Circulation of enterotoxigenic Escherichia coli (ETEC) isolates expressing CS23 from the environment to clinical settings.

IMPORTANCE: The importance of clean water cannot be overstated. It is a vital resource for maintaining health and well-being. Unfortunately, water sources contaminated with fecal discharges from animal and human origin due to a lack of wastewater management pose a significant risk to communities, as they can become a means of transmission of pathogenic bacteria like enterotoxigenic E. coli (ETEC). ETEC is frequently found in polluted water in countries with a high prevalence of diarrheal diseases, such as Bolivia. This study provides novel insights into the circulation of ETEC between diarrheal cases and polluted water sources in areas with high rates of diarrheal disease. These findings highlight the Choqueyapu River as a potential reservoir for emerging pathogens carrying antibiotic-resistance genes, making it a crucial area for monitoring and intervention. Furthermore, the results demonstrate the feasibility of a low-cost, high-throughput method for tracking bacterial pathogens in low- and middle-income countries, making it a valuable tool for One Health monitoring efforts.

Escherichia coli ST2797 Is Abundant in Wastewater and Might Be a Novel Emerging Extended-Spectrum Beta-Lactamase E. coli.

The increasing prevalence of antibiotic-resistant bacteria is an emerging threat to global health. The analysis of antibiotic-resistant enterobacteria in wastewater can indicate the prevalence and spread of certain clonal groups of multiresistant bacteria. In a previous study of Escherichia coli that were isolated from a pump station in Norway over 15 months, we found a recurring E. coli clone that was resistant to trimethoprim, ampicillin, and tetracycline in 201 of 3,123 analyzed isolates (6.1%). 11 representative isolates were subjected to whole-genome sequencing and were found to belong to the MLST ST2797 E. coli clone with plasmids carrying resistance genes, including blaTEM-1B, sul2, dfrA7, and tetB. A phenotypic comparison of the ST2797 isolates with the uropathogenic ST131 and ST648 that were repeatedly identified in the same wastewater samples revealed that the ST2797 isolates exhibited a comparable capacity for temporal survival in wastewater, greater biofilm formation, and similar potential for the colonization of mammalian epithelial cells. ST2797 has been isolated from humans and has been found to carry extended spectrum ß-lactamase (ESBL) genes in other studies, suggesting that this clonal type is an emerging ESBL E. coli. Collectively, these findings show that ST2797 was more ubiquitous in the studied wastewater than were the infamous ST131 and ST648 and that ST2797 may have similar abilities to survive in the environment and cause infections in humans. IMPORTANCE The incidence of drug-resistant bacteria found in the environment is increasing together with the levels of antibiotic-resistant bacteria that cause infections. The COVID-19 pandemic has shed new light on the importance of monitoring emerging threats and finding early warning systems. Therefore, to mitigate the antimicrobial resistance burden, the monitoring and early identification of antibiotic-resistant bacteria in hot spots, such as wastewater treatment plants, are required to combat the occurrence and spread of antibiotic-resistant bacteria. Here, we applied a PhenePlate system as a phenotypic screening method for genomic surveillance and discovered a dominant and persistent E. coli clone ST2797 with a multidrug resistance pattern and equivalent phenotypic characteristics to those of the major pandemic lineages, namely, ST131 and ST648, which frequently carry ESBL genes. This study highlights the continuous surveillance and report of multidrug resistant bacteria with the potential to spread in One Health settings.

Repeated Isolation of Extended-Spectrum-ß-Lactamase-Positive Escherichia coli Sequence Types 648 and 131 from Community Wastewater Indicates that Sewage Systems Are Important Sources of Emerging Clones of Antibiotic-Resistant Bacteria.

