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In Argentina, hemolytic uremic syndrome (HUS) caused by Shiga toxin-producing Escherichia coli (STEC-HUS) infection is endemic, and reliable data about prevalence and risk factors have been available since 2000. However, information about STEC-associated bloody diarrhea (BD) is limited. A prospective study was performed during the period October 2018-June 2019 in seven tertiary-hospitals and 18 referral units from different regions, aiming to determine (i) the frequency of STEC-positive BD cases in 714 children aged 1-9 years of age and (ii) the rate of progression of bloody diarrhea to HUS. The number and regional distribution of STEC-HUS cases in the same hospitals and during the same period were also assessed. Twenty-nine (4.1%) of the BD patients were STEC-positive, as determined by the Shiga Toxin Quik Chek (STQC) test and/or the multiplex polymerase chain reaction (mPCR) assay. The highest frequencies were found in the Southern region (Neuquén, 8.7%; Bahía Blanca, 7.9%), in children between 12 and 23 month of age (8.8%), during summertime. Four (13.8%) cases progressed to HUS, three to nine days after diarrhea onset. Twenty-seven STEC-HUS in children under 5 years of age (77.8%) were enrolled, 51.9% were female; 44% were Stx-positive by STQC and all by mPCR. The most common serotypes were O157:H7 and O145:H28 and the prevalent genotypes, both among BD and HUS cases, were stx 2a -only or -associated. Considering the endemic behavior of HUS and its high incidence, these data show that the rate of STEC-positive cases is low among BD patients. However, the early recognition of STEC-positive cases is important for patient monitoring and initiation of supportive treatment., (Copyright © 2023 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Shiga toxin-producing Escherichia coli (STEC) are a heterogeneous group of foodborne pathogens causing a broad spectrum of human disease, from uncomplicated diarrhea to hemolytic uremic syndrome (HUS). In this study, we report an HUS case associated with an O59:NM[H19] strain, harboring stx 2a , iha, lpfA O26 , lpfA O113 genes associated with STEC, and aatA, aap, pic, sigA, agg4A genes associated with enteroaggregative E. coli (EAEC), named Stx-EAEC. The strain showed low toxicity on Vero cells, and was resistant to streptomycin and trimethoprim/sulfonamides. The child carried the bacteria for more than 100 days. Since the large outbreak associated with Stx-EAEC O104:H4, many strains with similar profiles have been described. In Germany, an O59:NM[H19] strain, with comparable characteristics to the Argentine strain, was isolated from a bloody diarrhea case. In Argentina, this is the first report of an HUS case associated with a Stx-EAEC infection, and represents a new challenge for the surveillance system., (Copyright © 2019. Publicado por Elsevier España, S.L.U.)

On November 27, 2008, a foodborne disease outbreak associated with the consumption of chicken salad occurred in a kindergarten in the District of Hurlingham, Province of Buenos Aires. Thirty-seven children and 10 adults with gastrointestinal symptoms were affected. Five children were hospitalized with signs of dehydration, one of them requiring intensive care. Staphylococcus aureus subsp. aureus was isolated from the mentioned food in 4 out of 5 stool specimens from the patients, and in 3 out of 5 food handlers (nose of food handler #1, hands of food handlers #2 and 3). The isolates carried the genes coding for enterotoxins SEA and SED. The macrorestriction patterns showed 100% similarity by pulsed-field gel electrophoresis using the SmaI enzyme. A timely outbreak investigation allowed us to identify the causative agent of the food poisoning as well as the failures in food processing and to implement corrective measures., (Publicado por Elsevier España, S.L.U.)

