse, are resistant to multiple antibiotics of clinical importance, and carry known virulence factors.

GENOTIPIC CHARACTERIZATION OF SALMONELLA INFANTIS STRAINS ISOLATED IN MARCHE REGION

In this study thirty-eight strains of Salmonella enterica serovar Infantis isolated in Marche Region from human cases, food, animal and environmental samples were analyzed by Pulsed-Field Gel Electrophoresis (PFGE). All strains were typed by the DNA macro-restriction patterns obtained following PFGE of XbaI digests, while antimicrobial susceptibility testing of isolates was performed by the standardized disk diffusion method. The analysis of PFGE patterns by Bionumerics software demonstrated a strong similarity of S. infantis XbaI profiles, while the antimicrobial susceptibly testing showed less homogeneity.

OCCURRENCE AND CHARACTERIZATION OF STAPHYLOCOCCUS AUREUS ISOLATED FROM MEAT AND DAIRY PRODUCTS CONSUMED IN TURKEY

ABSTRACT A total of 413 samples of milk, dairy products, meat and meat products were analyzed for the presence of Staphylococcus aureus in a 2‐year survey (2004–2006). One hundred thirty‐eight samples were (33.4%) contaminated with S. aureus. A total of 138 S. aureus strains were isolated and identified. Enterotoksin profiles were determined by Staphylococcal Enterotoxin Test Reversed Passive Latex Agglutination. Their resistance to nine antibiotics were tested. The contamination rate of the meat product samples (48.7%) was significantly higher than milk and dairy products (23.2%). Most of the strains (60.1%) produced staphylococcal enterotoxins. Four strains produced staphylococcal enterotoxin C and staphylococcal enterotoxin D. All of the strains tested were resistant to many of the nine antibiotics. One strain isolated from feta cheese was resistant to all of the antibiotics tested. Penicillin G resistance was the highest (92.7%) among the antibiotics. Seventy‐one strains choosen randomly were compared by the DNA macrorestriction patterns obtained from pulsed field gel electrophoresis following Sma I digestion and 33 pulsotypes were determined. Seventy‐three strains were characterized by automated ribotyping using Eco RI and 8 ribotypes were determined. The same, as well as different pulsotypes, were identified among strains with the identical ribotyping profile. The presence of enterotoxigenic and antimicrobial resistant strains of S. aureus has become remarkably wide spread in the foods examined and poor sanitary conditions during processing may create a health risk for the consumers. The results of genotyping showed high diversity among the strains.PRACTICAL APPLICATIONSThese results suggests that the contamination of various food samples by Staphylococcus aureus indicates poor personel hygiene and/or cross contamination in Turkey. Potential of S. aureus strains to produce enteroxins represent a health hazard to the consumers. Prevalence of multiple resistance to the antibiotics of the strains suggests the restricted use of them in animal and human therapy and growth promotion in animals. Genotyping with pulsed field gel electrophoresis (PFGE) revealed more subtypes of S. aureus strains isolated from different food sources than automated ribotyping. Therefore, although both methods could be applied to demonsrate the source and route of the contamination, PFGE should be preferred in future studies of epidemiology of the S. aureus contamination.

Genetic heterogeneity of Escherichia coli strains isolated from raw milk, Minas Frescal cheese, and food handlers

From February 2004 to March 2005, 140 samples of food handlers - hands and nostrils - (92), raw milk (24), and minas frescal cheese (24) were analyzed for the presence of Escherichia coli in a dairy processing plant of Goiás State. Forty-seven E. coli strains were obtained and compared by DNA macrorestriction patterns obtained from pulsed-field gel electrophoresis following XbaI restriction in order to investigate the possible sources of cheese contaminations. Based on PFGE genotyping, one strain isolated from food the hands of a handler and five strains isolated from raw milk were identical or closely related to six strains from cheese suggesting, in these cases, the probable source of E. coli contamination in cheeses. No strain isolated from the nostrils was related to those found in cheeses or milk strains. The results showed high diversity among the strains, demonstrating a lack of predominance of an endemic clone in the dairy plant. This paper highlights the usefulness of PFGE as an epidemiological tool for determining the source of E. coli contamination in the food industry.