Antibiotic resistance in bacteria is an emerging problem globally. Resistant bacteria are found in human and animal microbiota, as well as in the environment. Wastewater receives bacteria from all these sources and thus can provide a measurement of abundance and diversity of antibiotic-resistant bacteria circulating in communities. In this study, water samples were collected from a wastewater pump station in a Norwegian suburban community over a period of 15 months. A total of 45 daily samples were cultured and analyzed for the presence of Escherichia coli Eighty E. coli-like colonies were collected from each daily sample and then phenotyped and analyzed for antibiotic resistance using the PhenePlate-AREB system. During the sampling period, two unique E. coli phenotypes with resistance to cefotaxime and cefpodoxime indicating carriage of extended-spectrum ß-lactamases (ESBL) were observed repeatedly. Whole-genome sequencing of 15 representative isolates from the two phenotypes identified these as two distinct clones belonging to the two globally spread E. coli multilocus sequence types (STs) ST131 and ST648 and carrying blaCTX-M-15 The number of ESBL-positive E. coli strains in the community wastewater pump station was 314 of 3,123 (10%) analyzed E. coli strains. Of the ESBL-positive isolates, 37% belonged to ST648, and 7% belonged to ST131. Repeated findings of CTX-M-15-positive ST648 and ST131 over time indicate that these STs are resident in the analyzed wastewater systems and/or circulate abundantly in the community.

Diversity and antibiotic resistance among Escherichia coli populations in hospital and community wastewater compared to wastewater at the receiving urban treatment plant.

Bacterial diversity and antimicrobial resistance patterns among the indicator organism Escherichia coli were monitored in wastewater samples collected over one year from a hospital (HW), a community (CW) and the receiving urban (UW) wastewater treatment plant (WWTP). We compared levels of antibiotic resistance in the different types of wastewater, and identified whether resistant strains were endemic in the wastewater system. If so, implementation of local treatment at certain resistance hotspots (e.g. hospital outlets) could be used to decrease the amount of resistant bacteria in the wastewater. E. coli from HW (n = 2644), CW (n = 2525) and UW (n = 2693) were analyzed by biochemical phenotyping (PhenePlate System) and antimicrobial susceptibility testing to nine antibiotics (AREB System). The phenotypic diversities of the total E. coli populations were similar for all three sites (Simpson's Diversity index, Di = 0.973), however for individual samples, HW showed low diversities (Median Di = 0.800) and the E. coli flora was often dominated by strains that may have originated from the fecal flora of single individuals. The diversities in CW samples was higher (Median Di = 0.936), and UW samples showed similar diversities as the whole collection of isolates (Median Di = 0.971). Resistance to at least one of the nine antibiotics was observed in 45% of the HW isolates, 44% of CW isolates, and 33% of UW isolates. Resistance to gentamicin and chloramphenicol was uncommon (3.2 and 5.3%, respectively), whereas resistance to tetracycline and ampicillin was most common (24% and 31%, respectively). Extended-spectrum beta-lactamase-producing E. coli (ESBL-EC) were more common in HW (11.5%) and in CW (6.9%) compared to UW (3.7%). A high diversity (Di = 0.974) was observed among ESBL-EC isolates from UW (n = 99), indicating absence of any clonal structure among these isolates. Common PhP types of ESBL-EC often dominated in each HW sample, but were not identified across different samples, whereas ESBL-EC in CW showed low diversity (Di = 0.857) and were dominated by a specific PhP type that was found across almost all CW samples. The antibiotic resistance rates were highest in hospital wastewater, but surprisingly they were also high in the studied community wastewater, compared to the urban wastewater. The relative contribution of HW seemed low in terms of dissemination of antibiotic resistant bacteria to the WWTP.

Persistence of Lactobacilli in Postmenopausal Women - A Double-Blind, Randomized, Pilot Study.

AIM: To investigate the ability of lactobacilli to persist in the genital area (vagina and labia) of women after the topical application of an ointment containing Lactobacillus gasseri LN40, L. fermentum LN99 and L. rhamnosus LN113. Secondary objectives were to study the presence of Escherichia coli and other contaminants, as well as subjective symptoms in the genital tract. METHODS: Eighteen healthy postmenopausal women were randomized to use either the study product or placebo for 10 days. Gynecological examinations, labial and vaginal samplings for bacterial cultivation were performed at baseline (visit 1), after treatment (visit 2), and at a 10-day follow-up (visit 3). LN strains were identified by specific cultivation methods. Subjective symptoms were evaluated by a self-administered questionnaire. RESULTS: The presence of LN99 was shown in 7 out of 8 women in the investigational group at visit 2 (p < 0.001 compared to placebo) and in 5 out of 8 at visit 3 (p < 0.05), whereas the presence of LN113 was shown in 2 out of 8 at visit 2 and in 1 out of 8 at visit 3. Subjective symptoms were significantly reduced (p < 0.01) at visits 2 and 3 for both products. CONCLUSION: Topical application of a probiotic ointment is feasible to achieve persistence of lactobacilli for at least 10 days.