The objectives of this study were: (1) to estimate STEC frequency in hide and carcass samples taken from beef slaughterhouses supplying the domestic market in Argentina, (2) to establish the pheno-genotypic characteristics of STEC and non-toxigenic Escherichia coli of serogroups O26, O45, O103, O121, O111, O145 or O157 isolated from the analyzed samples and, (3) to study their clonal relatedness. Sixty hides and 60 carcasses were analyzed. At the screening step, 48% of hide and 80% of carcass samples tested positive for the stx gene by endpoint PCR. The STEC isolation rate was 5% for hides and 8% for carcasses. The isolation rate of STEC-positive for O26, O45, O103, O111, O145 or O157 serogroups was 0% for hides and 2% for carcasses. With the purpose of studying the clonal relatedness of isolates, macrorestriction fragment analysis by pulsed-field gel electrophoresis was performed. The results indicated cross-contamination between hides and between carcasses of animals in the same lot and, that the origin of carcass contamination was their own hide, or the hides of other animals in the same lot. The high detection rate at the screening step, especially in carcasses, and the evidence of cross-contamination show the need to apply additional in-plant intervention strategies aimed at preventing carcass contamination., (Copyright © 2018 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Cattle are the main reservoir of Shiga toxin-producing Escherichia coli (STEC), and the strategies to prevent the transmission of these microorganisms are concentrated in the slaughtering plant. The aim of this study was to evaluate the hygienic-sanitary quality and the frequency of detection of STEC in beef carcasses in abattoirs from Tucuman province. Two hundred and seventy four beef carcass sponges were processed; the count of generic E. coli was marginal in 9 (3,3%) of them. Escherichia coli O157 was isolated in 4 (1,4%) samples; 2 of which were characterized as stx 2c(vh-a) /eae/ehxA whereas the other 2 were non-toxigenic strains. Non-O157 E. coli ONT:H49, stx 2a /ehxA/saa was isolated from 1 sample (0,4%). In this work the quality of the analyzed product indicates that the good practices of manufacture are fulfilled in slaughtering facilities in Tucumán province., (Copyright © 2017 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Enteropathogenic Escherichia coli (EPEC) is a major causative agent of acute diarrhea in children in developing countries. This pathotype is divided into typical EPEC (tEPEC) and atypical EPEC (aEPEC), based on the presence of the bfp virulence factor associated with adhesion, encoded in the pEAF plasmid. In the present study, the isolation of aEPEC O157:H16 from a bloody diarrhea case in a child and his household contacts (mother, father and sister) is described. The strain was characterized as E. coli O157:H16 eae-ɛ-positive, sorbitol fermenter with β-glucuronidase activity, susceptible to all antimicrobials tested, and negative for virulence factors stx1, stx2, ehxA and bfp. XbaI-PFGE performed on all isolates showed the AREXHX01.1040 macrorestriction pattern, with 100% similarity. These results highlight the importance of epidemiological surveillance of E. coli O157-associated diarrhea cases identified in children and their family contacts, as well as the incorporation of molecular techniques that allow the detection of the different E. coli pathotypes., (Copyright © 2015 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Groundwater is an important source of drinking water for many communities in Northern Argentina; particularly, in the province of Chaco, where about 14% of households use this natural resource. Enteroaggregative Escherichia coli is an emerging pathogen whose global importance in public health has increased in recent years. Despite the significant risk of disease linked to contaminated water exposure, the prevalence of E. coli pathotypes in aquatic environments is still not so well defined. The aim of the present study was to detect the presence of typical enteroaggregative E. coli through the recognition of its virulence factors aap, AA probe and aggR by molecular techniques. A total of 93 water samples from different small communities of Chaco were analyzed. E. coli was identified in 36 (38.7%) of the tested samples. Six strains isolated from different samples harbored the studied genes. Of these 6 isolates, 3 carried the aap gene, 2 the AA probe and the last one the combination of aap/aggR genes. The prevalence of E. coli isolates harboring enteroaggregative virulence genes in groundwater sources was 6.4%. This work represents the first contribution to the study of the presence and distribution of virulence genes of EAEC in groundwater sources in this region of Argentina., (Copyright © 2014 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Escherichia coli O157 is an emergent pathogen associated with diarrhea, hemorrhagic colitis and hemolytic uremic syndrome. Meat products constitute an important transmission source of this microorganism. The aims of this study were to characterize E. coli O157 isolated from cattle and meat products collected from abattoirs and retail stores, to establish the clonal relatedness among regional isolates and to compare them with those in the national database. Between 2004 and 2013, 169 minced meat, 35 sausage and 216 carcass samples were analyzed. Thirteen E. coli O157 isolates were identified; 6 of which were O157:H7 and characterized as stx2c(vh-a)/eae/ehxA (n = 5) and stx2/eae/ehxA (n = 1). The 7 remaining isolates were non-toxigenic E. coli strains, and serotyped as O157:NT (n = 4), O157:NM (n = 1), O157:ND (n = 1) and O157:H16 (n = 1). The strains yielded different XbaI-PFGE patterns. Compared to the E. coli O157 isolates in the National Database, none of these patterns have been previously detected in strains of different origin in Argentina., (Copyright © 2014 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

We describe the first isolation of an enteroaggregative Escherichia coli (EAEC) O104:H4 strain associated with an acute diarrhea case in Argentina. Two multiplex PCRs (mPCR) were performed as screening of genes mPCR1 (eae, lt, and st) and mPCR2 (IpaH, aggR, stx1 and stx2). A mPCR to detect the rfbO104, fliCH4 and terD genes, and PCR assays for the detection of pCVD432 plasmid, aaiC and lpfO113 genes were included. Biochemical and antimicrobial susceptibility assays as well as serotyping were performed. The identified E. coli strain was susceptible to all antimicrobials tested and harbored the aggR, aaiC, pCVD432 plasmid, lpfO113, rfbO104, fliCH4 and terD genes. Although serotype EAEC O104:H4 rarely spreads and sporadic cases have been reported, global concern increased after the large-scale outbreak in Europe in 2011. The finding of EAEC O104:H4 reinforces the need for improved methodologies for the detection of all E. coli pathotypes., (Copyright © 2014 Asociación Argentina de Microbiología. Publicado por Elsevier España. All rights reserved.)

STEC strains can infect extra-intestinal sites such as the human urinary tract and sometimes cause severe complications. We report two cases of urinary tract infection caused by STEC in two elderly women with comorbidities. Although both strains belonged to the O157:H7 serotype and carried genes associated with severe illness, none of the patients developed hemolytic uremic syndrome (HUS). These findings provide additional evidence for the presence of these agents in our country and in the region, and highlight the need to maintain an active surveillance system of HUS cases, placing special emphasis on the study of other sites of infection in patients with non-diarrheal HUS.

