Assessing reproductive effects on fish populations: an evaluation of methods to predict the reproductive strategy of fishes.

Many environmental monitoring programs include an assessment of the health of fish populations using a sentinel species and include an indicator of reproductive potential. Knowledge of the reproductive strategy of the fish species is critical for data interpretation but is not always known. The reproductive strategy of a species can be determined from detailed histological analyses of ovaries throughout the reproductive cycle; however, these studies can be costly and can delay the implementation of a monitoring program. Three quick and cost-effective methods of predicting the reproductive strategy (annual single spawning or annual multiple spawning) are evaluated in this study using predicted probabilities from binary logistic regression models as a means of classifying the reproductive strategies of 18 different fish species in Atlantic Canada. The first method was based on the hypothesis that the variability in the ovary weight-body weight relationship in prespawning females is higher in multiple spawners. This method did not have a good classification rate due to some multiple spawners having low variability. The other two methods involved predictor variables representing the proportion of oocytes in different stages of development and predictor variables representing the distribution of oocyte sizes during the prespawning season for 111 fish (25 different samples for species). Predicted probabilities from these regression models could be used to correctly classify the reproductive strategies of all 25 samples (development stage model) and all but one sample (oocyte size distribution model). These models can be used to estimate the reproductive strategy of a species from a single sample of fish collected during the prespawning period to support species selection and data interpretation in environmental monitoring programs.

Reproductive strategies and seasonal changes in the somatic indices of seven small-bodied fishes in Atlantic Canada in relation to study design for environmental effects monitoring.

Small-bodied fishes are more commonly being used in environmental effects monitoring (EEM) studies. There is a lack of understanding of the biological characteristics of many small-bodied species, which hinders study designs for monitoring studies. For example, 72% of fish population surveys in Canada's EEM program for pulp and paper mills that used small-bodied fishes were conducted outside of the reproductive period of the species. This resulted in an inadequate assessment of the EEM program's primary effect endpoint (reproduction) for these studies. The present study examined seasonal changes in liver size, gonad size, and condition in seven freshwater and estuarine small-bodied fishes in Atlantic Canada. These data were used to examine differences in reproductive strategies and patterns of energy storage among species. Female gonadal recrudescence in all seven species began primarily in the 2-month period in the spring before spawning. Male gonadal development was concurrent with females in five species; however, gonadal recrudescence began in the fall in male three-spined stickleback (Gasterosteus aculeatus) and slimy sculpin (Cottus cognatus). The spawning period for each species was estimated from the decline in relative ovary size after its seasonal maximum value in spring. The duration of the spawning period reflected the reproductive strategy (single vs multiple spawning) of the species. Optimal sampling periods to assess reproductive impacts in each species were determined based on seasonal changes in ovary size and were identified to be during the prespawning period when gonads are developing and variability in relative gonad size is at a minimum.

Negative consequences of using α = 0.05 for environmental monitoring decisions: a case study from a decade of Canada's Environmental Effects Monitoring Program.

Using the traditional α = 0.05 significance level for null hypothesis significance tests makes assumptions about relative costs of Type I vs relevant Type II errors and inflates their combined probabilities. We have examined the results of 1254 monitoring tests conducted under the Canadian Environmental Effects Monitoring (EEM) program from 1992 to 2003, focusing on how the choice of α affected the relative probabilities and implied costs of Type I and Type II errors. Using α = 0.05 resulted in implied relative costs of Type I vs Type II errors that were both inconsistent among monitoring end points and also inconsistent with the philosophy of the monitoring program. Using α = 0.05 also resulted in combinations of Type I and II error that were 15-17% larger than those for "optimal" α levels set to minimize Type I and II errors for each study, and 12% of all monitoring tests would have reached opposite conclusions had they used these optimal α levels for decision-making. Thus, if the Canadian EEM program used study-specific optimal α levels, they would reduce the incidence of relevant errors and eliminate inconsistent implied relative costs of these errors. Environmental research and monitoring programs using α = 0.05 as a decision-making threshold should re-evaluate the usefulness of this "one-size-fits-all" approach.

Evaluation of antimicrobial resistance phenotypes for predicting multidrug-resistant Salmonella recovered from retail meats and humans in the United States.