Surveillance of antimicrobial resistance among Escherichia coli in wastewater in Stockholm during 1 year: does it reflect the resistance trends in the society?

The resistance patterns of Escherichia coli in untreated (raw) urban wastewater (UW) was monitored by repeated sampling during 1 year. Comparison with data from wastewater samples collected from hospital wastewater (HW) in the same urban area was made. A total of 1326 E. coli isolates from 17 UW samples and 451 isolates from six HW samples were analysed by typing using the PhenePlate™ system, and their susceptibility towards 10 antibiotics was determined. Resistance to at least one antibiotic was observed in 34% of the UW isolates and 55% of the HW isolates. For UW isolates, phenotypic diversity was lower among antibiotic-susceptible than among antibiotic-resistant isolates, indicating a higher presence of clonal groups among susceptible isolates. Total antibiotic resistance measured as the Multiple Antibiotic Resistance (MAR) index was 0.08 for UW compared with 0.19 for HW, and increased over time for UW isolates, indicating increasing resistance among E. coli in the urban population during the studied time period. Resistance to all included ß-lactam antibiotics was detected in 2.4% of UW isolates and 14.0% of HW isolates, and 73/75 (97%) analysed isolates were confirmed to be extended-spectrum ß-lactamase (including plasmid-mediated AmpC)-producing E. coli. Thus, by cultivating samples from wastewater and analysing many independent isolates per sample, increasing frequencies of antibiotic resistance in UW were detected during 1 year that may reflect increasing faecal carriage of resistant bacteria in the society. Surveillance of antibiotic resistance in wastewater could be a valuable tool for screening of resistance trends on a population level.

Antibiotic resistance in environmental Escherichia coli - a simple screening method for simultaneous typing and resistance determination.

We describe a simple and standardised screening system (AREB) for surveillance of antibiotic resistant bacteria in the environment. The system consists of 96 well microplates containing eight sets of breakpoint amounts of 10 different antibiotics. The incubated microplates are read by a desktop scanner and the plate images are analysed by special software that automatically presents the resistance data. The AREB method is combined with a rapid typing method, the PhenePlate system, which yields information on the diversity of the bacteria in the studied samples, and on the possible prevalence of resistant clones. In order to demonstrate the usage of AREB, a comparative study on the resistance situation among 970 Escherichia coli isolates from sewage and recipient water in Sweden, Norway and Chile, was performed. Resistance rates to all antibiotics were markedly higher in hospital sewage than in other samples. Our data indicate that the AREB system is useful for comparing resistance rates among E. coli and other environmental indicator bacteria in different countries/regions. Simple handling and automatic data evaluation, combined with low cost, facilitate large studies involving several thousands of isolates.

Diarrheagenic Escherichia coli markers and phenotypes among fecal E. coli isolates collected from Nicaraguan infants.

We analyzed the prevalence of diarrheagenic Escherichia coli (DEC) markers and common phenotypes in 2,164 E. coli isolates from 282 DEC-positive samples. Enteropathogenic E. coli (EPEC) and enteroaggregative E. coli (EAEC) were very diverse and were not correlated with diarrhea. Enterotoxigenic E. coli (ETEC) estA and enterohemorrhagic E. coli (EHEC) belonged to a few phenotypes and were significantly correlated with diarrhea.

Ovine clone ST1464: a predominant genotype of Staphylococcus aureus subsp. anaerobius isolated from sheep in Sudan.