During March 2007 there was an epidemic outbreak associated with the consumption of ultrapasteurized milk. Four hundred people were affected and 60 required hospitalization. S. aureus subspecies aureus was isolated from 5 patients, 3 operators and 3 milk samples. All strains produced enterotoxins. Strains isolated from 3 patients, one operator and all the milk samples carried the genes encoding enterotoxins C (sec) and D (sed), and showed an indistinguishable macrorestriction pattern (SmaI-PFGE). Milk was identified as the source of intoxication and a production line operator as the source of contamination. This is the first foodborne outbreak reported in Paraguay whose agent was isolated, characterized and subtypified in the production plant, the food and the affected people.

Multiplex PCR for the detection of sea, seb, sec, sed and see genes of Staphylococcus aureus. Characterization of isolates from food. The presence of Staphylococcus aureus in food represents a potential risk to public health, being its enterotoxins the major virulence factor. Enterotoxin detection can be determined by ELISA, but only for the pool of enterotoxins SEA, SEB, SEC, SED and SEE. The main aims of this study were to optimize two PCR techniques for detection of S. aureus sea, seb, sec, sed and see, and to characterize Staphylococcus spp. isolates associated with food intoxication. Two PCR techniques were optimized and 115 Staphylococcus spp. isolates from Ciudad Aut6noma de Buenos Aires, and Buenos Aires, Córdoba, and Neuquén provinces were characterized. The characterization was performed by biochemical tests, ELISA and PCR. Sixty-eight isolates (59.1%) were positive by ELISA, while 61 (53%) were positive by PCR. Out of the positive PCR isolates, 34 (55.7%) carried the sea gene, 9 (14.8%) the seb gene, 5 (8.1%) the see gene, 4 (6.5%) the sec gene, 6 (9.9%) were positive for sea and seb genes, 2 (3.3%) for sea and sec genes, and 1 (1.7%) for sea and sedgenes. This is the first study of genotypic characterization of S. aureus isolates associated with food intoxication from different provinces of Argentina.

Diarrheal disease continues to be a serious health problem, especially in developing countries. Bloody diarrhea represents approximately 20-30% of all cases and has higher morbidity and mortality. Treatment with antibiotics is beneficial in cases of Shigella, Campylobacter, Yersinia and Salmonella infection, principally in those children with a higher risk of invasive disease. The aims of this study were to detect the bacterial agents associated with bloody diarrhea in children and to determine their antimicrobial susceptibility patterns. Between June 2001 and January 2008, 249 children with bloody diarrhea were studied. Shigella and Shiga toxin-producing Escherichia coli (STEC) were recovered from 48 (19.3%) and 3 (1.2%) of the total of cases, respectively. In 49 out of 249 children, in whom other enteropathogens were investigated, we recovered Campylobacter jejuni from 7 children (14.3%), Salmonella spp. from 2 (4.1%) and Aeromonas spp. from 1 (2%) in addition to Shigella from 7 children (14.3%). Thirty-four (70%) Shigella isolates showed resistance to ampicillin and 13 (27%) to trimethoprim-sulfamethoxazole. All Shigella isolates were susceptible to nalidixic acid, ciprofloxacin and ceftriaxone. Salmonella and STEC isolates were susceptible to all antibiotics assayed. Thus, the use of trimethoprim-sulfamethoxazole or ampicillin would not be appropriate for the empirical treatment of Shigella - associated diarrhea.

In the summer of 2006, an epidemic outbreak of acute gastrointestinal illness related to food consumption occurred in a small town in the province of Neuquén, Argentina. During a popular feast, approximately 800 local residents attended lunch held in the facilities of the Municipal Gymnasium. About three hours later, nearly 150 attendees sought medical assistance at the local hospital due to acute gastroenteritis. A case-control epidemiological investigation was conducted using representative non-probability sampling. The epidemiological investigation showed a common-source foodborne disease outbreak with a case-control ratio of 1:1.8. The main symptoms were abdominal cramps (88%), vomiting (73.5%) and diarrhea (60%). The cake was identified as the source of infection (OR 9.79; IC 95%, 2.66-36.00; p = 0.0001), and unsatisfactory hygienic conditions in food production, conservation and handling steps were identified. Coagulase positive, enterotoxigenic Staphylococcus aureus, subspecies aureus was detected in a piece of cake, with a count of 2.4 x 10(6) CFU/g, and in samples from the hands and nostrils of three people involved in food preparation and service. The strains isolated from both the cake and one of the food handlers carried the sea gene, and presented the same Smal-PFGE pattern. The foodborne disease outbreak was considered to be due to contamination in the preparation process of the cake consumed at the feast, which was related to inadequate hygienic conditions, lack of refrigeration and cold chain disruption.

We have assessed the frequency of Shiga toxin-producing Escherichia coil (STEC) in clinical and food samples as well as studied the genotypic and phenotypic characteristics of the recovered strains. One hundred ninety eight fecal samples from children with bloody diarrhea (BD), 14 from children with hemolytic uremic syndrome (HUS), 220 ground beef samples and 4 STEC isolates from other beef-derived products were analyzed. The STEC strains were isolated from 3 (1.5%) children with bloody diarrhea, 1 (7%) from a child with HUS and 4 (1.8%) from ground beef samples. All strains were eae and ehxA positive. The serotypes found were: O157:H7 (9 strains), O26:H11 (2), O111: NM (1) and O145:HNT (1). All O157:H7 STEC strains harbored the eae subtype gamma1, O26:H11 and O145:HNT strains, subtype beta1 and O111:NM strain, subtype gamma2/theta. The STEC strains of the same serogroup showed high genetic diversity. In Uruguay, STEC is not frequently isolated from cases of bloody diarrhea in children. However, all the recovered STEC strains carried the genes associated with severe disease and 2 out of 3 children infected with STEC developed HUS. Ground beef and other food products might be important vehicles for O157:H7 strains.

