A Comparative Study on Antimicrobial Resistance in Escherichia coli Isolated from Channel Catfish and Related Freshwater Fish Species.

Antimicrobial resistance (AMR) trends in 114 generic Escherichia coli isolated from channel catfish and related fish species were investigated in this study. Of these, 45 isolates were from commercial-sized channel catfish harvested from fishponds in Alabama, while 69 isolates were from Siluriformes products, accessed from the U.S. Department of Agriculture Food Safety and Inspection Service' (FSIS) National Antimicrobial Resistance Monitoring System (NARMS) program. Antibiotic susceptibility testing and whole genome sequencing were performed using the GenomeTrakr protocol. Upon analysis, the fishpond isolates showed resistance to ampicillin (44%), meropenem (7%) and azithromycin (4%). The FSIS NARMS isolates showed resistance to tetracycline (31.9%), chloramphenicol (20.3%), sulfisoxazole (17.4%), ampicillin (5.8%) and trimethoprim-sulfamethoxazole, nalidixic acid, amoxicillin-clavulanic acid, azithromycin and cefoxitin below 5% each. There was no correlation between genotypic and phenotypic resistance in the fishpond isolates, however, there was in NARMS isolates for folate pathway antagonists: Sulfisoxazole vs. sul1 and sul2 (p = 0.0042 and p < 0.0001, respectively) and trimethoprim-sulfamethoxazole vs. dfrA16 and sul1 (p = 0.0290 and p = 0.013, respectively). Furthermore, correlations were found for tetracyclines: Tetracycline vs. tet(A) and tet(B) (p < 0.0001 each), macrolides: Azithromycin vs. mph(E) and msr(E) (p = 0.0145 each), phenicols: Chloramphenicol vs. mdtM (p < 0.0001), quinolones: Nalidixic acid vs. gyrA_S83L=POINT (p = 0.0004), and ß-lactams: Ampicillin vs. blaTEM-1 (p < 0.0001). Overall, we recorded differences in antimicrobial susceptibility testing profiles, phenotypic-genotypic concordance, and resistance to critically important antimicrobials, which may be a public health concern.

Microbiota and Dose Response: Evolving Paradigm of Health Triangle.

SRA Dose-Response and Microbial Risk Analysis Specialty Groups jointly sponsored symposia that addressed the intersections between the "microbiome revolution" and dose response. Invited speakers presented on innovations and advances in gut and nasal microbiota (normal microbial communities) in the first decade after the Human Microbiome Project began. The microbiota and their metabolites are now known to influence health and disease directly and indirectly, through modulation of innate and adaptive immune systems and barrier function. Disruption of healthy microbiota is often associated with changes in abundance and diversity of core microbial species (dysbiosis), caused by stressors including antibiotics, chemotherapy, and disease. Nucleic-acid-based metagenomic methods demonstrated that the dysbiotic host microbiota no longer provide normal colonization resistance to pathogens, a critical component of innate immunity of the superorganism. Diverse pathogens, probiotics, and prebiotics were considered in human and animal models (in vivo and in vitro). Discussion included approaches for design of future microbial dose-response studies to account for the presence of the indigenous microbiota that provide normal colonization resistance, and the absence of the protective microbiota in dysbiosis. As NextGen risk analysis methodology advances with the "microbiome revolution," a proposed new framework, the Health Triangle, may replace the old paradigm based on the Disease Triangle (focused on host, pathogen, and environment) and germophobia. Collaborative experimental designs are needed for testing hypotheses about causality in dose-response relationships for pathogens present in our environments that clearly compete in complex ecosystems with thousands of bacterial species dominating the healthy superorganism.

Salmonella on Raw Poultry in Retail Markets in Guatemala: Levels, Antibiotic Susceptibility, and Serovar Distribution.