BACKGROUND: The aim of the present study was to examine the phenotypic and genotypic relatedness of 17 Staphylococcus aureus subsp. anaerobius isolates recovered from sheep abscesses in Khartoum state, Sudan, during the period 2007-2008. METHODOLOGY: This sample was characterised using antibiogram typing, biochemical typing with the commercial PhenePlate system (PhP-CS) and multilocus sequence typing (MLST). RESULTS: Low levels of resistance were noted to the 11 antimicrobial agents tested. All the isolates corresponded to a single PhP type, and to a single, novel, multilocus sequence type, designated ST1464. CONCLUSION: These results demonstrate that the vast majority of cases of sheep abscess disease in Khartoum state are caused by a single novel clone of S. aureus subsp. anaerobius.

Caracterización fenotípica y genotípica de la resistencia a meticilina de cepas de Staphylococcus aureus aisladas en Antofagasta / Phenotypic and genotypic characterization of methicillin resistance in strains of Staphylococcus aureus isolated in Antofagasta

Staphylococcus aureus resistant to methicillin (SARM) has been associated with nosocomial infections due to its capacity to develop resistance to multiple antibiotics. There is little information about the SARM which are found in the hospital services of Antofagasta. We studied the phenotypic and genotypic characteristics of methicillin resistance in 38 strains of S. aureus isolated in Antofagasta, identified by coagulase and API Staph tests and by a biochemical test (Ph-system). The susceptibility to antibiotics was studied using the agar dilution technique, identifying SARM strains with discs of oxacillin. Beta-lactamase with nitrocephine, and the gene mecA by means of PCR. Eighty nine percent (34 strains) were SARM with a high resistance to ampicillin, penicillin, erythromycin, claritromycin. gentiamycin, amikacine and ciprofloxacine. All isolates were susceptible to vancomycin and rifampicin. Beta-lactamase was demonstrated in 79% of the SARM strains. Strain typing and resistance patterns revealed a great diversity of PhP-types and antibiotypes in the isolates. Ninety seven percent of the SARM strains had the gene mecA. One PhP-type (C6) was dominant (5 SARM strains) all had the mecA gene, produced beta lactamase and had the same pattern of antibiotic resistance. We conclude that the dominant phenotypes of SARM strains which have the mecA gene and multiple resistance to antibiotics are present in the hospitals of Antofagasta, and sound the alert on the risk of nosocomial transmission of epidemic clones of SARM.

Staphylococcus aureus resistentes a meticilina (SARM) han sido asociados con infecciones nosocomiales por su capacidad para desarrollar resistencia a múltiple antibióticos, existiendo escasa información acerca de SARM que están circulando en los servicios hospitalarios de Antofagasta. Se estudió características fenotIpicas y genotípicas de la resistencia a meticilina en 38 cepas de S. aureus aisladas en Antofagasta, identificadas por tests de coagulasa y API Staph y por tipificación bioquímica (Ph-Sistem). La susceptibilidad a antibióticos se realizo por técnica de dilución en agar, las cepas SARM fueron identificadas con discos de oxacilina, beta-lactamasa por nitrocefina y gen mecA fue detectado pot PCR. El 89% (34 cepas), fueron SARM con una alta resistencia a ampicilina, penicilina, eritromicina, gentamicina, amikacina y ciprofloxacino. Todos los aislados fueron susceptibles a vancomocina y rifampicina. Beta lactamasa fue demostrada en 79% de las cepas SARM. La tipificación y los patrones de resistencia revelaron una alta diversidad de PhP tipos y antibioticos en los aislamientos. El 97% de las cepas SARM albergaban el gen mecA. Un PhP tipo (C6) fue dominante. (5 cepas SARM), todos presentando el gen mecA, produciendo beta lactamasa y mostrando el mismo patrón de resistencia antibiótica. Se concluye que los fenotipos dominantes de cepas SARM que albergan el gen mecA y resistencia múltiples alos antibióticos están circulando en los hospitales de Antofagasta, alertando sobre el riesgo de transmisión intranosocomial de clones epidémicos de SARM.

Diversity of intestinal Escherichia coli populations in Nicaraguan children with and without diarrhoea.

