Prevalence of intramammary antibiotic usage in dairy farming.

This research communication describes the lactating intramammary (IMM) antibiotic formulation most used by Irish dairy farmers at farm level through interviewing 202 dairy farmers. The IMM antibiotic usage data is not easily available to the researcher and farming community. This study determined that three commercial formulations (Synulox ™, Tetra Delta™ and Terrexine) made up 81% of the products used at farm level. The formulation Synulox™ was the most used at 34% first preference and 32% second preference and contains amoxicillin/clavulanic, a standard broad spectrum antibiotic, for which mastitis pathogen resistance remains low. The aminoglycosides were used in four of the IMM formulations analysed, including Tetra Delta™ and Terrexine. Of the 12 antibiotics identified in the IMM formulations studied, three including cefalexin, benzylpenicillin and penethamate are classified as highly important antibiotics (HIA) by the World Health Organisation (WHO) whilst the other 8 (dihydrostreptomycin, streptomycin, neomycin, framycetin, kanamycin, amoxicillin/clavulanic acid, and cefquinome), are considered critically important (CIA) for use in human health. This study has generated knowledge of the preferences of lactating IMM formulations used at farm level.

The impact of meteorology on the occurrence of waterborne outbreaks of vero cytotoxin-producing Escherichia coli (VTEC): a logistic regression approach.

This study analyses the relationship between meteorological phenomena and outbreaks of waterborne-transmitted vero cytotoxin-producing Escherichia coli (VTEC) in the Republic of Ireland over an 8-year period (2005-2012). Data pertaining to the notification of waterborne VTEC outbreaks were extracted from the Computerised Infectious Disease Reporting system, which is administered through the national Health Protection Surveillance Centre as part of the Health Service Executive. Rainfall and temperature data were obtained from the national meteorological office and categorised as cumulative rainfall, heavy rainfall events in the previous 7 days, and mean temperature. Regression analysis was performed using logistic regression (LR) analysis. The LR model was significant (p < 0.001), with all independent variables: cumulative rainfall, heavy rainfall and mean temperature making a statistically significant contribution to the model. The study has found that rainfall, particularly heavy rainfall in the preceding 7 days of an outbreak, is a strong statistical indicator of a waterborne outbreak and that temperature also impacts waterborne VTEC outbreak occurrence.

Microbiological assessment of private groundwater-derived potable water supplies in the Mid-West Region of Ireland.

Determining the likelihood that groundwater contains faecal coliforms can aid water resource management in facilitating the protection of drinking water supplies. This study assesses the incidence of the faecal indicator organism Escherichia coli in 125 private water supplies (PWSs) serving individual houses in the Mid-West Region of Ireland. Two factors, aquifer type and rainfall (mm), were chosen as independent variables that can affect the vulnerability of a groundwater body. Using a geographical information system, the relative hydrogeological and climatological features unique to each sampling location were derived. Utilising this information, a logistic regression (LR) model was used to predict the probability of contamination of PWSs with E. coli. The model contained two independent variables: rainfall (mm; p < 0.001) and aquifer characteristics (p = 0.001). The full model, containing both predictors, was statistically significant at p < 0.001, indicating that the model distinguished between the independent variables' relationship to the incidence of contamination. The likelihood of E. coli contamination is greater with increased rainfall and in areas where a bedrock aquifer is dominant. The LR model explained between 27.4% (Cox and Snell R squared) and 36.8% (Nagelkerke R squared) of the variance in contamination and correctly classified 75.2% of cases.

Development of real-time PCR assays for detection and quantification of Bacillus cereus group species: differentiation of B. weihenstephanensis and rhizoid B. pseudomycoides isolates from milk.

Quantitative real-time PCR (qRT-PCR) offers an alternative method for the detection of bacterial contamination in food. This method provides the quantitation and determination of the number of gene copies. In our study, we established an RT-PCR assay using the LightCycler system to detect and quantify the Bacillus cereus group species, which includes B. cereus, B. anthracis, B. thuringiensis, B. weihenstephanensis, B. mycoides, and B. pseudomycoides. A TaqMan assay was designed to detect a 285-bp fragment of the motB gene encoding the flagellar motor protein, which was specific for the detection of the B. cereus group species, excluding B. pseudomycoides, and the detection of a 217-bp gene fragment of a hypothetical protein specific only for B. pseudomycoides strains. Based on three hydrolysis probes (MotB-FAM-1, MotB-FAM-2, and Bpm-FAM-1), it was possible to differentiate B. weihenstephanensis from the B. cereus group species with nonrhizoid growth and B. pseudomycoides from the whole B. cereus group. The specificity of the assay was confirmed with 119 strains belonging to the Bacillus cereus group species and was performed against 27 other Bacillus and non-Bacillus bacteria. A detection limit was determined for each assay. The assays performed well not only with purified DNA but also with DNA extracted from milk samples artificially contaminated with bacteria that belong to the B. cereus group species. This technique represents an alternative approach to traditional culture methods for the differentiation of B. cereus group species and differentiates B. weihenstephanensis and B. pseudomycoides in one reaction.