Listeria monocytogenes isolated from vegetable salads sold at supermarkets in Santiago, Chile: Prevalence and strain characterization

Between 2000 and 2005, 717 samples of three types of salads were analysed for Listeria monocytogenes in Santiago, Chile in order to provide information to Chilean health authorities on the presence of the pathogen in vegetable salad samples and to ascertain the risk of these products for consumers. L. monocytogenes isolates were found in 88 out of 347 (25.4%) samples of frozen vegetable salads and in 22 out of 216 (10.2%) freshly supermarkets prepared, cooked or raw ready-to-eat vegetable salads; no Listeria was isolated from 154 samples of raw minimally processed salads industrially prepared. Enumeration of L. monocytogenes was done by plate count for 20 positive frozen samples, randomly chosen. Most of them (90%) had < 10 cfu/g. MPN technique was performed for 34 another positive samples; 12 had ≥ 1100/g, five ranged between 240 and 93, eight between 23 and three and nine had < 3.0. No L. monocytogenes was recovered after cooking 12 contaminated frozen samples. Isolation of strains was done using three selective agars. Sixty-two L. monocytogenes were isolated from lithium chloride phenylethanol moxalactam agar, 95 from Listeria selective agar Oxford formulation, and 103 from polymixin acriflavine lithium chloride ceftazidime aesculin mannitol agar. Fifty isolates (45.5%) belong to PCR group IIb (including strains serovar 1/2b), 41 (37.3%) to PCR group IVb (including strains serovar 4b), 17 (15.5%) to PCR group IIa (including strains serovar 1/2a), and 2 (1.8%) to PCR group IIc. With the use of DNA macrorestriction patterns analysis, 17 different clusters were detected among 71 isolates, with P10, the most frequent with 25 isolates (35.2%) of PCR group IIb.

Dissemination of imipenem-resistant Acinetobacter baumannii strains carrying the ISAba1–bla OXA-23 genes in a Chinese hospital

An outbreak of 95 clinical infections with imipenem-resistant Acinetobacter baumannii in a Chinese hospital was investigated and the carbapenemase-encoding genes and their relationship with ISAba1 of these and a further 16 isolates recovered from the intensive care unit (ICU) environment were analysed. Almost all isolates were resistant to a wide range of antimicrobials; the lowest resistance rates were found for polymyxin E (17.1 %), cefoperazone/sulbactam (30.6 %) and ampicillin/sulbactam (67.6 %). Six pattern types defined by DNA macrorestriction patterns were distinguished among the clinical isolates with dissemination of pattern A (50 isolates) to patients in seven hospital units and pattern B (35 isolates) to eight units; the environmental isolates from ICUs were also of pattern A. All isolates were positive for the bla(OXA-66) and bla(OXA-23) genes. The OXA-23-encoding gene was located 34 bp downstream of ISAba1. No plasmids were detected and conjugal transfer of resistance was not demonstrated. The bla(OXA-23) probe hybridized with 200 and 220 kb ApaI chromosomal fragments for type patterns A and B, respectively.

Comparison of Staphylococcus aureus isolates from food handlers, raw bovine milk and Minas Frescal cheese by antibiogram and pulsed-field gel electrophoresis following SmaI digestion

During a year (from February/2004 to March/2005) 140 samples of food handlers (92), raw milk (24) and Minas Frescal cheese (24) were analyzed for the presence of Staphylococcus aureus in a dairy processing plant of Goiás State, Brazil. Seventy-three S. aureus isolates were obtained and compared by antimicrobial susceptibility and by the DNA macrorestriction patterns obtained from pulsed-field gel electrophoresis following SmaI digestion in order to investigate the possible sources of cheeses contamination. The results showed high diversity among the strains, demonstrating a lack of predominance of an endemic clone in the dairy plant. Based on the results, the raw milk appears to be the most probable source of S. aureus contamination in cheeses.