Escherichia coli remains an important aetiological agent of infantile diarrhoea in Nicaragua. However, little is known about whether there is a high prevalence of endemic strains or whether infection is due to the epidemic spread of virulent clones. This study was undertaken to determine the diversity and distribution of clonal groups in a population of intestinal E. coli isolated from the faeces of children from León, Nicaragua, with (n=381) and without (n=145) diarrhoea, between March 2005 and September 2006. All samples had been screened previously for the presence of diarrhoeagenic E. coli (DEC) markers by multiplex PCR. From each sample, 8 E. coli colonies (where available) were analysed by biochemical fingerprinting (PhP-RE system), yielding a total of 4009 tested isolates. On average, three different biochemical phenotypes (BPTs) were found among the eight colonies analysed from each sample. The total diversity, measured as Simpson's diversity index (Di), was 0.97 among all 4009 isolates studied. Cluster analysis of data from all 4009 isolates revealed 24 common BPTs (identified in at least 1 % of the isolates) and 234 less common BPTs. Similar Di values were obtained among isolates from infants with and without diarrhoea, indicating that no widespread outbreak of DEC had occurred. Moreover, among samples that were positive for the DEC types enteroaggregative E. coli, enteropathogenic E. coli and enterotoxigenic E. coli (ETEC) carrying the eltB gene, the diversities were almost as high as among non-DEC samples, whereas samples positive for ETEC carrying estA, enteroinvasive E. coli and enterohaemorrhagic E. coli showed lower diversities, indicating the prevalence of virulent clonal groups among these samples. The PhenePlate patterns of the 24 common BPTs identified here were compared with those obtained from E. coli isolated in a cohort infant study performed in 1991-1992 in the same area. Only 4 % of the isolates from the 1990s were similar to any of the common BPTs found in the present study.

Prevalence and transmission of antimicrobial resistance among Aeromonas populations from a duckweed aquaculture based hospital sewage water recycling system in Bangladesh.

In order to investigate the influence of a duckweed aquaculture based hospital sewage water recycling plant on the prevalence and dissemination of antibiotic resistance, we made use of an existing collection of 1,315 Aeromonas isolates that were previously typed by the biochemical fingerprinting PhP-AE system. In these treatment plant, hospital raw sewage water is first collected in a settlement pond (referred to as sewage water in this study) and is then transferred to a lagoon, where the duckweed (Lemnaceae) is grown (referred to as lagoon). The duckweed is harvested and used as feed for the fish in a separate pond (referred to as fish pond). From this collection, representatives of 288 PhP types were subjected to antibiotic susceptibility testing for eight antimicrobials by broth microdilution method. The overall resistance rates among Aeromonas isolates from the treatment plant were highest for ampicillin (87%) and erythromycin (79%) followed by cephalothin (58%), nalidixic acid (52%), streptomycin (51%), tetracycline (31%), chloramphenicol (13%) and gentamicin (8%). A significantly lower prevalence of antibiotic resistance was found in Aeromonas from environmental control water, patient stool samples, duckweed and fish compared to sewage water isolates. The prevalence of resistance in the sewage water was not significantly reduced compared to the lagoon water and fish pond. Throughout the treatment system, the frequencies of resistant strains were found to diminish during the sewage water purification process, i.e. in the lagoon where sewage water is used to grow the duckweed. However, the frequency of resistant strains again increased in the fish pond where sewage grown duckweed is used for aquaculture. Among the selected isolates, two multiresistant clonal groups of Aeromonas caviae HG4 were identified that exhibited indistinguishable PhP and amplified fragment length polymorphism fingerprints and shared a common plasmid of approximately 5 kb. Representatives of both groups were recovered from almost every part of the sewage treatment plant but not in the control ponds nor in human samples, which suggests that specific multiresistant Aeromonas clones are able to persist and spread throughout the entire purification process.

Asymptomatic bacteriuria in the elderly: high prevalence and high turnover of strains.

Asymptomatic bacteriuria (ASB) was followed in repeated prevalence surveys in a cohort of non-institutionalized residents (n=330), aged>or=80 y. Urine samples were collected at baseline, and at 6, and at 18 months. Phenotyping (PhenePlate) was performed on isolates of Escherichia coli to evaluate strain relatedness. ASB occurred in 19.0, 19.4, and 19.9% in women, and in 9.4, 9.6 and 7.9% in men, at baseline and at the 6- and 18-months follow-up, respectively, and ASB was found at least once in 37% of women and in 20% of men. Of those with ASB at baseline, 60% also had ASB in the 2 subsequent surveys. Among those with persisting E. coli bacteriuria, 76% and 40%, respectively, carried the same strain at the 6- and 18-months follow-ups. In women, we found that the risk of developing a symptomatic urinary tract infection within 24 months was higher among those with ASB at baseline than in those without bacteriuria (p=0.019). ASB is common and often persistent, but we found a high turnover of strains, indicating a high rate of recolonization.