The objective of this study was to determine Salmonella numbers on retail raw chicken carcasses in Guatemala and to phenotypically characterize the isolates (serotyping and antibiotic susceptibility). In total, 300 chicken carcasses were collected from seven departments in Guatemala. Salmonella numbers were determined using the most-probable-number method following the U. S. Department of Agriculture's Food Safety and Inspection Service protocol. In total, 103 isolates were obtained, all of which were tested for antibiotic susceptibility, whereas 46 isolates were serotyped. Overall, Salmonella prevalence and mean number (mean log most probable number per carcass) was 34.3% and 2.3 (95% confidence interval: 2.1 to 2.5), respectively. Significant differences (P < 0.05) in Salmonella prevalence were found by storage condition (refrigerated or ambient temperature), market type (wet markets, supermarkets, and independent poultry stores), chicken production system (integrated or nonintegrated production company), and chicken skin color (white or yellow). Chickens produced by integrated companies had lower Salmonella numbers (P < 0.05) than nonintegrated companies, and white-skin carcasses had lower numbers (P < 0.05) than yellow-skin carcasses. Among 13 different Salmonella serovars identified, Paratyphi B (34.8%) was most prevalent, followed by Heidelberg (16.3%) and Derby (11.6%). Of all the Salmonella isolates, 59.2% were resistant to one to three antibiotics and 13.6% to four or more antibiotics. Among all the serovars obtained, Salmonella Paratyphi B and Heidelberg were the most resistant to the antibiotics tested. Salmonella levels and antibiotic resistant profiles among isolates from raw poultry at the retail market level were high relative to other reports from North and South America. These data can be used by Guatemalan stakeholders to develop risk assessment models and support further research opportunities to control transmission of Salmonella spp. and antibiotic-resistant isolates from chicken meat to humans.

Counts, serotypes, and antimicrobial resistance of Salmonella isolates on retail raw poultry in the People's Republic of China.

The objective of this study was to determine Salmonella counts, serotypes, and antimicrobial resistance profiles in retail raw chicken meat in the People's Republic of China. Salmonella counts were determined according to the most-probable-number (MPN) method for 300 whole chicken carcasses. These samples were collected from large, small, and wet (open) markets in Guangdong, Shaanxi, and Sichuan provinces. Salmonella isolates were serotyped and tested for antimicrobial susceptibility. Of the 300 chicken carcasses, 43.3% were positive for Salmonella, with an overall mean of 1.7 log MPN per carcass (95% confidence interval, 1.6 to 1.8 log MPN per carcass). No significant differences (P > 0.05) were detected for storage temperature (i.e., chilled, frozen, or ambient), market type (large, small, or wet), province, or location (capital or noncapital city). Seventy-eight serotypes were identified among the 1,094 Salmonella isolates. The top five most common Salmonella serotypes on raw chicken carcasses were Enteritidis (19.2%), Indiana (15.2%), Typhimurium (14.6%), Agona (7.1%), and Thompson (6.6%). Salmonella isolates (n = 779) were most frequently resistant to sulfisoxazole (74.1%) and tetracycline (71.1%) and least resistant to ceftriaxone (22.5%) and cefoxitin (19%). Only 4% of the isolates were susceptible to all 15 antimicrobial agents, 45% were resistant to 1 to 5 agents, 29% were resistant to 6 to 10 agents, and 22% were resistant to 11 to 15 agents. Our findings revealed that Salmonella contamination was common in retail raw poultry in China, and the counts on contaminated carcasses were mostly low. Salmonella isolates were diverse in their serotype distribution and antimicrobial susceptibility profiles, with more than half of the isolates resistant to more than five antimicrobial agents. These data may be used in risk assessment models to reduce the transmission of Salmonella via chicken meat to humans in China.

Counts, serovars, and antimicrobial resistance phenotypes of Salmonella on raw chicken meat at retail in Colombia.

