Dissemination Of Antimicrobial-resistant Bacteria Research Articles

Identifying antibiotic-resistant strains via cell sorting and elastic-light-scatter phenotyping

The proliferation and dissemination of antimicrobial-resistant bacteria is an increasingly global challenge and is attributed mainly to the excessive or improper use of antibiotics. Currently, the gold-standard phenotypic methodology for detecting resistant strains is agar plating, which is a time-consuming process that involves multiple subculturing steps. Genotypic analysis techniques are fast, but they require pure starting samples and cannot differentiate between viable and non-viable organisms. Thus, there is a need to develop a better method to identify and prevent the spread of antimicrobial resistance. This work presents a novel method for detecting and identifying antibiotic-resistant strains by combining a cell sorter for bacterial detection and an elastic-light-scattering method for bacterial classification. The cell sorter was equipped with safety mechanisms for handling pathogenic organisms and enabled precise placement of individual bacteria onto an agar plate. The patterning was performed on an antibiotic-gradient plate, where the growth of colonies in sections with high antibiotic concentrations confirmed the presence of a resistant strain. The antibiotic-gradient plate was also tested with an elastic-light-scattering device where each colony’s unique colony scatter pattern was recorded and classified using machine learning for rapid identification of bacteria. Sorting and patterning bacteria on an antibiotic-gradient plate using a cell sorter reduced the number of subculturing steps and allowed direct qualitative binary detection of resistant strains. Elastic-light-scattering technology is a rapid, label-free, and non-destructive method that permits instantaneous classification of pathogenic strains based on the unique bacterial colony scatter pattern.Key points• Individual bacteria cells are placed on gradient agar plates by a cell sorter• Laser-light scatter patterns are used to recognize antibiotic-resistant organisms• Scatter patterns formed by colonies correspond to AMR-associated phenotypes

Diagnostic ultrasonography and antimicrobial resistance of different pathogens associated with canine and feline lower urinary tract disorders

There is a significant issue concerning the dissemination of antimicrobial-resistant bacteria within companion animals. Urinary tract infections (UTIs) are a common problem in veterinary medicine for which empirical antibiotics are utilized. This study aimed to investigate the antimicrobial resistance of different uropathogens associated with UTIs in canine and feline cases. A total of 146 dogs and 162 cats suffered from lower urinary tract disorders were subjected to ultrasonographic and microbiological examination. Cystitis, urinary sediment, and cystic calculi are the most common ultrasonographic abnormalities associated with bacterial UTIs. Bacterial UTIs were obtained in 36.98 % and 25.92 % of cases in dogs and cats, respectively. A low rate of mixed infection was detected in canine cases (3.7 %). E. coli was the most prevalent pathogen isolated from 46.4 % and 66.7 % of canine and feline isolates, respectively followed by Proteus spp. in canine isolates (16.1 %) and Klebsiella spp. in feline isolates (14.3 %). Staphylococcus spp. was isolated from canine cases only with the detection of methicillin-resistant Staphylococcus pseudintermedius (MRSP) strains at 3.6 %. The majority of the isolated strains were resistant to various antibiotic classes, particularly β-lactams. All gram-negative bacteria were susceptible to amikacin, whereas gram-positive strains exhibited 100 % sensitivity to nitrofurantoin and linezolid. Different bacterial species displayed low resistance to carbapenems and fluoroquinolones. Multi-drug resistance was reported in canine and feline strains at 64.3 % and 54.8 %, respectively. These findings prove the crucial necessity to restrict antibiotic consumption depending on urine culture and antibiotic sensitivity tests.