Clonal heterogeneity of clinical isolates of vancomycin-resistant Enterococcus faecium with unique vanS.

OBJECTIVE: To examine the clonal diversity of vancomycin-resistant enterococci (VRE). METHODS: A total of 900 clinical isolates of enterococci were obtained, and VRE isolates were subjected to antimicrobial susceptibility tests, biochemical fingerprinting with the PhPlate system (PhP), ribotyping and pulsed-field gel electrophoresis (PFGE) typing. RESULTS: Forty-nine of all enterococcal isolates were resistant to high levels of vancomycin (MIC >or= 128) and identified as Enterococcus faecium. Biochemical fingerprinting with PhP showed that the VRE isolates were highly diverse (diversity index, D(i) = 0.93) and belonged to 24 PhP-types. The VRE could be separated into 34 and 27 types with PFGE and ribotyping, giving diversity indices of 0.98 and 0.97, respectively. The PFGE method was more discriminatory than ribotyping and PhP system for E. faecium isolates. A combination of either of the two typing methods resulted in at least 44 types. Furthermore, sequencing analysis of vanS of Tn1546 showed one nucleotide mutation (C-->A) at position 5727 in comparison with the prototype BM4147, which was found to be unique in all Iranian VRE isolates. CONCLUSION: The isolated clinical VRE strains were highly diverse in Tehran.

Persistence, transmission, and virulence characteristics of Aeromonas strains in a duckweed aquaculture-based hospital sewage water recycling plant in Bangladesh.

The persistence and transmission of Aeromonas in a duckweed aquaculture-based hospital sewage water treatment plant in Bangladesh was studied. A total of 670 samples from different sites of the hospital sewage water treatment plant, from feces of hospitalized children suffering from diarrhea, from environmental control ponds, and from feces of healthy humans were collected over a period of three years. In total, 1,315 presumptive Aeromonas isolates were biochemically typed by the PhenePlate rapid screening system (PhP-AE). A selection of 90 representative isolates was further analyzed with PhenePlate (PhP) extended typing (PhP-48), fatty acid methyl ester analysis, and amplified fragment length polymorphism (AFLP) fingerprinting. In addition, the prevalence of the putative virulence factors hemolysin and cytotoxin and the presence of the cytolytic enterotoxin gene (AHCYTOEN) were analyzed. Aeromonas was found at all sites of the treatment plant, in 40% of the samples from environmental control ponds, in 8.5% of the samples from hospitalized children suffering from diarrhea, and in 3.5% of samples from healthy humans. A significantly high number of Aeromonas bacteria was found in duckweed, which indicates that duckweed may serve as a reservoir for these bacteria. PhP-AE typing allowed identification of more than 192 distinct PhP types, of which 18 major PhP types (MTs) were found in multiple sites and during several occasions. AFLP fingerprinting revealed the prevalence of genotypically indistinguishable Aeromonas isolates among certain PhP MTs recovered from different sampling occasions and/or at multiple sites. Hemolytic and cytotoxic activities were observed in 43% of the tested strains, whereas 29% possessed the cytolytic enterotoxin gene AHCYTOEN. Collectively, two specific MTs associated with diarrhea were shown to exhibit high cytotoxicity. Furthermore, all tested isolates of these major types were positive for the cytolytic enterotoxin gene. In conclusion, our data indicate that certain phenotypically and genotypically stable clonal lineages of Aeromonas have persisted in the treatment system for a prolonged period and might spread from the hospitalized children suffering from diarrhea to fish produced for human consumption through the sewage water treatment system.

Integrated analysis of established and novel microbial and chemical methods for microbial source tracking.