Genetic diversity of Listeria monocytogenes recovered from infected persons and pork, seafood and dairy products on retail sale in France during 2000 and 2001

Growth of the food-borne human pathogen Listeria monocytogenes to large numbers in ready-to-eat food products greatly increases the risk of disease for susceptible consumers. A better knowledge of the population structure of L. monocytogenes present in retailed food could allow better prevention strategies to be developed. We present the analysis of 450 L. monocytogenes isolates, 179 responsible for sporadic human cases of listeriosis and 271 isolated from foods collected from retailers. All isolates were investigated by multiplex PCR (food isolates), allowing serovar predictions, or serotyped (human isolates), and DNA macrorestriction patterns were determined. Isolates from different sources were significantly differently distributed into PCR groups. PCR group IIa, corresponding to serovars 1/2a and 3a, was predominant in food isolates (58%; OR = 3.19; P < 1 × 10 − 7 ). A larger proportion of human isolates belonged to PCR group IVb, corresponding to serovars 4b, 4d and 4e (44%; OR = 5.69; P < 1 × 10 − 7 ). DNA macrorestriction pattern analysis of PCR group IIa isolates showed that isolates from pork products had a very low diversity (ID = 0.905) whereas isolates from humans were more diverse (ID = 0.976). Furthermore, 78% of the pork product isolates belonging to PCR group IIa exhibited only two AscI profiles, a 1 and a 2, which were very similar (94%). DNA array analysis of representative isolates showed that isolates with a 1 and a 2 profiles constitute a homogeneous population, whereas isolates exhibiting non a 1– a 2 profiles are more diverse. Six of the isolates with a 1 and a 2 profiles were selected and investigated for their gene content using a DNA array. With respect to 295 strains present in our data collection, a specific pattern of the presence and absence of 15 genes was identified. Five are predicted to encode internalins and cell surface proteins, and eight of the genes were missing in this group. They code for cell surface proteins, transcriptional regulators, an acylase, a sugar phosphorylase and proteins of unknown functions. The ability of strains to multiply in different niches may be determined by the presence or absence of these genes.

Use of DNA arrays for the analysis of outbreak-related strains of Listeria monocytogenes

We evaluated the discriminative power and usefulness of the DNA array technology as compared to DNA macrorestriction pattern analysis for monitoring epidemiologically related clusters of Listeria monocytogenes strains that differ slightly in DNA macrorestriction patterns. We show that this approach allows clarifying the genetic basis of the pattern variations. In the reported outbreak, the differences were due to phage excision, showing the power of this technique in epidemiological studies.

Extreme weather events and environmental contamination are associated with case-clusters of melioidosis in the Northern Territory of Australia

Melioidosis, the infection due to the environmental organism Burkholderia pseudomallei, is endemic to northern Australia and South East Asia. It is associated with exposure to mud and pooled surface water, but environmental determinants of this disease are poorly understood. We defined case-clusters in northern Australia, determined their contribution to the observed rate of melioidosis, and explored clinical features and associated environmental factors. Using geographical information systems data, we examined clustering of melioidosis cases in time and geographical space in the Top End of the Northern Territory of Australia between 1990 and 2002 using a scan statistic. DNA macrorestriction analysis, resolved by pulsed field gel electrophoresis, was performed on isolates from patients. We defined five case-clusters involving 27 patients that occurred within 7-28 days and/or a radius of 100-300 km. Clustered cases were associated with extreme weather events or environmental contamination; no difference in the clinical pattern of disease was noted from other patients not involved in clusters. Isolates from patients linked to environmental contamination were caused by isolates with similar DNA macrorestriction patterns, but isolates from patients linked to severe weather events had more diverse DNA macrorestriction patterns. Case-clusters of melioidosis where isolates exhibit diverse DNA macrorestriction patterns in our region are linked to extreme weather events and outbreaks where isolates are predominantly of the same DNA macrorestriction pattern are linked with contamination of an environmental source.