Shiga toxin-producing Escherichia coli is an emergent pathogen, being the Shiga toxin (Stx) the main virulence factor. These toxins are classified into 6 types (1,2, 2c, 2d, 2e and 2f) and 22 variants. In Argentina, two PCR for stx gene detection, PCR-MK and multiplex-PCR, were validated. The aim of this work was to analyze, by using bioinformatic tools, the stx variants that could be amplified by these PCRs, and to experimentally show the amplification of 8 stx variants. Twenty-five nucleotide sequences were collected from GenBank corresponding to 21 stx variants. The BLAST 2 sequences program was used to analyze the complementarities between the nucleotide sequence of the variants and the primers corresponding to the PCR studied. PCR-MK could detect types stx1, stx2, stx2c, stx2d and stx2f, but not type stx2e and three type stx2c variants. On the other hand, the multiplex-PCR could detect types stx1, stx2, stx2c, stx2d, but not stx2e and stx2f types. It was experimentally determined that both PCRs can detect those variants that cause severe disease in humans.

Shiga toxin (Stx)-producing Escherichia coli (STEC) is an emergent pathogen associated with foodborne diseases, especially foodstuffs of animal origin. A total of 250 beef samples (ground beef and hamburgers) obtained from retail outlets in Santa Fe and Santo Tomé cities, and 150 milk samples from bulk tank milk from dairy barns of the region were analyzed by selective enrichment and immunomagnetic separation. Escherichia coli O157:H7 stx2, eae and ehxA positive strains were isolated from three (1.2%) beef samples. The strains could be differentiated by pulsed-field gel electrophoresis, phagetyping and genotyping of stx. The milk samples were negative for STEC O157. These findings confirm the role of food of animal origin in the epidemiology of E. coli O157:H7 - associated diseases.

In this report we describe the detection and duration of fecal shedding of Shiga toxin-producing Escherichia coil (STEC) O157 and non-O157 in symptomatic and asymptomatic cases during four events occurred among children in day-care centers in Argentina. In each event, the cases were identified among children, family contacts and staff members of the Institution. The isolates were characterized by pheno-genotyping and subtyping methods. The STEC fecal shedding was prolonged and intermittent. Strains O157:H7 (1st event); O26:H11 (2nd event); O26:H11 (3rd event) and O145:NM (4th event) were shed during 23-30, 37, 31 and 19 days, respectively. Considering the possibility of STEC intermittent long-term shedding, symptomatic and asymptomatic individuals should be excluded from the Institution until two consecutive stool cultures obtained at least 48 h apart, test negative.

Typeability, reproducibility, and discriminatory power of ERIC-PCR and Apal-PFGE to establish the genetic relation of P. multocida strains were determined. Forty-nine strains of different source, biotype, capsular group, somatic serotype, and resistance to antimicrobials were studied. By ERIC-PCR, 31 patterns were defined with 10 to 14 bands in a rank of 0.2 and 1.2 kb. By Apal-PFGE, 37 restriction patterns were established with 7 to 15 bands of 34 to 450 kb. Typeability was 100% (T=1) for ERIC-PCR, and 94% (T = 0.94) for Apal-PFGE. Reproducibility of both techniques was 100% (R=1). Discriminatory power was 93% (D = 0.93) for ERIC-PCR, and 98% (D = 0.98) for Apal-PFGE. By using both techniques, epidemiologically related strains were grouped, and unrelated strains were clearly differentiated. The value of ERIC-PCR and Apal-PFGE as complements to epidemiologic studies was demonstrated, especially when both techniques were used to analyze the strains.

Thirty Pasteurella multocida strains isolated in Argentina from human and animal samples were identified, biotypified and characterized. Twenty-two (73%) strains were identified as P. multocida subsp. multocida, 5 (17%) as P. multocida subsp. gallicida, and 3 (10%) as P. multocida subsp. septica. All strains were grouped in 8 biotypes, and 70% of the strains presented capsular type A. The most frequent somatic serotypes were 1 (n:11) and 3 (n:9). P. multocida strains from swine source were resistant to tiamulin, streptomycin and tetracycline. Characterization of P. multocida strains isolated in Argentina is the first step to conduct future studies intended for the prevention and treatment of pasteurellosis in human and veterinary medicine.

From October 15 to November 8, 2003, a gastrointestinal outbreak occurred at a day care center in a Hospital in Mar del Plata City. Fourteen out of 80 (17.5%) children, mean age 23.6 +/- 13.9 months, and the mother of one of them had diarrhea. One case developed hemolytic uremic syndrome. No conclusive evidence of the origin of the outbreak was found, but the epidemic curve suggested person-to-person spread. The usual practices at the place where infant milk formula was prepared at the day care center, together with the inadequate infrastructure conditions and hygiene practices at the kitchen of the hospital, were considered risk factors. One case had Shiga toxin-producing Escherichia coli (STEC) O103:H2 infection and other STEC O26:H11. The duration of shedding for the child with O26:H11 infection was 37 days. In the other symptomatic children, the pathogen was not recovered from fecal samples collected 6 or more days after the onset of the illness. This emphasizes that the collection of early samples is necessary to recover STEC strains. In order to prevent and control enteric diseases in day care facilities the following measures are necessary: optimal hygiene standards, early case reporting, and exclusion of those who remain culture-positive.