Several microbes and chemicals have been considered as potential tracers to identify fecal sources in the environment. However, to date, no one approach has been shown to accurately identify the origins of fecal pollution in aquatic environments. In this multilaboratory study, different microbial and chemical indicators were analyzed in order to distinguish human fecal sources from nonhuman fecal sources using wastewaters and slurries from diverse geographical areas within Europe. Twenty-six parameters, which were later combined to form derived variables for statistical analyses, were obtained by performing methods that were achievable in all the participant laboratories: enumeration of fecal coliform bacteria, enterococci, clostridia, somatic coliphages, F-specific RNA phages, bacteriophages infecting Bacteroides fragilis RYC2056 and Bacteroides thetaiotaomicron GA17, and total and sorbitol-fermenting bifidobacteria; genotyping of F-specific RNA phages; biochemical phenotyping of fecal coliform bacteria and enterococci using miniaturized tests; specific detection of Bifidobacterium adolescentis and Bifidobacterium dentium; and measurement of four fecal sterols. A number of potentially useful source indicators were detected (bacteriophages infecting B. thetaiotaomicron, certain genotypes of F-specific bacteriophages, sorbitol-fermenting bifidobacteria, 24-ethylcoprostanol, and epycoprostanol), although no one source identifier alone provided 100% correct classification of the fecal source. Subsequently, 38 variables (both single and derived) were defined from the measured microbial and chemical parameters in order to find the best subset of variables to develop predictive models using the lowest possible number of measured parameters. To this end, several statistical or machine learning methods were evaluated and provided two successful predictive models based on just two variables, giving 100% correct classification: the ratio of the densities of somatic coliphages and phages infecting Bacteroides thetaiotaomicron to the density of somatic coliphages and the ratio of the densities of fecal coliform bacteria and phages infecting Bacteroides thetaiotaomicron to the density of fecal coliform bacteria. Other models with high rates of correct classification were developed, but in these cases, higher numbers of variables were required.

Pheno- and genotyping of Staphylococcus epidermidis isolated from bovine milk and human skin.

The purpose of this study was to improve our knowledge concerning the epidemiology and strain diversity of Staphylococcus epidermidis isolated from bovine milk in commercial dairy herds. A total of 341 S. epidermidis isolates obtained from cows' milk (317), farmers (17) and patients (7) were characterized. Of these 105 isolates were from cows' milk in two farms, where also 17 isolates were sampled from farmers. The remaining 212 isolates from cows' milk were from 170 farms. All isolates were examined by antimicrobial susceptibility, whereas 202 were examined by pulsed-field gel electrophoresis (PFGE) and 122 by ribotyping. PFGE showed single patterns in the human strains with one exception; one strain was categorised as the same clone as four of the milk strains. PFGE divided 73 of the milk strains into 62 different patterns. The PFGE method had high discriminatory power and shows that many different S. epidermidis types exist in milk samples. Antibiotic resistance patterns matched the SmaI profiles closely in the two herds, but poorly in the routinely collected milk samples. Isolates from herd 1 showed one to five patterns, depending on the typing method used. Isolates from the milker's skin showed one pattern, which was identical to the most common pattern found in the milk isolates. Isolates from herd 2 showed three to four patterns, two of these being identical to skin isolates from the milker. As dairy cows are not a natural host for S. epidermidis the results suggest a human source of these udder infections.

[Prevalence of antibiotic resistant Enterococcus spp in waste waters in the north of Chile]. / Prevalencia de Enterococos resistentes a antibióticos en aguas servidas en el norte de Chile.