The changing prevalence of drug-resistant Escherichia coli clonal groups in a community: evidence for community outbreaks of urinary tract infections

A multidrug-resistant clonal group (CgA) of Escherichia coli was shown to cause half of all trimethoprim-sulphamethoxazole (TMP-SMZ)-resistant urinary tract infections (UTIs) in a college community between October 1999 and January 2000. This second study was conducted to determine the fate of CgA. Urine E. coli isolates from women with UTI, collected between October 2000 and January 2001, were tested for antibiotic susceptibility, O serogroup, ERIC2 PCR and DNA macrorestriction patterns using pulsed-field gel electrophoresis. The proportion of UTIs caused by CgA declined by 38% (P<0.001) but the prevalence of resistance to TMP-SMZ did not change. Six additional clonal groups were identified and these were responsible for 32% of TMP-SMZ-resistant UTIs. The temporal decline in the proportion of UTIs caused by CgA provides evidence that CgA caused a community outbreak of UTI. The fluctuation and occurrence of other E. coli clonal groups in this community suggest that a proportion of community-acquired UTIs may be caused by E. coli disseminated from one or more point sources.

A new phenotype of resistance to lincosamide and streptogramin A-type antibiotics in Streptococcus agalactiae in New Zealand

To characterize a new type of resistance to clindamycin in Streptococcus agalactiae. Nineteen erythromycin-susceptible, clindamycin-resistant S. agalactiae isolates from New Zealand were studied. MICs of macrolide, lincosamide and streptogramin antibiotics were determined. Clindamycin and streptogramin resistance genes were searched for by PCR. Isolates were compared by serotyping and by DNA macrorestriction patterns determined by PFGE. Conjugative transfer of resistance traits to recipient strains of S. agalactiae and Enterococcus faecium was assayed. The 19 S. agalactiae isolates were intermediate or resistant to clindamycin (MIC range: 0.5-2 mg/L) and lincomycin (MIC range: 1-8 mg/L) and had high MICs of dalfopristin (4-32 mg/L), a streptogramin A-type antibiotic, compared with controls. By contrast, the strains were susceptible to macrolides and quinupristin, a streptogramin B-type antibiotic. This new phenotype was called LSA (lincosamide-streptogramin A). Clindamycin resistance could not be transferred to recipient strains. Thirteen isolates belonged to serotype III and to a single PFGE genotype A, and five isolates belonged to serotype I and to genotype B. One isolate was non-typeable and belonged to a distinct genotype C. We have characterized a new LSA phenotype in S. agalactiae. Analysis of restriction patterns of S. agalactiae chromosomal DNA showed that the resistance was spread in a minimum of three bacterial clones. The genetic and biochemical basis for the resistance remains unknown.

Molecular and Ultrastructural Characterization of Porcine Hippurate-Negative Brachyspira pilosicoli

Brachyspira pilosicoli, the causative agent of porcine intestinal spirochetosis, usually has hippurate-cleaving capacity. We have regularly isolated hippurate-negative B. pilosicoli from cases of porcine diarrhea. In this study, we show that these biochemically atypical B. pilosicoli isolates can be classified as B. pilosicoli. 16S ribosomal DNA was partially sequenced from eight hippurate-negative and two hippurate-positive B. pilosicoli-like isolates from seven herds. The differences in nucleotide sequence with B. pilosicoli P43/6/78 type strain were not associated with hippurate cleavage. In 877 bp, the hippurate-negative isolates had a similarity of 98.63 to 100% to the type strain, with the corresponding figures for the two hippurate-positive isolates being 98.86 and 100%. The nucleotide sequences of hippurate-positive isolates were identical to the respective sequences of hippurate-negative isolates from one herd. The DNA macrorestriction patterns of a total of 20 hippurate-negative and -positive B. pilosicoli isolates were diverse, and no clustering in conjunction with the hippurate reaction was found. In two herds, hippurate-positive and -negative B. pilosicoli isolates had a common macrorestriction pattern. The ultrastructure of hippurate-negative isolates was similar to the type strain. In conclusion, B. pilosicoli can be either hippurate positive or negative and, thus, the scheme for biochemical differentiation of porcine Brachyspira should be revised to include identification of hippurate-negative B. pilosicoli.