RAW BOVINE MILK AS A RESERVOIR OF MULTI-DRUG RESISTANT, BETA-LACTAMASE-PRODUCING Klebsiella

The transmission of zoonotic bacteria through consumption of raw milk is complicated by the dissemination of antimicrobial-resistant bacteria. The present study was conducted to detect the occurrence of antimicrobial-resistant bacteria (ESBL-/AmpC-producing Klebsiella spp.) in cow’s milk originating from healthy or infected (mastitis) cattle in India. In total, 450 milk samples were collected from apparently healthy cattle and cattle suffering from clinical or sub-clinical mastitis. Out of 455 Klebsiella spp., 67 (14.73%) isolates were found to be ESBL producers in the double-disc diffusion test. The occurrence of ESBL-producing Klebsiella spp. was significantly (p < 0.05) higher in milk samples collected from cattle suffering with mastitis than in healthy cattle. Among the ESBL-producing Klebsiella spp., 56 (83.6%) isolates were also detected that produced AmpC β-lactamases. All the ESBL and AmpC-producing Klebsiella spp. possessed blaCTX-M (100%) and blaAmpC (100%), respectively. The present study revealed a higher occurrence of class 1 integron in ESBL-producing Klebsiella spp. isolates. All ESBL-producing- Klebsiella spp. isolates were multi-drug resistant. The ciprofloxacin- and/or levofloxacin-resistant Klebsiella spp. isolates possessed the quinolone resistance gene (qnrS). The co-trimoxazole-resistant isolates possessed the sul1 and sul2 genes. Phylogenetic analysis of the studied isolates revealed that strains isolated from the same location had a clonal relationship. The study increases consumer awareness of the need to avoid raw milk consumption to prevent the spread of antimicrobial resistance in the community.

Proximity to human settlement is directly related to carriage of critically important antimicrobial-resistant Escherichia coli and Klebsiella pneumoniae in Silver Gulls

Human population and activities play an important role in dissemination of antimicrobial resistant bacteria. This study investigated the relationship between carriage rates of critically important antimicrobial-resistant (CIA-R) Escherichia coli and Klebsiella pneumoniae by Silver Gulls and their proximity to human populations. Faecal swabs (n = 229) were collected from Silver Gulls across 10 southern coastline locations in Western Australia (WA) traversing 650 kms. The sampling locations included main town centres and remote areas. Fluoroquinolone and extended-spectrum cephalosporin-resistant E. coli and K. pneumoniae were isolated and tested for antimicrobial sensitivity. Genome sequencing was performed on n = 40 subset out of 98 E. coli and n = 14 subset out of 27 K. pneumoniae isolates to validate phenotypic resistance profiles and determine the molecular characteristics of strains. CIA-R E. coli and K. pneumoniae were detected in 69 (30.1 %) and 20 (8.73 %) of the faecal swabs respectively. Two large urban locations tested positive for CIA-R E. coli (frequency ranging from 34.3 % to 84.3 %), and/or for CIA-R K. pneumoniae (frequency ranging from 12.5 % to 50.0 %). A small number of CIA-R E. coli (3/31, 9.7 %) were identified at a small tourist town, but no CIA-R bacteria were recovered from gulls at remote sites. Commonly detected E. coli sequence types (STs) included ST131 (12.5 %) and ST1193 (10.0 %). Five K. pneumoniae STs were detected which included ST4568, ST6, ST485, ST967 and ST307. Resistance genes including blaCTX-M-3, blaCTX-M-15 and blaCTX-M-27 were identified in both bacterial species. High-level colonisation of CIA-R E. coli and K. pneumoniae in Silver Gulls in and around urban areas compared to remote locations substantiates that anthropogenic activities are strongly associated with acquisition of resistant bacteria by gulls.