Although multidrug-resistant (MDR) non-Typhi Salmonella (NTS) strains are a concern in food production, determining resistance to multiple antimicrobial agents at slaughter or processing may be impractical. Single antimicrobial resistance results for predicting multidrug resistance are desirable. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value were used to determine each antimicrobial agent's ability to predict MDR phenotypes of human health significance: ACSSuT (resistance to at least ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, tetracycline) in NTS isolates, and MDR-AmpC-SN (resistance to ACSSuT, additional resistance to amoxicillin-clavulanate and to ceftiofur, and decreased susceptibility [MIC >= 2 microg/ml] to ceftriaxone) in NTS serotype Newport. The U.S. National Antimicrobial Resistance Monitoring System determined MICs to 15 or more antimicrobial agents for 9,955 NTS isolates from humans from 1999 to 2004 and 689 NTS isolates from retail meat from 2002 to 2004. A total of 847 (8.5%) human and 26 (3.8%) retail NTS isolates were ACSSuT; 995 (10.0%) human and 16 (2.3%) retail isolates were serotype Newport. Among Salmonella Newport, 204 (20.5%) human and 9 (56.3%) retail isolates were MDR-AmpC-SN. Chloramphenicol resistance provided the highest PPVs for ACSSuT among human (90.5%; 95% confidence interval, 88.4 to 92.3) and retail NTS isolates (96.3%; 95% confidence interval, 81.0 to 99.9). Resistance to ceftiofur and to amoxicillin-clavulanate and decreased susceptibility to ceftriaxone provided the highest PPVs (97.1, 98.1, and 98.6%, respectively) for MDR-AmpC-SN from humans. High PPVs for these agents applied to retail meat MDR-AmpC-SN, but isolate numbers were lower. Variations in MIC results may complicate ceftriaxone's predictive utility. Selecting specific antimicrobial resistance offers practical alternatives for predicting MDR phenotypes. Chloramphenicol resistance works best for ACSSuT-NTS, and resistance to ceftiofur, amoxicillin-clavulanate, or chloramphenicol works best for MDR-AmpC-SN.

Effects of pulp and paper mill effluent on fish: a temporal assessment of fish health across sampling cycles.

The Canadian environmental effects monitoring (EEM) program is a regulated, cyclical, industry-funded program designed to determine whether receiving water impacts exist when a mill is in compliance with its discharge limits. The results from three cycles of the fish monitoring program (1992 to 2004) are available from over 200 surveys of fish compared between sites located upstream and downstream of pulp and paper mill effluent outfalls. Previous meta-analyses have shown a national average response pattern across cycles characterized by an increase in endpoints measuring energy storage and growth and a decrease in a reproductive endpoint, consistent with a response of nutrient enrichment in combination with some form of metabolic disruption. Although the national average pattern of effects was temporally consistent, there was some variability in the magnitude of effects among cycles. Questions were raised as to whether the intercycle variability was due to changes in effluent quality or due, at least in part, to other factors. The present study compares responses over the first three cycles, and shows that the choice of sentinel species is likely to be a major contributing factor to the variability in observed effects. Subset analyses using studies from mills that used the same sentinel species across cycles reveal fairly uniform responses and little evidence of significant improvements in overall fish health from cycles one to three. However, a meta-analysis using 1991 data collected from 10 mills before the implementation of the EEM program and data from the same mills collected during cycles one to three of the program reveal significantly reduced effects on relative liver weight and potential improvements in other endpoints.

Dealing with heterogeneous regression slopes in analysis of covariance: new methodology applied to environmental effects monitoring fish survey data.

Analysis of covariance (ANCOVA) is a powerful statistical method which incorporates one or more covariates into the analysis to reduce error associated with measurement. ANCOVA (modeling response as a function of fish size) is frequently used to analyze environmental effects monitoring (EEM) fish survey data. In approximately 12% of fish survey data sets taken from cycles 1 to 3 of Environment Canada's EEM database for pulp and paper mills, the standard assumption of parallel regression slopes is not met. For the first three cycles of the EEM program, these data sets were classified as indicating a mill effect, but for the most part were excluded from subsequent analyses aimed at quantifying the effect. We present two different methods for initially dealing with data sets that exhibit heterogeneous slopes so that they can be analyzed using the parallel slope model. The first method identifies data sets where heterogeneous slopes are forced by a few high-influence observations. The second approach identifies data sets where a model with heterogeneous slopes is statistically, but not practically, significant: with a high coefficient of determination for the parallel slope model. These new methodologies are applied to EEM pulp and paper data sets and about 55% of cases with heterogeneous slopes can be described by a parallel slope model. We also discuss a third method that can be used to describe mill effects when regression slopes remain heterogeneous even after applying the above two methods, enabling comparison with a critical effect size. These new methodologies could benefit the EEM program by enabling more data sets to be incorporated into meta-analyses and be used to make more equitable mill monitoring decisions in the future.