Genotypic and phenotypic diversity of Ralstonia pickettii and Ralstonia insidiosa isolates from clinical and environmental sources including High-purity Water. Diversity in Ralstonia pickettii.

BACKGROUND: Ralstonia pickettii is a nosocomial infectious agent and a significant industrial contaminant. It has been found in many different environments including clinical situations, soil and industrial High Purity Water. This study compares the phenotypic and genotypic diversity of a selection of strains of Ralstonia collected from a variety of sources. RESULTS: Ralstonia isolates (fifty-nine) from clinical, industrial and environmental origins were compared genotypically using i) Species-specific-PCR, ii) PCR and sequencing of the 16S-23S rRNA Interspatial region (ISR) iii) the fliC gene genes, iv) RAPD and BOX-PCR and v) phenotypically using biochemical testing. The species specific-PCR identified fifteen out of fifty-nine designated R. pickettii isolates as actually being the closely related species R. insidiosa. PCR-ribotyping of the 16S-23S rRNA ISR indicated few major differences between the isolates. Analysis of all isolates demonstrated different banding patterns for both the RAPD and BOX primers however these were found not to vary significantly. CONCLUSIONS: R. pickettii species isolated from wide geographic and environmental sources appear to be reasonably homogenous based on genotypic and phenotypic characteristics. R. insidiosa can at present only be distinguished from R. pickettii using species specific PCR. R. pickettii and R. insidiosa isolates do not differ significantly phenotypically or genotypically based on environmental or geographical origin.

Specific PCR to identify the heavy-metal-resistant bacterium Cupriavidus metallidurans.

The aim of this study is to develop a polymerase chain reaction (PCR) assay for rapid detection of Cupriavidus metallidurans. PCR primers targeting the Signal transduction histidine kinase gene were designed and designated Cm-F1/Cm-R1. Strains of C. metallidurans were positively identified. The size of the PCR products was 437 bp, as expected. This PCR method enables monitoring of industrial, environmental and clinical sources for presence of C. metallidurans.

Novel Tn4371-ICE like element in Ralstonia pickettii and genome mining for comparative elements.

BACKGROUND: Integrative Conjugative Elements (ICEs) are important factors in the plasticity of microbial genomes. An element related to the ICE Tn4371 was discovered during a bioinformatic search of the Ralstonia pickettii 12J genome. This element was analysed and further searches carried out for additional elements.A PCR method was designed to detect and characterise new elements of this type based on this scaffold and a culture collection of fifty-eight Ralstonia pickettii and Ralstonia insidiosa strains were analysed for the presence of the element. RESULTS: Comparative sequence analysis of bacterial genomes has revealed the presence of a number of uncharacterised Tn4371-like ICEs in the genomes of several beta and gamma- Proteobacteria. These elements vary in size, GC content, putative function and have a mosaic-like structure of plasmid- and phage-like sequences which is typical of Tn4371-like ICEs. These elements were found after a through search of the GenBank database. The elements, which are found in Ralstonia, Delftia, Acidovorax, Bordetella, Comamonas, Acidovorax, Congregibacter, Shewanella, Pseudomonas Stenotrophomonas, Thioalkalivibrio sp. HL-EbGR7, Polaromonas, Burkholderia and Diaphorobacter sp. share a common scaffold. A PCR method was designed (based on the Tn4371- like element detected in the Ralstonia pickettii 12J genome) to detect and characterise new elements of this type. CONCLUSION: All elements found in this study possess a common scaffold of core genes but contain different accessory genes. A new uniform nomenclature is suggested for ICEs of the Tn4371 family. Two novel Tn4371-like ICE were discovered and characterised, using the novel PCR method described in two different isolates of Ralstonia pickettii from laboratory purified water.

Evaluation of an Environmental Monitoring Program for the Microbial Safety of Air and Surfaces in a Dairy Plant Environment.