The antimicrobial resistance profiles of Escherichia coli and Pseudomonas aeruginosa isolated from private groundwater wells in the Republic of Ireland

The role of the natural environment in the dissemination of antimicrobial resistant bacteria has been increasingly recognised in the literature. However, knowledge surrounding the critical factors and mechanisms mediating their occurrence is still limited, particularly in relatively ‘pristine’ groundwater environments. In the Republic of Ireland (RoI), a country characterised by high groundwater reliance, household-based (unregulated) wells provide drinking water to 11% of the population. These private wells are generally located in rural areas, where the risk of microbiological contamination is high due to intensive agricultural practices and high reliance on domestic wastewater treatment systems; both of which are also potential sources of antimicrobials and antimicrobial resistant bacteria. Accordingly, the current research sought to elucidate current rates of antimicrobial resistant bacteria and the principal factors associated with their presence in private wells in the RoI. A total of 250 samples (from 132 wells nationwide) were assessed for the presence of faecal (Escherichia coli) and environmental (Pseudomonas aeruginosa) bacteria, with single isolates from each contaminated sample tested phenotypically against 18 and 9 antimicrobials, respectively. Findings show that while 16.7% of E. coli (n = 8/48) were categorically resistant to ≥1 antimicrobial, with a further 79.2% classified as intermediately resistant, no categorical resistance was found among P. aeruginosa isolates (n = 0/6), with just one intermediately resistant isolate detected. Multivariate regression modelling indicates significantly higher odds of resistant E. coli detection in concurrence with elevated cattle density (OR = 1.028, p = 0.032), aligning with findings of highest resistance rates to veterinary antimicrobials (e.g., streptomycin = 14.6%, tetracycline = 12.5%, and ampicillin = 12.5%). Multivariate model results also suggest overland flow culminating in direct wellhead ingress as a primary ingress mechanism for resistant E. coli. Study findings may inform groundwater source protection initiatives and antimicrobial resistance surveillance moving forward.

Residual concentrations of antimicrobial growth promoters in poultry litter favour plasmid conjugation among Escherichia coli.

Considering that plasmid conjugation is a major driver for the dissemination of antimicrobial resistance in bacteria, this study aimed to investigate the effects of residual concentrations of antimicrobial growth promoters (AGPs) in poultry litter on the frequencies of IncFII-FIB plasmid conjugation among Escherichia coli organisms. A 2 × 5 factorial trial was performed invitro, using two types of litter materials (sugarcane bagasse and wood shavings) and five treatments of litter: non-treated (CON), herbal alkaloid sanguinarine (SANG), AGPs monensin (MON), lincomycin (LCM) and virginiamycin (VIR). E.coli H2332 and E.coli J62 were used as donor and recipient strains, respectively. The presence of residues of monensin, lincomycin and virginiamycin increased the frequency of plasmid conjugation among E.coli in both types of litter materials. On the contrary, sanguinarine significantly reduced the frequency of conjugation among E.coli in sugarcane bagasse litter. The conjugation frequencies were significantly higher in wood shavings compared with sugarcane bagasse only in the presence of AGPs. Considering that the presence of AGPs in the litter can increase the conjugation of IncFII-FIB plasmids carrying antimicrobial resistance genes, the real impact of this phenomenon on the dissemination of antimicrobial resistant bacteria in the poultry production chain must be investigated.

Environmental Antimicrobial Resistance in a Small Urban Mediterranean River: A Focus on Endemic Beta-Lactamases in Clinically Relevant Bacteria

Aquatic ecosystems subjected to anthropic pressures are likely hotspots for emergence or dissemination of antimicrobial resistant bacteria. The city of Montpellier is located on a Mediterranean climate watershed that undergoes strong demographic pressures. The aim of the study is to explore antimicrobial resistance, particularly those of clinical concern, in urban rivers flowing in this urban area. The method developed herein to explore antimicrobial resistance is based on cultural and molecular approaches completed by hydrological, hydrogeological, climatic, and physico-chemical data. Hospital vicinity and urbanization density significantly increase cultivable bacterial community, fecal bacteria from human origin, and prevalence of β-lactamases and extended-spectrum β-lactamases encoding-genes without an increase in 16S rDNA gene abundance. A total of 22 multidrug Enterobacterales have been isolated. All Escherichia coli (n = 10) and Klebsiella pneumoniae (n = 6) isolated on a made-house media carried β-lactamases genes, blaCTX-M being the most prevalent (87%), followed by blaTEM (56%) and blaSHV (37%), 56% of these strains carrying two or three of these genes. In urban settings, water quality and infectious risk are generally linked to wastewater treatment plants effluents. This study shows that running waters in urbanized area contribute to the dissemination of antimicrobial resistance, making these environments a reservoir for resistant bacteria with important consideration.