Non-indigenous predators threaten ecosystem engineers: Interactive effects of green crab and oyster size on American oyster mortality.

Non-indigenous green crabs (Carcinus maenas) are emerging as important predators of autogenic engineers like American oysters (Crassostrea virginica) throughout the eastern seaboard of Canada and the United States. To document the spreading distribution of green crabs, we carried out surveys in seven sites of Prince Edward Island during three fall seasons. To assess the potential impact of green crabs on oyster mortality in relation to predator and prey size, we conducted multiple predator-prey manipulations in the field and laboratory. The surveys confirmed an ongoing green crab spread into new productive oyster habitats while rapidly increasing in numbers in areas where crabs had established already. The experiments measured mortality rates on four sizes of oysters exposed to three sizes of crab, and lasted 3-5 days. The outcomes of experiments conducted in Vexar® bags, laboratory tanks and field cages were consistent and were heavily dependent on both crab size and oyster size: while little predation occurred on large oysters, large and medium green crabs preyed heavily on small sizes. Oysters reached a refuge within the 35-55 mm shell length range; below that range, oysters suffered high mortality due to green crab predation and thus require management measures to enhance their survival. These results are most directly applicable to aquaculture operations and restoration initiatives but have implications for oyster sustainability.

Testing against "normal" with environmental data.

Normal ranges are some fraction of a reference distribution deemed to represent an expected condition, typically 95%. They are frequently used as the basis for generic criteria for monitoring programs designed to test whether a sample is outside of "normal," as in reference-condition approach studies. Normal ranges are also the basis for criteria for more classic environmental effects monitoring programs designed to detect differences in mean responses between reference and exposure areas. Limits on normal ranges are estimated with error that varies depending largely on sample size. Direct comparison of a sample or a mean to estimated limits of a normal range will, with some frequency, lead to incorrect conclusions about whether a sample or a mean is inside or outside the normal range when the sample or the mean is near the limit. Those errors can have significant costs and risk implications. This article describes tests based on noncentral distributions that are appropriate for quantifying the likelihood that samples or means are outside a normal range. These noncentral tests reverse the burden of evidence (assuming that the sample or mean is at or outside normal), and thereby encourage proponents to collect more robust sample sizes that will demonstrate that the sample or mean is not at the limits or beyond the normal range. These noncentral equivalence and interval tests can be applied to uni- and multivariate responses, and to simple (e.g., upstream vs downstream) or more complex (e.g., before vs after, or upstream vs downstream) study designs. Statistical procedures for the various tests are illustrated with benthic invertebrate community data collected as part of the Regional Aquatics Monitoring Program (RAMP) in the vicinity of oil sands operations in northern Alberta, Canada. An Excel workbook with functions and calculations to carry out the various tests is provided in the online Supplemental Data. Integr Environ Assess Manag 2017;13:188-197. © 2016 SETAC.

Plasmid-mediated quinolone resistance in non-Typhi serotypes of Salmonella enterica.

BACKGROUND: Serious infections with Salmonella species are often treated with fluoroquinolones or extended-spectrum beta-lactams. Increasingly recognized in Enterobacteriaceae, plasmid-mediated quinolone resistance is encoded by qnr genes. Here, we report the presence of qnr variants in human isolates of non-Typhi serotypes of Salmonella enterica (hereafter referred to as non-Typhi Salmonella) from the United States National Antimicrobial Resistance Monitoring System for Enteric Bacteria. METHODS: All non-Typhi Salmonella specimens from the United States National Antimicrobial Resistance Monitoring System for Enteric Bacteria collected from 1996 to 2003 with ciprofloxacin minimum inhibitory concentrations > or = 0.06 microg/mL (233 specimens) and a subset with minimum inhibitory concentrations < or = 0.03 microg/mL (102 specimens) were screened for all known qnr genes (A, B, and S) by polymerase chain reaction. For isolates with positive results, qnr and quinolone resistance-determining region sequences were determined. Plasmids containing qnr genes were characterized by conjugation or transformation. RESULTS: Conjugative plasmids harboring qnrB variants were detected in 7 Salmonella enterica serotype Berta isolates and 1 Salmonella enterica serotype Mbandaka isolate. The S. Mbandaka plasmid also had an extended-spectrum beta -lactamase. Variants of qnrS on nonconjugative plasmids were detected in isolates of Salmonella enterica serotype Anatum and Salmonella enterica serotype Bovismorbificans. CONCLUSIONS: Plasmid-mediated quinolone resistance appears to be widely distributed, though it is still uncommon in non-Typhi Salmonella isolates from the United States, including strains that are quinolone susceptible by the criteria of the Clinical and Laboratory Standards Institute (formerly the National Committee for Clinical Laboratory Standards). The presence of this gene in non-Typhi Salmonella that causes infection in humans suggests potential for spread through the food supply, which is a public health concern.