Microbiological hazards can occur when foodstuffs come into contact with contaminated surfaces or infectious agents dispersed by air currents in the manufacturing environment. An environmental monitoring program (EMP) is a critical aspect of sustainable and safe food manufacturing used to evaluate the effectiveness of the microbial controls in place. An effective EMP should be based on risk analysis, taking into account previous sampling history to determine the selection of the sampling points, the scope of the test, and the frequency of analysis. This study involved evaluation of the environmental monitoring regime and microbiological status of a medium-sized dairy plant manufacturing food ingredients, e.g., proteins, milk powders, and dairy fats. The data specific to microbial tests ( n = 3,468), recorded across 124 fixed sampling locations over a 2-year period (2014 to 2015) from air ( n = 1,787) and surfaces ( n = 1,681) were analyzed. The aim of this study was to highlight the strengths and weaknesses of the EMP in a select dairy processing plant. The results of this study outline the selection of sampling locations, the scope of the test, and the frequency of analysis. An analysis of variance revealed subsections of the manufacturing areas with high risk factors, especially the packaging subsection specified for bulk packaging, the atomizer, and the fluidized bed. The temporal and spatial analysis showed the potential to reduce or relocate the monitoring effort, most notably related to total coliforms and Staphylococcus aureus, across the dairy plant due to homogeneity across the sampling subsections with little or no deviations. The results suggest a need to reevaluate the current EMP and the corrective action plan, especially with regard to detection of pathogens. Recommendations for optimization of the EMP are presented to assist the dairy industry with reviewing and revising the control measures and hazard assessment with regard to existing contamination issues.

Development of a hierarchical model for predicting microbiological contamination of private groundwater supplies in a geologically heterogeneous region.

Private groundwater sources in the Republic of Ireland provide drinking water to an estimated 750,000 people or 16% of the national population. Consumers of untreated groundwater are at increased risk of infection from pathogenic microorganisms. However, given the volume of private wells in operation, remediation or even quantification of public risk is both costly and time consuming. In this study, a hierarchical logistic regression model was developed to 'predict' contamination with E. coli based on the results of groundwater quality analyses of private wells (n = 132) during the period of September 2011 to November 2012. Assessment of potential microbial contamination risk factors were categorised into three groups: Intrinsic (environmental factors), Specific (local features) and Infrastructural (groundwater source characteristics) which included a total of 15 variables. Overall, 51.4% of wells tested positive for E. coli during the study period with univariate analysis indicating that 11 of the 15 assessed risk factors, including local bedrock type, local subsoil type, septic tank reliance, 5 day antecedent precipitation and temperature, along with well type and depth, were all significantly associated with E. coli presence (p < 0.05). Hierarchical logistic regression was used to develop a private well susceptibility model with the final model containing 8 of the 11 associated variables. The model was shown to be highly efficient; correctly classifying the presence of E. coli in 94.2% of cases, and the absence of E. coli in 84.7% of cases. Model validation was performed using an external data set (n = 32) and it was shown that the model has promising accuracy with 90% of positive E. coli cases correctly predicted. The developed model represents a risk assessment and management tool that may be used to develop effective water-quality management strategies to minimize public health risks both in Ireland and abroad.

Evaluation of the Complex Nomenclature of the Clinically and Veterinary Significant Pathogen Salmonella.

Salmonella encompasses a vast and highly related population of clinically and veterinary significant pathogens. The genus is responsible for an array of diseases such as typhoid fever and salmonellosis (a variety of illnesses including gastroenteritis), which cause public health issues globally. Even with the global recognition of Salmonella as a significant human and veterinary pathogen, the highly complex and evolving nomenclature system of Salmonella is problematic for clinicians, veterinarians, and microbiologists to comprehend. The present paper offers a review of the ever developing nomenclature for this bacterial species.

Assessment of the Bacterial Diversity of Aircraft Water: Identification of the Frequent Fliers.

The aim of this study was to determine and identify bacteria inhabiting the supply chain of an airline's drinking water using phenotypic and 16S rDNA sequence-based analysis. Water samples (n = 184) were sourced from long-haul and short-haul aircraft, the airline water source and a water service vehicle. In total, 308 isolates were characterised and their identity determined, which produced 82 identified bacterial species belonging to eight classes: γ-Proteobacteria; ß-Proteobacteria; α-Proteobacteria; Bacilli; Actinobacteria; Flavobacteria; Sphingobacteria and Cytophaga. Statistical differences in bacterial diversity were found to exist across sampling locations (X2 = 39.220, p = 0.009) and furthermore, differences were observed (X2 = 15.475, p = 0.030) across aircraft type (long- or short-haul). This study demonstrates the diverse nature of microorganisms within the aircraft drinking water supply chain. To the best of our knowledge, this is the most extensive study undertaken to date of microbial diversity in aircraft drinking water.

Bacteria that Travel: The Quality of Aircraft Water.