The objective of this study was to determine Salmonella counts, serovars, and antimicrobial-resistant phenotypes on retail raw chicken carcasses in Colombia. A total of 301 chicken carcasses were collected from six departments (one city per department) in Colombia. Samples were analyzed for Salmonella counts using the most-probable-number method as recommended by the U.S. Department of Agriculture, Food Safety Inspection Service protocol. A total of 378 isolates (268 from our previous study) were serotyped and tested for antimicrobial susceptibility. The overall Salmonella count (mean log most probable number per carcass ± 95% confidence interval) and prevalence were 2.1 (2.0 to 2.3) and 37%, respectively. There were significant differences (P < 0.05) by Salmonella levels (i.e., counts and prevalence) by storage temperature (i.e., frozen, chilled, or ambient), retail store type (wet markets, supermarkets, and independent markets), and poultry company (chicken produced by integrated or nonintegrated company). Frozen chicken had the lowest Salmonella levels compared with chicken stored at other temperatures, chickens from wet markets had higher levels than those from other retail store types, and chicken produced by integrated companies had lower levels than nonintegrated companies. Thirty-one Salmonella serovars were identified among 378 isolates, with Salmonella Paratyphi B tartrate-positive (i.e., Salmonella Paratyphi B dT+) the most prevalent (44.7%), followed by Heidelberg (19%), Enteritidis (17.7%), Typhimurium (5.3%), and Anatum (2.1%). Of all the Salmonella isolates, 35.2% were resistant to 1 to 5 antimicrobial agents, 24.6% to 6 to 10, and 33.9% to 11 to 15. Among all the serovars obtained, Salmonella Paratyphi B dT+ and Salmonella Heidelberg were the most antimicrobial resistant. Salmonella prevalence was determined to be high, whereas cell numbers were relatively low. These data can be used in developing risk assessment models for preventing the transmission of Salmonella from chicken to humans in Colombia.

Quantification, serovars, and antibiotic resistance of salmonella isolated from retail raw chicken meat in Vietnam.

The objectives of this study were to quantify Salmonella counts on retail raw poultry meat in Vietnam and to phenotypically characterize (serovars and antibiotic resistance) the isolates. A total of 300 chicken carcasses were collected from two cities and two provinces in Vietnam. Salmonella counts on the samples were determined according to the most-probable-number (MPN) method of the U.S. Department of Agriculture, Food Safety and Inspection Service (USDA-FSIS). A total of 457 isolates were serotyped and tested for antibiotic susceptibility. Overall, 48.7% of chicken samples were Salmonella positive with a count of 2.0 log MPN per carcass. There were no significant differences (P > 0.05) in log MPN per carcass by the study variables (market type, storage condition, and chicken production system). There was a significant difference (P < 0.05) in Salmonella-positive prevalence by chicken production system. Among the 22 Salmonella serovars identified, Albany was the most frequent (34.1%), followed by Agona (15.5%) and Dabou (8.8%). Resistance to at least one antibiotic was common (i.e., 73.3%), with high resistance to tetracycline (59.1%) and ampicillin (41.6%). Resistance to three antibiotics was the most frequently found multidrug resistance profile (17.7%, n = 81); the profile that was resistant to the highest number of drugs was resistant to nine antibiotics (0.7%, n = 3). Only Salmonella Albany posed phenotypic resistance to ceftriaxone (a drug of choice to treat severe cases of salmonellosis). The data revealed that, whereas Salmonella prevalence on raw poultry was high (48.7%), counts were low, which suggests that the exposure risk to Salmonella is low. However, improper storage of raw chicken meat and cross-contamination may increase Salmonella cell counts and pose a greater risk for infection. These data may be helpful in developing risk assessment models and preventing the transmission of foodborne Salmonella from poultry to humans in Vietnam.

Prevalence of Salmonella on chicken carcasses from retail markets in Vietnam.

This study was conducted to estimate the prevalence of Salmonella on chicken carcasses collected from six regions in Vietnam. A total of 1,000 whole, dressed chicken carcasses were collected from five cities and seven provinces across the six regions in Vietnam. Of these, 900 samples were collected from wet markets and 100 from supermarkets. All samples were analyzed for the presence of Salmonella according to a method recommended by the U. S. Department of Agriculture, Food Safety and Inspection Service. The overall Salmonella prevalence was 45.9%. There was no significant difference (P > 0.05) in Salmonella prevalence by (i) location (Ha Noi city, 51.1%; Hai Phong city, 45.6%; Da Nang and Can Tho cities, 45.5%; Bac Ninh province and Ho Chi Minh city, 44.7%; Dong Nai province, 44.6%; Ha Tinh province, 44.4%; Phu Tho province, 43.8%; Lao Cai province, 43.5%; Kien Giang province, 41.9%; and Lam Dong province, 40.9%), (ii) market type (wet market, 46.2%; supermarket samples, 43.0%), and (iii) storage temperature at retail (ambient storage, 46.4%; chilled storage, 45.1%). Hence, Salmonella presence on poultry meat in Vietnam was not associated with a specific city or province, market type, or storage temperature at retail. Strategies to reduce Salmonella levels on raw poultry in Vietnam should be undertaken to improve the safety of poultry products and reduce the incidence of human salmonellosis from poultry consumption.