The Emergence, Persistence, and Dissemination of Antimicrobial-Resistant Bacteria in Environmental Hajj Settings and Implications for Public Health

The emergence of antimicrobial resistance is causing the loss of what was once considered the miracle cure. The transmission of antimicrobial resistance during mass gathering is a potential threat in addition to other infectious diseases. Here, we review the English language literature on the rate and the acquisition of antimicrobial resistance during the Hajj. There is a variable incidence of methicillin-resistant Staphylococcus aureus, Escherichia coli, and Enterobacteriaceae. There had been no report of multi-drug-resistant Mycobacterium tuberculosis. Continued surveillance of antimicrobial resistance coupled with public health measures are needed to decrease the rate of emergence of resistance.

Antimicrobial use and detection of cefotaxime-resistant Enterobacteriaceae in the pig production chain, Ogun State, Nigeria

Cefotaxime belongs to the third-generation cephalosporin group of antimicrobials, which are classified as critical for the treatment of infections in humans. The upsurge in the incidence of cefotaxime-resistant (C-R) bacteria from animal sources is of global public health importance. This study investigated the pres- ence of C-R Enterobacteriaceae in the pig production chain in Ogun State, Nigeria, and examined C-R isolates for production of extended-spectrum b-lactamase (ESBL). The knowledge, attitude and practices of pig farmers regarding antimicrobial usage were also investigated. C-R bacteria were detected in 54 (17.8%) out of 303 samples. C-R isolates were identified as Escherichia coli (n = 22), Klebsiella spp. (n = 17), Enterobacter aerogenes (n = 10) and Citrobacter freundii (n = 5). The organisms were present in feces from on-farm pigs (15/109; 13.7%), fresh pork at slaughterhouses (19/40; 47.5%), frozen pork at retail shops (7/28; 25.0%), cutting surfaces of butchers’ tools (7/52; 13.5%), and abattoir effluent water (6/41; 14.6%). No C-R bacteria were detected in ready-to-eat pork. Three isolates of Es. coli and one of K. pneumoniae were ESBL-producers and pos- sessed blaCTX-M-15 ESBL gene variant. ESBL-producing Es. coli belonged to phylo- genetic group A. All C-R isolates were resistant to more than three antimicrobials from different classes of antimicrobials. Tetracycline, ampicillin, amoxicillin, ciprofloxacin and enrofloxacin were among the commonly used antimicrobials in pig production, whereas cephalosporins were rarely used. Farmers knew that pigs could serve as reservoirs of pathogenic bacteria transmissible to humans. However, they were not aware that the use of antimicrobials in pig production could lead to the development and proliferation of antimicrobial-resistant bacteria in pigs. Efforts should be made to improve awareness among farmers on the roles of antimicrobial use in the emergence and dissemination of antimicrobial-resistant bacteria in animal production.

The level of antimicrobial resistance of sewage isolates is higher than that of river isolates in different Escherichia coli lineages

The dissemination of antimicrobial-resistant bacteria in environmental water is an emerging concern in medical and industrial settings. Here, we analysed the antimicrobial resistance of Escherichia coli isolates from river water and sewage by the use of a combined experimental phenotypic and whole-genome-based genetic approach. Among the 283 tested strains, 52 were phenotypically resistant to one or more antimicrobial agents. The E. coli isolates from the river and sewage samples were phylogenetically indistinguishable, and the antimicrobial-resistant strains were dispersedly distributed in a whole-genome-based phylogenetic tree. The prevalence of antimicrobial-resistant strains as well as the number of antimicrobials to which they were resistant were higher in sewage samples than in river samples. Antimicrobial resistance genes were more frequently detected in strains from sewage samples than in those from river samples. We also found that 16 river isolates that were classified as Escherichia cryptic clade V were susceptible to all the antimicrobials tested and were negative for antimicrobial resistance genes. Our results suggest that E. coli strains may acquire antimicrobial resistance genes more frequently and/or antimicrobial-resistant E. coli strains may have higher rates of accumulation and positive selection in sewage than in rivers, irrespective of their phylogenetic distribution.