Shiga toxin-producing Escherichia coli (STEC) cause non-bloody or bloody diarrhea, hemorrhagic colitis and hemolytic uremic syndrome (HUS) in humans. The aim of the present study was to validate a multiplex PCR for the STEC diagnosis based on the detection of stx1, stx2 and rfbO157 genes. The multiplex PCR validation was carried out in two independent laboratories in a parallel way. Work range, selectivity and robustness were established. The PCR performance was evaluated using different concentrations of two STEC strains harboring different target genes. The work range depended on the strain analyzed, the maximum and the minimum values were 6.6 x 10(7) and 1.0 x 10(4) CFU/50 microl. The detection limit was 1.0 x 10(4) CFU/50 microl and the cut limit 1.0 x 10(5) CFU/50 ml. A good robustness was observed when different variables were introduced. Inclusivity, exclusivity, positive predictivity, negative predictivity and analytical accuracy were of 100%. Interference was not shown when different concentrations of STEC strains, carrying different genes, were used. The validated technique is an appropriate alternative for detection and confirmation of STEC O157 and non-O157 strains from bacterial cultures.

Culture media, reagents, and commercial kits were compared on artificially contaminated food samples. The objective was to find an isolation method for Escherichia coli O157:H7 sensitive, specific and accessible in terms of cost, requirements of equipments and qualification of the analyst. The adopted scheme consisted in a selective enrichment at 42 degrees C during 18 to 24 h, using an appropriate medium, in accordance with the nature of the sample, followed by a step of immunomagnetic separation and simultaneous isolation on a chromogenic agar and MacConkey sorbitol agar with potassium tellurite and cefixime, during 18-24 h at 37 degrees C. The presumptive colonies were confirmed as E. coli O157 by serological and biochemical tests. Secondly, this methodology was applied to food samples, water, bovine gastric content and manure. A total of 410 samples were studied: 279 from meat, 54 milk and dairy products, 6 from vegetables, 27 water samples and 44 bovine gastric content and manure. The frequency of isolation of E. coli O157:H7 was of 3.9%. The phenotypic and genotypic characterization of the isolates was performed. A simple isolation methodology for E. coli O157 was developed, which proved accessible to food laboratories of lower complexity. This methodology allowed the detection of this pathogen in food and environmental samples in Gualeguaychú City. The role of water as vehicle of infection was also established. The strains harbored the same virulence factors as those recovered from human disease.

Shiga toxin producing-Escherichia coli (STEC), an important emerging foodborne pathogen, has been associated with bloody and non-bloody diarrhea, hemorrhagic colitis, hemolytic uremic syndrome (HUS) and thrombotic thrombocytopenic purpura. The cattle have been shown to be a major reservoir of STEC and raw foods such as ground beef and milk are the most common vehicles of infection. In the present study, the prevalence of STEC in 95 samples of frozen hamburgers and in 114 samples of soft cheese was established in 8.4% and 0.9%, respectively. The genotypic and phenotypic characteristics of the strains were determined. The virulence genes stx1, stx2, eaeA and EHEC-hlyA were identified by PCR and by colony blot hybridization assays. Serotyping, antimicrobial susceptibility and production of Stx using specific cytotoxicity assays on Vero cells were also determined. All STEC strains were characterized as eaeA-/EHEC-hlyA+. The stx2 genotype was prevalent (77.8%), and four different O:H serotypes were found, comprising: O8:H19 (5 strains), O113:H21 (1), O8:H16 (1), and O39:H49 (1). One STEC strain was nontypable. Although soft cheese complimented the microbiological quality controls for the coliform counts, the detection of STEC in one sample raises doubts concerning the effectiveness of the current quality controls. These data contribute to the implementation of strategies for the prevention and control of HUS.

The present study was focused on the isolation and characterization of Escherichia coli, particularly the serotype O157:H7, from five combined-sewer outflows waters, which drain into the beaches of Mar del Plata. Seventeen hemolytic uremic syndrome cases were reported in Mar del Plata during the sampling period (May 1995-April 1996) in children between 9 month- and 5 year-old, and 3 deaths were recorded. E. coli identification was carried out by biochemical tests. E. coli was detected in 75% of the samples and a total of 98 strains were selected, with 11 sorbol non-fermenting strains. The strains belonged to the O1, O6, O44, O86a, O119 and O168 serogroups. As none of the known virulence factors was detected, strains could not be grouped within any of the diarrheagenic E. coli categories. None of the E. coli strains belonged to the O157:H7 serotype, but E. coli isolation showed fecal contamination in the combined-sewer outflows. Since their waters drain into beaches for recreational use, it is necessary to emphasize the detection of E. coli that would cause severe human illness. Bacterial pollution in combined-sewer outflows draining into Mar del Plata coasts might represent a high risk for human health.