Prevalence of Salmonella on retail chicken meat in Russian Federation.

The objective of this study was to estimate the prevalence of Salmonella on raw retail chicken meat in Russia. Broiler chicken carcasses (n = 698) were collected from three regions of Russia: central (i.e., Moscow area), northwest (i.e., St. Petersburg area), and southern (i.e., Krasnodar area). In each region, samples were collected to represent various cities and districts, as well as different types of retail stores and carcass storage temperatures (i.e., chilled and frozen). All chicken samples were analyzed for the presence of Salmonella using a whole-carcass rinse method. The overall Salmonella prevalence was 31.5%. There were significant differences (P < 0.05) in Salmonella prevalence by (i) region-29.3% (n = 464) in Moscow, 38.5% (n = 192) in St. Petersburg, and 23.8% (n = 42) in Krasnodar; (ii) retail store type-28.8% (n = 236) in hypermarkets, 31.9% (n = 260) in supermarkets (part of chain stores), 44.3% (n = 61) in independent supermarkets, 42.9% (n = 28) in independent minimarkets, and 26.6% (n = 113) in wet markets; and (iii) poultry company-34.3% (n = 545) on chickens produced by integrated companies compared with 22.9% (n = 118) on chickens produced by nonintegrated companies. Strategies such as good agriculture and management practices should be enhanced to reduce Salmonella prevalence on raw poultry in Russia and therefore increase the safety of chicken products.

Prevalence of Salmonella on retail broiler chicken meat carcasses in Colombia.

A cross-sectional study was performed to estimate the prevalence of Salmonella on retail market chicken carcasses in Colombia. A total of 1,003 broiler chicken carcasses from 23 departments (one city per department) were collected via a stratified sampling method. Carcass rinses were tested for the presence of Salmonella by conventional culture methods. Salmonella strains were isolated from 27 % of the carcasses sampled. Logistic regression analysis was used to determine potential risk factors for Salmonella contamination associated with the chicken production system (conventional versus free-range), storage condition (chilled versus frozen), retail store type (supermarket, independent, and wet market), poultry company (integrated company versus nonintegrated company), and socioeconomic stratum. Chickens from a nonintegrated poultry company were associated with a significantly (P < 0.05) greater risk of Salmonella contamination (odds ratio, 2.0) than were chickens from an integrated company. Chilled chickens had a significantly (P < 0.05) higher risk of Salmonella contamination (odds ratio, 4.3) than did frozen chicken carcasses.

Prevalence of Salmonella on raw poultry at retail markets in China.

Data regarding Salmonella on raw poultry are very limited in China. The objective of this study was to determine the prevalence of Salmonella on raw poultry at the retail level in six provinces and two national cities in China. Whole chicken carcasses (n = 1,152) were collected from three types of retail markets (large, small, and wet). All samples were analyzed for the presence of Salmonella by using the U.S. Department of Agriculture, Food Safety Inspection Service method. Of 1,152 chicken samples, overall Salmonella prevalence was 52.2%. The highest prevalence was observed in Guangxi Province (65.3%), next in Guangdong Province (64.6%), and then in Beijing (63.9%), Shaanxi Province (50.7%), Henan Province (47.9%), Shanghai (44.4%), and Fujian Province (42.4%), and lowest prevalence was observed in Sichuan Province (38.9%). Salmonella prevalence was significantly different among the six provinces and two national cities. Salmonella prevalence was highest in the wet markets (54.4%) compared with the large markets (50.3%) and the small markets (52.1%), but differences were not significant (P > 0.05). Good manufacturing practices, good agricultural practices, and hazard analysis critical control point systems for Salmonella control in poultry production at the farm, processing, and retail level should be implemented.