The clonal spread of multidrug-resistant non-typhi Salmonella serotypes.

Non-typhoid Salmonella are one of the most important organisms causing food-borne diseases worldwide. There have been significant increases in developed countries in recent years in the occurrence of resistance, in particular multidrug resistance phenotypes, in non-typhoid Salmonella spp. Such increases have been observed in many countries, not only within the European community but also the Americas and Southeast Asia. Of particular concern is the increasing detection of Salmonella isolates displaying resistance to key antimicrobials, notably fluoroquinolones and third-generation cephalosporins. An important factor associated with this increase in multidrug resistance among particular Salmonella spp. is the national and international spread of certain clonal genotypes, the most recent being the global epidemic spread of multidrug-resistant S. Typhimurium DT104, since the early 1990s. In this review, we describe examples where particular antimicrobial-resistant Salmonella serotypes emerged, persisted for periods of time, and then quickly decreased in prevalence.

Salmonella resistant to extended-spectrum cephalosporins: prevalence and epidemiology.

Salmonella resistant to extended-spectrum cephalosporins (ESCs) have emerged worldwide since 1988. By 2004, 43 countries had reported this public health problem. Resistance was mediated by classical extended-spectrum beta-lactamases, plasmid-mediated cephalosporinases, and recently a class A carbapenemase. Of these, CMY-2 is the most widely disseminated enzyme. Salmonella enterica serotype Typhimurium and S. enterica serotype Enteritidis are the most common serovars associated with ESC resistance in human infections. Many outbreaks in humans have been reported, most often among children and neonates. ESC-resistant Salmonella is frequently recovered from animals and food, with poultry as primary food source, suggesting that humans are often infected by these routes.

Surveillance for human Salmonella infections in the United States.

Surveillance for human Salmonella infections plays a critical role in understanding and controlling foodborne illness due to Salmonella. Along with its public health partners, the Centers for Disease Control and Prevention (CDC) has several surveillance systems that collect information on Salmonella infections in the United States. The National Salmonella Surveillance System, begun in 1962, receives reports of laboratory-confirmed Salmonella infections through state public health laboratories. Salmonella outbreaks are reported by state and local health departments through the Foodborne Disease Outbreak Reporting System, which became a Web-based, electronic system (eFORS) in 2001. PulseNet facilitates the detection of clusters of Salmonella infections through standardized molecular subtyping (DNA "fingerprinting") of isolates and maintenance of "fingerprint" databases. The National Antimicrobial Resistance Monitoring System for Enteric Bacteria (NARMS) monitors antimicrobial resistance in Salmonella by susceptibility testing of every 20th Salmonella isolate received by state and local public health laboratories. FootNet is an active surveillance system that monitors Salmonella infections in sentinel areas, providing population-based estimates of infection rates. Efforts are underway to electronically link all of the Salmonella surveillance systems at CDC to facilitate optimum use of available data and minimize duplication.

Invasive infection with multidrug-resistant Salmonella enterica serotype typhimurium definitive type 104 among HIV-infected adults.

BACKGROUND: Multidrug-resistant Salmonella enterica serotype Typhimurium definitive type 104 (MRDT104), with resistance to at least ampicillin, chloramphenicol, streptomycin, sulfamethoxazole, and tetracycline (R-type ACSSuT), was first detected in the United States in 1985, and the prevalence increased to account for nearly 7% of Salmonella infections in 1998. METHODS: A retrospective study of S. Typhimurium infections in an urban health care system assessed whether infection with an antibiotic-resistant strain--and specifically MRDT104--was associated with invasive disease or HIV infection. Sixty cases of S. Typhimurium infection were identified. RESULTS: Of the 50 isolates available for analysis, 30 (60%) were MRDT104. Pathogens were isolated from blood in 25 (83%) of 30 patients infected with MRDT104, compared with 10 (50%) of 20 patients who were infected with non-MRDT104 strains (P = .01). Among isolates obtained from 32 HIV-infected patients, 19 (95%) of 20 MRDT104 isolates were from blood specimens, compared with 8 (66%) of 12 non-MRDT104 isolates (P = .05). CONCLUSIONS: MRDT104 accounted for the majority of S. Typhimurium infections in this patient population, and MRDT104 infections were more invasive than non-MRDT104 infections, particularly in HIV-infected persons.

Quantifying natural variability as a method to detect environmental change: Definitions of the normal range for a single observation and the mean of m observations.