We studied the differential kinetic patterns for Shiga toxin (Stx) production (i.e. Stx1, Stx2 and Stx2c) in different reference Escherichia coli strains and in those isolated from hemolytic uremic syndrome (HUS) patients. These results were correlated with those obtained by specific cytotoxic activity assays on Vero cells and hybridization tests with DNA probes for Stx1 and Stx2. Strains cultured in Penassay broth were sampled at 1.5; 3; 5; 9 and 24 hours to determine bacterial growth and its association with cell-bound and free cytotoxicity. Stx1 showed an intracellular/extracellular concentration ratio (ic/ec) between 32 and 200 times after 3 h-growth. At 24 h both Stx1 concentrations were equal or, in some strains, the ec resulted 2-fold higher that the ic. The ic-Stx1 was equal or just 2-fold higher that ec after 3 h-growth. However, at 24 h the released toxin level was 16 to 32 times higher that cell-bound toxin. The ec-Stx2c increased logarithmically, with maximal yields at 5 h, remaining constant up to 24 h. At that time ic-toxin was 2-fold higher than the released one. When the same experiments were performed on strains isolated from HUS patients they showed that the kinetic patterns obtained corresponded to Stx2. These results were confirmed by hybridization assays. In this study we have shown that Stx1 production decreases dramatically during stationary phase while Stx2 is detected at high level at that time. This could explain the higher frequency of association of Stx2-producing E. coli strains and HUS in some countries, including Argentina.

Forty-one sporadic cases of non-O group 1 Vibrio cholerae gastroenteritis were detected in Orán, Salta, between February 1992 and February 1995. The frequency of isolation was 0.9% of the diarrhea cases. Out of 41 patients, 21 (51.2%) were older than 15 years and 25 (60.9%) were male. All the patients had diarrhea, 24 (58.5%) had watery stools and 6 (14.6%) cholera-like diarrhea; 10 (24.4%) presented vomiting and 12 (29%) mild dehydration. Six malnourished children who suffered from diarrhea with moderate dehydration for more than a week, were hospitalized. V. cholerae non O1 and Shigella flexneri were isolated from one patient, during the first outbreak and V. cholerae non O1 and Salmonella IV 50:b:- were recovered simultaneously from another patient during the fourth outbreak. A 72 year old woman died during the second cholera outbreak. The symptoms were: watery diarrhea, vomiting, fever and mild dehydration. A strain of V. cholerae O5, that did not produce cholera toxin, heat-stable enterotoxin, Kanagawa-like hemolysin or verocitotoxin was detected. It was positive for El Tor hemolysin and D-mannose and L-fucose resistant cells-associated hemagglutinins. Among the 41 isolates studied, all were oxidase and indole positive, fermented glucose, saccharose and mannitol. They were all motile, produced lysine and ornithine decarboxylases but not arginine dihydrolase or hydrogen sulfide. They were sensitive to O129 vibriostatic compound. None of them belonged to O1 or O139 serogroup and they did not produce cholera troxin. Among the V. cholerae non O1 strains isolated, 9.5% were resistant to ampicillin and 4.9% to trimethoprim-sulfamethoxazole. Active surveillance had shown that V. cholerae non-O1 is not an important agent of diarrhea in Orán, Salta.

Sixteen Escherichia coli strains isolated from diarrhea patients, characterized as enteroinvasive (EIEC) by the keratoconjunctivitis test (Sereny test), and 15 non-EIEC strains, were studied. The aim of this study was to establish a possible correlation between the Sereny test and several other tests as enzyme-linked immunosorbent assay (ELISA) and biochemical markers, which are more easily accessible to general practice laboratories. Strains were assayed for the virulence marker antigen (the outer membrane proteins) by direct ELISA, biochemical tests, antimicrobial susceptibility profile and serotyping. Mean optical density (OD) values in the ELISA were 2.018 +/- 0.336 for EIEC strains, and 0.569 +/- 0.283 for non-EIEC strains. Difference between the OD values of the two groups was found to be significant at p < 0.01 by the one-tailed Student's test. One (6.2%) of the 16 Sereny test-positive strains was negative by ELISA. Considered as a group, the EIEC strains are rather heterogeneous in their biochemical characteristics. Lysine decarboxylase test appeared as the most relevant assay, since all EIEC strains were negative for this enzyme, whereas 66.7% of non-EIEC strains were positive. Other helpful tests performed for strain characterization were: gas production from glucose and from glycerol, sucrose and salicin fermentation, and esculin hydrolysis (p < 0.01). However, strains behaviour was variable with these tests. Differences between both groups of strains with regard to mucate utilization and dulcitol fermentation was significative only to a 5% level. Antibiotic sensitivity patterns showed differences between EIEC and non-EIEC strains: 37.5% of the former vs 80% of the latter, were resistant to at least four of the antibiotics tested.(ABSTRACT TRUNCATED AT 250 WORDS)

Eighty-one morphologic and biochemical features of seventy Bacillus strains were studied. Techniques of Numerical Taxonomy were used for this purpose. One method of coding and four others of clustering methods were used. Clustering methods were: weighted an unweighted pair-groups methods using arithmetic averages (WPGMA and UPGMA, respectively) and those of simple and complete linkage. With this methodology four dendrograms were obtained. The highest cophenetic correlation coefficient (0,718) was obtained from UPGMA. With UPGMA a classification in eleven groups and fourteen subgroups was obtained. Group was defined as the cluster of strains linked with similarity coefficients greater than 0.700 and subgroup as the cluster of strains linked with similarity coefficients greater than 0.800. It is important to take into account that in WPGMA dendrogram a very similar distribution was observed. As in classical taxonomy, the characteristic morphologic and physiological variabilities of the genus Bacillus were observed.