Introduction to the ILSI Research Foundation/Risk Science Institute reports from the expert working group on neurodevelopmental endpoints.

In 2004 the International Life Sciences Institute (ILSI) Risk Science Institute established an expert working group to assess the lessons learned from the implementation of standardized tests for developmental neurotoxicity in experimental animals. This introduction summarizes the working group process and the four reports from the expert working group addressing: the use of positive controls, understanding variability, appropriate statistical techniques, and interpretation. The reports address the 1991 US Environmental Protection Agency standardized protocol for evaluation of developmental neurotoxicity (DNT) and the 2007 Organisation for Economic Co-operation and Development (OECD) Test Guidelines for DNT. The EPA protocol is comprised of tests for evidence of deficits in neurobehavioral function, including auditory startle habituation, motor activity, associative learning and memory, and neuropathologic examination, including simple morphometric analysis.

Framework for identification and collection of data useful for risk assessments of microbial foodborne or waterborne hazards: a report from the International Life Sciences Institute Research Foundation Advisory Committee on data collection for microbial risk assessment.

A microbial risk assessment (MRA) can provide the scientific basis for risk management decision making. Much data are needed to complete an MRA, including quantitative data for pathogens in foods. The purpose of this document was to provide information on data needs and data collection approaches for MRAs that will be useful for national governments, particularly in developing countries. A framework was developed, which included the following activities: (i) identify the purpose of data collection-this should include stating the specific question(s) to be addressed; (ii) identify and gather existing data-this should include a determination of whether the data are sufficient to answer questions to be addressed; (iii) develop and implement a data collection strategy; (iv) analyze data and draw conclusions; and (v) use data to answer questions identified at the start of the process. The key data needs identified for an MRA were as follows: (i) burden of foodborne or waterborne disease; (ii) microbial contamination of foods; and (iii) consumption patterns. In addition, dose-response data may be necessary, if existing dose-response data cannot be used to estimate dose response for the population of interest. Data should be collected with a view to its use in risk management decision making. Standard sampling and analysis methods should be used to ensure representative samples are tested, and care should be taken to avoid bias when selecting data sets. A number of barriers to data collection were identified, including a lack of clear understanding of the type of data needed to undertake an MRA, which is addressed in this document.

Framework for use of toxicity screening tools in context-based decision-making.

One of the principal applications of toxicology data is to inform risk assessments and support risk management decisions that are protective of human health. Ideally, a risk assessor would have available all of the relevant information on (a) the toxicity profile of the agent of interest; (b) its interactions with living systems; and (c) the known or projected exposure scenarios: to whom, how much, by which route(s), and how often. In practice, however, complete information is seldom available. Nonetheless, decisions still must be made. Screening-level assays and tools can provide support for many aspects of the risk assessment process, as long as the limitations of the tools are understood and to the extent that the added uncertainty the tools introduce into the process can be characterized and managed. Use of these tools for decision-making may be an end in itself for risk assessment and decision-making or a preliminary step to more extensive data collection and evaluation before assessments are undertaken or completed and risk management decisions made. This paper describes a framework for the application of screening tools for human health decision-making, although with some modest modification, it could be made applicable to environmental settings as well. The framework consists of problem formulation, development of a screening strategy based on an assessment of critical data needs, and a data analysis phase that employs weight-of-evidence criteria and uncertainty analyses, and leads to context-based decisions. Criteria for determining the appropriate screening tool(s) have been identified. The choice and use of the tool(s) will depend on the question and the level of uncertainty that may be appropriate for the context in which the decision is being made. The framework is iterative, in that users may refine the question(s) as they proceed. Several case studies illustrate how the framework may be used effectively to address specific questions for any endpoint of toxicity.

Role of quantitative risk assessment and food safety objectives in managing Listeria monocytogenes on ready-to-eat meats.