Epidemiological relatedness of methicillin-resistant Staphylococcus aureus from a tertiary hospital and a geriatric institution in Spain

From January 2000 to June 2002, 24 Staphylococcus aureus isolates were recovered from decubitus ulcers of patients in a geriatric institution, of which 17 (70.8%) were methicillin-resistant S. aureus (MRSA). Antibiotic resistance and DNA macrorestriction (pulsed-field gel electrophoresis; PFGE) patterns of the MRSA isolates were compared with a collection of 161 MRSA isolates from patients admitted to the institution's reference hospital. PFGE revealed the presence of five clonal types (found also in hospitalised patients) among the 17 MRSA isolates. The findings suggest nosocomial acquisition of the MRSA strains by five patients, with subsequent dissemination of the strains within the institution. The high rate of MRSA highlights the need for epidemiological analysis to control the dissemination of MRSA in long-term care facilities.

Clonal Spread of Imipenem-Resistant Pseudomonas aeruginosa in the Intensive Care Unit of a Turkish Hospital

Pseudomonas aeruginosa may cause life-threatening infections, especially in nosocomial settings. Although carbapenems are considered as one of the most effective alternatives in antipseudomonal therapy, resistance to the car-bapenem group of antibacterials is a growing problem. In the first 6 months of 1997, P. aeruginosa isolates that were resistant to almost all antipseudomonal agents including imipenem were recovered from various specimens from intensive care unit (ICU) patients. Isolates with the same antibiogram profile caused a small outbreak in May 1997. A retrospective case-control study revealed that the major risk factors for infection/colonization with multiresistant P. aeruginosa were prolonged stay in the ICU (p< 0.001), previous and lengthy imipenem usage (p<0.001 and p<0.0001, respectively), and mechanical ventilation (p<0.001). Analytical isoelectric focusing of the sonicates prepared from the isolates showed that each isolate produced 1-5 beta-lacta-mases, enzymes with isoelectric points (pIs) of 5.1, 6.4, 8.5-8.7 being the most prevalent. DNA macrorestriction patterns of imipenem-resistant isolates were distinct from those of the imipenem-sensitive isolates recovered from ICU patients during the same interval and from the environmental isolates (controls). Thus, our results indicate that colonized patients appear to be the major source for cross-contamination of other patients and if imipenem is selected for empirical therapy, emergence of resistant strains should be anticipated and appropriate precautions taken.

A 5-year epidemiological study of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae isolates in a medium- and long-stay neurological unit.

Thirty-eight different strains of extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae (ESBL Kp), isolated from urine and pus samples of 38 patients hospitalized in a medium- and long-stay neurology department between 1 January 1992 and 31 December 1996, were analysed by antibiotic resistance phenotyping, DNA macrorestriction by pulsed-field electrophoresis and isoelectric focusing of beta-lactamases. An epidemiological survey was conducted to identify risk factors for infection by ESBL Kp in this setting. The 38 isolates were distributed into 13 antibiotypes, three of which predominated (13, six and six isolates). The DNA macrorestriction pattern identified 15 genotypes, four of which predominated (11, six, four and four isolates). A combination of the two typing methods revealed several epidemic clones that emerged consecutively. Two main types of ESBL (SHV-2 and CTX-1) were identified by isoelectric focusing, the former predominating. The case-control study showed that the length of hospital stay, degree of malnutrition and dependency, and urinary sphincter status were the main factors significantly associated with ESBL Kp isolation.

Eradication of vancomycin resistant Enterococcus faecium from a paediatric oncology unit and prevalence of colonization in hospitalized and community-based children.

We previously reported an outbreak of vancomycin resistant enterococci (VRE) in a paediatric oncology unit in December 1995 which was associated with widespread environmental contamination of the unit with VRE. We undertook this study to evaluate the effectiveness of the infection control policy instituted subsequent to the outbreak and to investigate the underlying prevalence of VRE colonization in hospitalized, outpatient and community-based children. We sought to establish the molecular similarity of VRE isolates from the study. Stool specimens were obtained from outpatients at risk of VRE, hospital inpatients and from healthy community-based children. VRE colonization was eradicated from the inpatient unit within 11 months, but in outpatients, 16 months after the outbreak, 4 of 137 (2.9 %) attending oncology outpatients, 5 of 65 (7.7%) with cystic fibrosis and 1 of 12 (8.3 %) with liver disease were found to be colonized with VRE. The isolates were all Enterococcus faecium, Van A phenotype except one E. casseliflavus of the Van C phenotype. All were unique in SmaI DNA macrorestriction patterns with the exception of two isolates, which were similar to the original outbreak strain and three further isolates of a single strain but which differed from the outbreak strain. Of 315 hospital inpatients, 2.5 % were colonized with VRE of the Van C resistance phenotype but VRE was not detected in 116 healthy, community-based children. We conclude that effective strategies can successfully control spread of VRE but despite a low prevalence of VRE colonization in hospital patients and in community-based children, outbreaks can occur when infection control practices are not optimal. Continued vigilance to detect VRE and limit spread within hospitals is therefore necessary.