BACKGROUND: There is little information available in Chile on the distribution of Enterococcus spp in waste water and its implications in transmission of antibiotic resistance through the water cycle. Enterococcus spp are common in nosocomial infections and may spread antibiotic resistance through the food chain. AIM: To determine the presence of antibiotic resistant Enterococcus spp in the sewage of Antofagasta, Chile. MATERIAL AND METHODS: Samples of sewage from two sewage treatment plants and from the Public Hospital of Antofagasta collector were obtained. Enterococcus spp were isolated on m-Enterococcus agar containing ampicillin, vancomycin and streptomycin. The isolates were identified and subjected to biochemical typing (PhPlate). Minimal inhibitory concentration determination was performed by agar dilution technique. RESULTS: High counts of resistant Enterococcus spp were found on the streptomycin plates, lower on ampicillin and very low on vancomycin plates. A total of 63 Enterococcus spp strains were typed and the identification showed 5 different species; E faecalis (65%), E faecium (14%), E hirae (13%), E durans (6%) and E gallinarum (2%). The typing revealed a high diversity among the isolates. Two biochemical phenotypes were predominant, C1 (21 strains) and C6 (7 strains). Both were highly resistant to gentamycin and streptomycin; moderately resistant to ampicillin, chloramphenicol, tetracycline and ciprofloxacin, and with intermediate susceptibility to vancomycin. Both phenotypes were found in the sewage of the hospital collector and in the treatment plants. CONCLUSIONS: In the sewage of Antofagasta we found dominating phenotypes of multiresistant Enterococcus spp. Sewage could be an important way of transmission of these microorganisms.

Prevalencia de Enterococos resistentes a antibióticos en aguas servidas en el norte de Chile / Prevalence of antibiotic resistant Enterococcus spp in waste waters in the north of Chile

Background: There is little information available in Chile on the distribution of Enterococcus spp in waste water and its implications in transmission of antibiotic resistance through the water cycle. Enterococcus spp are common in nosocomial infections and may spread antibiotic resistance through the food chain. Aim: To determine the presence of antibiotic resistant Enterococcus spp in the sewage of Antofagasta, Chile. Material and Methods: Samples of sewage from two sewage treatment plants and from the Public Hospital of Antofagasta collector were obtained. Enterococcus spp were isolated on m-Enterococcus agar containing ampicillin, vancomycin and streptomycin. The isolates were identified and subjected to biochemical typing (PhPlate). Minimal inhibitory concentration determination was performed by agar dilution technique. Results: High counts of resistant Enterococcus spp were found on the streptomycin plates, lower on ampicillin and very low on vancomycin plates. A total of 63 Enterococcus spp strains were typed and the identification showed 5 different species; E faecalis (65%), E faecium (14%), E hirae (13%), E durans (6%) and E gallinarum (2%). The typing revealed a high diversity among the isolates. Two biochemical phenotypes were predominant, C1 (21 strains) and C6 (7 strains). Both were highly resistant to gentamycin and streptomycin; moderately resistant to ampicillin, cloramphenicol, tetracycline and ciprofloxacin, and with intermediate susceptibility to vancomycin. Both phenotypes were found in the sewage of the hospital collector and in the treatment plants. Conclusions: In the sewage of Antofagasta we found dominating phenotypes of multiresistant Enterococcus spp. Sewage could be an important way of transmission of these microorganisms.

Occurrence and relatedness of vancomycin-resistant enterococci in animals, humans, and the environment in different European regions.

Vancomycin-resistant enterococci (VRE) in Europe are thought to have emerged partly due to the use of the glycopeptide avoparcin in animal husbandry. We compared the occurrence of VRE in geographical regions of Europe in which until 1997 large amounts of avoparcin were used (Spain, United Kingdom, and Denmark) with the occurrence of VRE in Sweden, where avoparcin was banned in 1986. We also studied the relatedness between VRE strains from different regions and habitats. In total, 2,580 samples were collected from humans, animals, and the environment (soil, sewage, recipient water). VRE resistant to 20 microg/ml vancomycin were identified in 8.2% of the samples and were found most frequently in raw and treated urban sewage samples (means, 71% and 36% of the samples, respectively), pig manure (17%), and hospital sewage (16%). The proportions of VRE-positive sewage samples were similar in Sweden, Spain, and the United Kingdom, whereas pig feces and manure were more often positive in Spain than in Sweden (30% versus 1%). Most VRE were Enterococcus faecium carrying vanA, and computerized biochemical phenotyping of the isolates of different ecological origins showed a high degree of polyclonality. In conclusion, it seems that animal-associated VRE probably reflect the former use of avoparcin in animal production, whereas VRE in human-associated samples may be a result of antibiotic use in hospitals. Since there seems to be a reservoir of the resistance genes in all countries studied, precautions must be taken to limit the use of antibiotics and antibiotic-like feed additives.