The efficiency of the pooling method for the detection of enterotoxigenic Escherichia coli (ETEC) strains associated with children acute diarrhea, was evaluated. This study involved the analysis of 6989 E. coli strains corresponding to 1485 cases, coming from 7 hospitals at different geographic locations. Three to five strains from each case were inoculated in pool in Casamino-acids-yeast extract-salts medium plus lincomycin (30 micrograms/ml) and incubated at 37 degrees C during 18 hours with shaking. Polymyxin sulphate B (2200 U/ml) was added to the cultures, and incubation with shaking continued for additional 30 minutes. The culture was then centrifuged and the supernatant tested by a) the enzyme-linked immunosorbent assay to detect heat-labile enterotoxin (LT) and b) the suckling mouse assay for the detection of heat-stable enterotoxin (ST). Strains from a pool were individually studied in all positive cases as shown by the pooling methodology. Also, in 1 out of every 15 negative cases by the pool method, their component strains were individually analyzed, to confirm that there were no false negatives. Fifty seven LT-ETEC, 61 ST-ETEC and 15 LT-ST cases were detected. From the 89 negatives selected by the pool method, 2 cases were positive when the component strains were individually tested (p1 = 2.25%, 95% confidence interval from 0.27 to 7.88%). In both cases one of the non ETEC strains inhibited the ETEC strain. In other 5 negative cases (p2 = 5.62%, 95% confidence interval from 1.85% to 12.63%), ETEC strains were detected when analyzed individually, and gave a positive result when the pooling method was repeated.(ABSTRACT TRUNCATED AT 250 WORDS)

Numerical Taxonomy's techniques were employed on twenty one anaerobic Gram positive bacilli strains taking account of eighty morphologic and biochemical features. Two methods of coding (weighted and non weighted) and four clustering methods (unweighted pair-group method using arithmetic averages; weighted pair-group method using arithmetic averages; single linkage and complete linkage) were used. Eight dendrograms were obtained and they showed that C. haemolyticum (strains 1, 2, 4, 7); C. subterminale (strains 8, 19); C. botulinum (strain 14) and C. sporogenes (strain 15) belong to a different group than the other strains. Another group would be formed with C. perfringens (strains 3, 5, 6, 12, 16, 21) and C. paraperfringens (strains 9, 10, 11). Of the remaining strains the two classified as Propionibacterium acnes (strains 17, 18) have a very high degree of agreement. C. histolyticum (strain 20) adjoins at a very low level, while C. butyricum (strain 13) shows a unfixed behaviour. The dendrograms display a disposition according with Classical Taxonomy.

Eighteen lots of processed cheese and processed cheese spread were analyzed. Results obtained for the yeasts and molds count (RHL) and the total coliforms count (RCT) were 88.9% inferior to 100 per gram, in the 90 subsamples studied. The fecal coliforms count (RCF) and the S. aureus count were, respectively, 96.6% and 94.4% inferior to 10 per gram, for the same subsamples. Taking into account the format presented by the International Commission on Microbiological Specifications for Foods and the results of our survey, a three class plan is proposed for the following microbiological tests: RHL (n = 5, c = 2, m = 100, M = 1000); RCT (n = 5, c = 2, m = 100, M = 1000); RCF (n = 5, c = 2, m = 10, M = 100); and S. aureus count (n = 5, c = 1, m = 100, M = 1000). Therefore, applying these microbiological specifications, 4 lots (22.2%) would be rejected.

The production of heat-labile enterotoxin (LT) by E. coli strains was studied. Four strains isolated from acute infant diarrhea cases in Argentina and one reference strain, all of them producers of different levels of LT, were employed. GM1-ELISA was used as titration method. The production was assayed in two culture media; Evans (Ev) and Trypticase Soy Broth (CTS) with or without glucose and with different concentrations of lincomycin (0; 30; 45 y 90/micrograms/ml). The effects of pH and inoculum concentration were also examined. Higher levels of LT were found to be produced in Ev (mean production value X = 233.1 ng/ml) than in CTS (X = 133.8 ng/ml), although the difference was statistically not significant (P = 0.22). Lincomycin addition stimulated toxin synthesis and release in both media, and these effects were found to increase with the concentration of the antibiotic (P less than 0.01). Glucose increased the LT levels of low-producing strains and also the growth rate, cellular yield and LT levels of 40T and CC2e strains. Levels of LT released were found to be independent of inoculum concentration. In CTS without pH control, the LT production was one half the levels attained with regulated pH. This was not an effect on the release of the toxin as the determination included both the extracellular LT and the intracellular toxin accumulated in the periplasmic space, which is released by polymyxin B treatment. A pH-dependent inactivation of the synthetized and release toxin was demonstrated.