Listeria monocytogenes may be found on ready-to-eat (RTE) meats, posing a public health risk. To minimize the public health impact, an appropriate level of protection (ALOP) can be established for a population with respect to L. monocytogenes, and ideally should be based on a scientific assessment of the risk, as well as societal and economic factors. Food safety systems can be based on meeting the ALOP. Food safety objectives (FSO) provide a link between the ALOP and performance objectives that are established to control a foodborne hazard. An FSO can be used as a risk management tool for L. monocytogenes in RTE meats, as the FSO establishes the stringency of the measures being used to control the hazard, by specifying the frequency and/or cell number of the pathogen in the food that should not be exceeded at the time of consumption. Typically, this requires setting performance objectives or performance criteria at an earlier point in the food chain, to ensure that the product will meet the FSO. Establishing an FSO requires an assessment of the risk of the hazard to the population of interest. Risk management strategies such as use of HACCP systems and Good Manufacturing Practices can then be used to ensure that the FSO is met.

Direct dosing of preweaning rodents in toxicity testing and research: deliberations of an ILSI RSI Expert Working Group.

Laboratory animal studies designed to assess the effects of exposure of a test substance during postnatal development are commonly utilized in basic research and to evaluate potential hazard to children for chemical and pharmaceutical regulation. Direct dosing, defined here as the administration of a test substance directly to a preweaning mammal, has been identified as a useful tool that can be used in the conduct of such studies for regulatory purposes. The International Life Sciences Institute Risk Science Institute (ILSI RSI) convened an Expert Working Group to develop guidance on the design and implementation of direct dosing regulatory studies on preweaning mammals, which was published as an ILSI monograph in 2003 (Zoetis and Walls, Principles and Practices for Direct Dosing of Pre-Weaning Mammals in Toxicity Testing and Research, Washington, DC: ILSI Press, 2003). A summary of the Working Group conclusions regarding direct dosing studies with laboratory rodents are presented here, although the ILSI monograph also includes rabbits, canines, swine and nonhuman primates. Issues to be considered when designing the protocol include selection of the test species, the route of administration, dose levels, and the timing of dosing. Knowledge of the maturational status of the test species and information on critical windows of development are important in creating a valid study design. Most common routes of administration (e.g., oral, inhalation, injection) are possible with typical laboratory species; however, adjustments may be necessary due to practical considerations. Information on the pharmacokinetic profile in young animals versus adults and in the test species versus humans is very useful for determining dosing parameters. The conduct of the study and the interpretation of the data will be improved by an understanding of confounding factors as well as statistical and biological issues specific for postnatal studies. Ultimately, the success of the study will depend upon careful preparation, including thorough training of the technical staff.

Validation of Predictive Mathematical Models Describing the Growth of Listeria monocytogenes.

Growth of Listeria monocytogenes and Listeria innocua in commercially available sterile homogeneous foods was investigated at different temperatures, pH values, and NaCl concentrations. Growth data were fitted to the Gompertz equation and the resulting growth kinetics were compared with predictions from the Pathogen Modeling Program and Food MicroModel. In general, good agreement was obtained when comparing growth rates and generation times for both models. Differences were observed when comparing lag phases, which ranged from 117 h shorter to 4.9 h longer than predicted for L. monocytogenes . Despite differences in lag phase, under most conditions, the models gave good predictions of microbial growth. Predictive modeling appears to be a useful tool in determining growth rates of Listeria in foods.

Validation of Predictive Mathematical Models Describing Growth of Staphylococcus aureus.

An investigation was performed on the growth of Staphylococcus aureus in a commercially available, sterile, homogeneous food at 12°C with 1.2 and 5.9% NaCl; at 25°C with 10.4% NaCl; and at 20 and 35°C with 1.2, 5.3, 12.5, and 15.8% NaCl; over a pH range of 5.5 to 7.5. Growth data were fitted to the Gompertz equation and the resulting growth kinetics were compared with predictions from the Pathogen Modeling Program (PMP) and Food MicroModel (FMM). For the PMP, predicted lag-phase durations varied from 0.5 to 130 h longer than the observed values. In general, close agreement with growth rates was obtained but there was a 10-fold difference in one case. For FMM, predicted lag-phase durations ranged from 27 h shorter to 47 h longer than the observed values. Again, close agreement with growth rates was obtained, but in one case a fivefold difference was observed. In general, for the sterile foods used under the growth conditions tested, the models underestimated the growth of S. aureus . This implies that while the models can be used as a guide to indicate growth rates in foods they should not be relied upon as the sole determinant of the product's safety.