Differences in genomic macrorestriction patterns of arabinose-positive (Burkholderia thailandensis) and arabinose-negative Burkholderia pseudomallei.

We reported previously two biochemically and antigenically distinct biotypes of Burkholderia pseudomallei. These two distinct biotypes could be distinguished by their ability to assimilate L-arabinose. Some B. pseudomallei isolated from soil samples could utilize this substrate (Ara+), whereas the other soil isolates and all clinical isolates could not (Ara-). Only the Ara isolates were virulent in animals and reacted with monoclonal antibody directed at the surface envelope, most likely the exopolysaccharide component. In the present study, pulsed-field gel electrophoresis was employed for karyotyping of these previously identified B. pseudomallei strains. We demonstrate here that the DNA macrorestriction pattern allows the differentiation between B. pseudomallei, which can assimilate L-arabinose, and the proposed B. thailandensis, which cannot do so. Bacterial strains from 80 melioidosis patients and 33 soil samples were examined by genomic DNA digestion with NcoI. Two major reproducible restriction patterns were observed. All clinical (Ara-) isolates and 9 Ara- soil isolates exhibited macrorestriction pattern I (MPI), while 24 soil isolates (Ara+) from central and northeastern Thailand displayed macrorestriction pattern II (MPII). The study here demonstrated pulsed-field gel electrophoresis to be a useful tool in epidemiological investigation possibly distinguishing virulent B. pseudomallei from avirulent B. thailandensis or even identifying closely related species of Burkholderia.

Use of pulse field gel electrophoresis for the epidemiological characterisation of coagulase positive Staphylococcus isolated from meat workers and beef carcasses

Coagulase positive staphylococci isolated at an Australian abattoir, from beef carcasses, hands, and environmental samples, were typed by the DNA macrorestriction patterns obtained following PFGE of SmaI digests. The predominant PFGE pattern of isolates collected before evisceration was different from the dominant pattern of isolates collected after evisceration. PFGE patterns for isolates from workers’ hands and from concordant carcasses were indistinguishable. PFGE types among isolates collected from non-evisceration abattoir personnel and clerical staff were distinct from patterns among isolates collected from the slaughter floor personnel. Twenty-nine (85%) of the 34 isolates collected from carcasses after evisceration exhibited PFGE patterns that were 93% homologous. During evisceration the carcasses were handled extensively and it is proposed that the hands of workers at this abattoir are the primary source of staphylococcal contamination of carcasses on the slaughter floor. This paper highlights the usefulness of PFGE as an epidemiological tool for determining the source of staphylococcal contamination in the food industry.

Evaluation of an rDNA Listeria probe for Listeria monocytogenes typing

A Listeria monocytogenes DNA fragment, identified as part of the 23S rRNA gene and called B17, was used to type 266 L. monocytogenes strains and 43 strains of other Listeria species. Results were compared with those obtained: i) with pBA2 (which consists of a 2.3 kb Bacillus subtilis DNA fragment encoding 16S rRNA, inserted into the HindIII site of pBR322), a probe previously used for Listeria and L. monocytogenes ribotyping, and ii) with DNA macrorestriction profiles analysis. Twenty profiles were identified for L. monocytogenes using pB 17, three of which accounted for 87% of strains. This new rDNA probe had greater discriminatory power for serogroups 1/2 or 3 strains than for serogroup 4 strains. The number of varieties and the discrimination index were higher with this new probe than with pBA2, but DNA macrorestriction patterns analysis gave better discrimination between strains.