A total of 23 lots of dehydrated and instant products for babies and children were analyzed. Neither Salmonella, C. perfringens, B. cereus, St. aureus, Group A beta-hemolytic streptococci, fecal coliforms, E. coli nor aflatoxins could be detected in the 115 samples studied. With the exception of the cereals, results obtained for the Aerobic Plate Count (RAP), the Moulds and Yeast Count (RHL) and the Total Coliforms Count (RCT) were, respectively, 91% equal of lower then 3 per gram, and 92% lower than 100 per gram. The results obtained justify the proposition of consultants of FAO/WHO, to limit determination to RAP, RCT, and investigation of Salmonella. According to specifications of FAO/WHO, 10,5% of the lots examined would be considered non acceptable. With regard to the cereals, the lower sensitivity of the used techniques, due to the physical sample characteristics, makes it advisable to adopt RHL here, too.

A dichotomous key is proposed for the identification of eight species of Clostridium: Clostridium botulinum, C. butyricum, C. haemolyticum, C. histolyticum, C. paraperfringens, C. perfringens, C. sporogenes y C. subterminale, on the basis of the Gram staining, catalase production, growth on nutrient agar, glucose utilization, motility test, gelatin hydrolysis, lecithinase production, human blood hemolysis and the test of mice toxicity.

A two episodes case of CSF ventriculo-atrial shunt infection due to B. circulans and B. larvae is presented. B. circulans was first isolated from 4 blood cultures and CSF (shunt valve tap). The patient showed a brain damage syndrome reversible with antibiotic treatment. Lethal toxin production was demonstrated for the B. circulans strain in a mouse model. This strain was found to be a variant of Gordon's description as it produced urease and was tolerant to 7% NaCl. The patient recovered after cefotaxime, cotrimoxazole and rifampicin treatment. A second infection due to B. larvae was detected two months later. The shunt system was removed due to obstruction and a scanning electron microscopy study was performed. Confluent masses of white blood cells and rods were observed on the inner surface of the catheter. As for as we know, this is the first case of human infection due to B. larvae.

A total of 118 Staphylococcus aureus strains isolated from routine sampling, samples from food poisoning outbreaks and human clinical specimens were examined for the production of enterotoxins A, B, C, D and E. The toxic properties of strains were compared with other biochemical characteristics and with the sensitivity to antibiotics. Of the total strains examined, 17.8% (21 strains) produced enterotoxins, and of the toxigenic strains, 81% (17 strains) produced just one type of enterotoxin and 19% (4 strains) two types. Enterotoxin A production was found in 52.4% strains, the other enterotoxins detected in decreasing order of frequency were: C; B; AD; D; AB and BD. All the strains examined produced catalases, coagulases, thermonuclease and fermented glucose; 81 and 89.7% for toxigenic and non-toxigenic strains, respectively, fermented mannitol; 47.6 and 54.6% hydrolyzed casein and 47.6 and 52.6% gelatin; 85.7 and 92.8% produced yellow or orange pigment. Mixed acid fermentation was carried out in 100% and in 96.9%; acetoin was produced by 57.1 and 47.4%; one or more hemolysins were released by 85.7 and 92.8% of the toxigenic and non-toxigenic strains, respectively. Sensitivity to antibiotics was widespread among all the strains. No relation was found between enterotoxin B production and methicillin and tetracycline resistance. Neither the biochemical properties nor the sensitivity to antibiotics has been shown to correlate reliably with toxin production.

A total of 70 samples of ice-creams, milk, milk cream and water or fruit based were obtained from 7 different industrial producers. Of these samples, 14.3% were positive for Staphylococcus aureus in 1 g: 97.2% showed less than 10 St. aureus in 1 g, 1.4% between 101-1,000 and 1.4% between 1,001-10,000. In 21 strains enterotoxigenicity, was investigated, as well as the response to different biochemical tests and the susceptibility to antibiotics: 9.5% of the strains were toxigenic (2 strains), 1 produced enterotoxin A and 1 enterotoxin B. There was no difference between toxigenic and nontoxigenic strains when the biochemical tests were performed (catalase test, coagulase test, thermostable nuclease production, anaerobic utilization of glucose and mannitol, gelatin and casein hydrolysis, production of hemolysins and pigments). All the strains showed a wide susceptibility when they were tested with different antibiotics. Our results confirm the limit introduced to the article 1078 of the Argentine Food Code for ice-creams of industrial production (100 per g) but there is no specification of the count method to be used.

Sixty seven samples of ice-creams were taken from seven different semi-industrial and small-scale shops. 43.3% of the samples were positive for the investigation of Staphylococcus aureus in 1 g; 88% showed less than 10 St. aureus in 1 g; 10.5% between 10 and 100 and 1.5% between 101 and 1,000. In 29 strains was investigated enterotoxigenicity, as well as the response to different biochemical tests and the susceptibility to antibiotics: 21% of the strains were toxigenic (4 produced enterotoxin A, 1 enterotoxin B and 1 enterotoxins A and D). There was no difference between the toxigenic and non-toxigenic strains when biochemical tests were performed (catalase test, coagulase test, thermostable nuclease production, anaerobic utilization of glucose and mannitol, gelatin and casein hydrolysis, production of hemolysins and pigments). All the strains showed susceptibility to different antibiotics. On the basis of the present study it is proposed to incorporate in article 1075 of the Argentine Food Code the limit of 100 Staphylococcus aureus in 1 g of ice creams.