The travelling population is increasing globally year on year. International tourist arrival figures reached 1087 million in 2013 and 1133 million in 2014; of which 53% and 54% respectively accounted for air transport. The water on board aircraft is sourced from surface or ground water; piped to a central filling point and distributed to each aircraft by water service vehicles at the home base or at the destination airport. The purpose of this study was to ascertain the microbial, chemical (pH; Total and Free chlorine) and physical (temperature) quality of water from two aircraft, long- and short-haul, as well as from the original water source and the water service vehicle. A total of 154 water samples were collected and analysed. Long-haul flights were found to be significantly poorer in terms of microbial quality than short haul flights (p = 0.015). Furthermore, correlation and regression analysis showed that the water service vehicle was a significant source of increased microbial load in aircraft. Microbial diversity was also demonstrated, with 37 bacterial species identified belonging to eight classes: γ-Proteobacteria; ß-Proteobacteria; α-Proteobacteria; Bacilli; Actinobacteria; Flavobacteria; Sphingobacteria and Cytophaga; using phenotypic and 16S rDNA sequence-based analysis. We present a novel quantified study of aircraft-related potable water supplies.

A study of cross-contamination of food-borne pathogens in the domestic kitchen in the Republic of Ireland.

To date, there have been no published information and empirical data available on the role played by the food preparer in the domestic kitchen in the Republic of Ireland. In this study, we have looked at the incidence of potential food pathogens and their cross-infection in the domestic kitchen during the preparation of a Sunday roast chicken lunch. Key contact sites in the domestic kitchen were sampled, including the chicken carcass before and after the preparation of a roast chicken meal. Twelve contact sites in twenty-five domestic kitchens were analysed and tested for aerobic plate count, Salmonella, Campylobacter, Escherichia coli and Staphylococcus aureus. Our findings identified the ability of food-borne disease microorganisms to become disseminated from infected foods, such as fresh chickens, to hand and food contact surfaces in the domestic kitchen, reiterating the need for consumer awareness and knowledge of effective hygiene procedures in the domestic kitchen.

Past, Present and Future of Sensors in Food Production.

Microbial contamination management is a crucial task in the food industry. Undesirable microbial spoilage in a modern food processing plant poses a risk to consumers' health, causing severe economic losses to the manufacturers and retailers, contributing to wastage of food and a concern to the world's food supply. The main goal of the quality management is to reduce the time interval between the filling and the detection of a microorganism before release, from several days, to minutes or, at most, hours. This would allow the food company to stop the production, limiting the damage to just a part of the entire batch, with considerable savings in terms of product value, thereby avoiding the utilization of raw materials, packaging and strongly reducing food waste. Sensor systems offer major advantages over current systems as they are versatile and affordable but need to be integrated in the existing processing systems as a process analytical control (PAT) tool. The desire for good selectivity, low cost, portable and usable at working sites, sufficiently rapid to be used at-line or on-line, and no sample preparation devices are required. The application of biosensors in the food industry still has to compete with the standard analytical techniques in terms of cost, performance and reliability.

The antibiotic susceptibility of water-based bacteria Ralstonia pickettii and Ralstonia insidiosa.

Ralstonia pickettii and Ralstonia insidiosa are waterborne bacteria that can survive and grow in various water sources, and that are emerging pathogens in hospital settings. Not much is known about the antibiotic resistance of these bacteria. Previous reports of antimicrobial susceptibility have been largely limited to a few clinical strains with no accounting for genotypic or phenotypic diversity or that these species could vary from the set breakpoints. Etests and disc diffusion tests were carried out to compare the antimicrobial susceptibilities to 12 different antibiotics of 68 different isolates of R. pickettii (53) and R. insidiosa (15) from varying environments, which have previously been well characterized both phenotypically and genetically. The majority of the R. pickettii and R. insidiosa isolates showed susceptibility to most of the antibiotics tested in this study. The most effective were found to be the quinolones and sulfamethoxazole/trimethoprim. Antibiotic susceptibility was also found not to vary between environmental niches for R. pickettii and R. insidiosa isolates.

Characterization of Salmonella enterica serotype Typhimurium isolates from human, food, and animal sources in the Republic of Ireland.

A potential epidemic clone of Salmonella enterica serotype Typhimurium DT104, and the possible emergence of S. enterica serotype Typhimurium DT104b, has been identified from the characterization of 67 S. enterica serotype Typhimurium strains from three sources, human gastroenteritis isolates, isolates from food samples, and veterinary isolates, by antimicrobial resistance profiling, phage typing, and pulsed-field gel electrophoresis. Resistance to ampicillin, chloramphenicol, streptomycin, sulfonamides, and tetracycline was found in 77.6% of these strains.