The normal range has been defined as the range that encloses 95% of reference values; in practice this range has been defined as the reference mean ± 2 standard deviations (SD). When sample sizes are small and reference data are not normally distributed, the mean ± 2 SDs do not enclose 95% of data values. Prediction intervals (PI) calculated using sample statistics are used in the present study to define the normal range for a single observation and the mean of m observations. The PIs provide confidence limits for the next randomly selected observation (or mean of m observations) from a population. The PIs are defined using normally distributed reference data; normality can typically be achieved with transformations of the data. Covariates can be used to explain some of the variability in the reference distribution, increasing the ability to detect change. When assumptions of normality are not met, alternative methods of defining the normal range are provided. The normal range can be used to quantify natural variability and assess change from the reference distribution. It can be used as an early warning indicator of change in environmental monitoring to identify the need for further investigation.

Reevaluating fluoroquinolone breakpoints for Salmonella enterica serotype Typhi and for non-Typhi salmonellae.

Salmonella enterica infections cause considerable morbidity and mortality worldwide. Antimicrobial therapy may be life-saving for patients with extraintestinal infections with S. enterica serotype Typhi or non-Typhi salmonellae. Because antimicrobial resistance to several classes of traditional first-line drugs has emerged in the past several decades, the quinolone antimicrobial agents, particularly the fluoroquinolones, have become the drugs of choice. Recently, resistance to nalidixic acid has emerged among both Typhi and non-Typhi Salmonella serotypes. Such Salmonella isolates typically also have decreased susceptibility to fluoroquinolones, although minimum inhibitory concentrations of the fluoroquinolones usually are within the susceptible range of the interpretive criteria of the NCCLS. A growing body of clinical and microbiological evidence indicates that such nalidixic acid-resistant S. enterica infections also exhibit a decreased clinical response to fluoroquinolones. In this article, we recommend that laboratories test extraintestinal Salmonella isolates for nalidixic acid resistance, we recommend that short-course fluoroquinolone therapy be avoided for infection with nalidixic acid-resistant extraintestinal salmonellae, and we summarize existing data and data needs that would contribute to reevaluation of the current NCCLS fluoroquinolone breakpoints for salmonellae.

Prior antimicrobial agent use increases the risk of sporadic infections with multidrug-resistant Salmonella enterica serotype Typhimurium: a FoodNet case-control study, 1996-1997.

Several strains of multidrug-resistant (MDR) Salmonella serotype Typhimurium, including MDR S. Typhimurium definitive type 104, cause almost 10% of Salmonella infections among humans in the United States. To determine the risk factors for acquiring sporadic MDR S. Typhimurium infection, we conducted a population-based, case-control study using data from the Foodborne Diseases Active Surveillance Network (FoodNet) during 1996-1997. S. Typhimurium isolates from 5 FoodNet surveillance areas (California, Connecticut, Georgia, Minnesota, and Oregon) were tested for antimicrobial resistance and phage typing. Telephone interviews were conducted with ill persons and matched control subjects. Compared with both control subjects and patients infected with pansensitive strains of S. Typhimurium, patients with MDR S. Typhimurium infection were significantly more likely to have received an antimicrobial agent, particularly an agent to which the Salmonella isolate was resistant, during the 4 weeks preceding illness onset. Prudent antimicrobial agent use among humans and among veterinarians and food-animal producers is necessary to reduce the burden of drug-resistant salmonellosis in humans.

Antimicrobial use in agriculture: controlling the transfer of antimicrobial resistance to humans.

Salmonella and Campylobacter infections occur commonly in children. Some of these infections are severe, requiring treatment with antimicrobial agents. Many classes of antimicrobial agents that are used in humans also are used in food animals for growth promotion, disease prevention, and therapy. The use of such antimicrobial agents in food animals increases the likelihood that human bacterial pathogens that have food animal reservoirs, such as Salmonella or Campylobacter, will develop cross-resistance to drugs approved for use in human medicine. Resistance determinants also may be transmitted from food animals to humans through the food supply with bacteria that usually are commensal, such as Escherichia coli and enterococci. Clinicians should be aware that antimicrobial resistance is increasing in food-borne pathogens and that patients who are taking antimicrobial agents for any reason are at increased risk for acquiring antimicrobial-resistant food-borne infections. Several European countries have demonstrated that restricting the use of antimicrobial agents in food animals can be followed by a decrease in antimicrobial resistance in humans without compromising animal health or significantly increasing the cost of production. Appropriate use of antimicrobial agents in humans and food animals is an important factor in maintaining their effectiveness.