Characterization of antimicrobial resistance genes in Enterobacteriaceae carried by suburban mesocarnivores and locally owned and stray dogs.

The role of wildlife in the dissemination of antimicrobial-resistant bacteria and antimicrobial resistance genes (ARGs) in the environment is of increasing concern. We investigated the occurrence, richness and transmissibility potential of ARGs detected in the faeces of three mesocarnivore species: the coyote (Canis latrans), raccoon (Procyon lotor) and Virginia opossum (Didelphis virginiana), and of stray and owned dogs in suburban Chicago, IL, USA. Rectal swabs were collected from live-captured coyotes (n=32), raccoons (n=31) and Virginia opossums (n=22). Fresh faecal samples were collected from locally owned (n=13) and stray dogs (n=18) and from the live-captured mesocarnivores, when available. Faecal samples and rectal swabs were enriched to select for Enterobacteriaceae and pooled by mesocarnivore species and dog type (owned or stray). Pooled enriched samples were then analysed for the presence of ARGs using shotgun sequencing. The three mesocarnivore and stray dog samples had twice as many unique ARGs compared to the owned dog sample, which was partly driven by a greater richness of beta-lactamase genes (genes conferring resistance to penicillins and cephalosporins). Raccoon and stray dog samples had the most ARGs in common, suggesting possible exposure to similar environmental sources of ARGs. In addition to identifying clinically relevant ARGs (e.g. blaCMY and qnrB), some ARGs were linked to the class 1 integrase gene, intI1, which may indicate anthropogenic origin. Findings from this pilot investigation suggest that the microbial communities of suburban mesocarnivores and stray dogs can host ARGs that can confer resistance to several antimicrobials used in human and veterinary medicine.

In-Depth Genomic Characterization of a Meropenem-nonsusceptible Pseudomonas otitidis Strain Contaminating Chicken Carcass

Background: The indiscriminate use of antibiotics in food-animal production has a major impact on public health, particularly in terms of contributing to the emergence and dissemination of antimicrobial resistant bacteria in the food-animal production chain. Although Pseudomonas species are recognized as important spoilage organisms in foodstuff, they are also known as opportunistic pathogens associated with hospital-acquired infections. Furthermore, Pseudomonas can play a role as potential reservoirs of antimicrobial resistance genes, which may be horizontally transferred to other bacteria. Considering that cephalosporins (3rd and higher generations) and carbapenems are critically important beta-lactam antimicrobials in human medicine, this study reports the occurrence and genomic characterization of a meropenem-nonsusceptible Pseudomonas otitidis strain recovered from a chicken carcass in Brazil.Materials, Methods & Results: During the years 2018-2019, 72 frozen chicken carcasses were purchased on the retail market from different regions in Brazil. Aliquots from individual carcass rinses were screened for Extended Spectrum Beta-lactamase (ESBL)-producing bacteria in MacConkey agar supplemented with 1mg.L-1 cefotaxime. Phenotypically resistant isolates were further tested for resistance to other antimicrobials and confirmed as ESBL-producers by means of disk-diffusion method using Müller-Hinton agar. Only one meropenen-nonsusceptible isolate was detected and submitted to whole genome sequencing (WGS) in Illumina Miseq. The strain was identified as Pseudomonas otitidis by local alignment of the 16S rRNA sequence using BLASTn and confirmed by Average Nucleotide Identity (ANI) analysis using JspeciesWS database. Genes encoding for antimicrobial resistance were detected by means of Resfinder and Comprehensive Antibiotic Resistance Database (CARD) databases. The phenotypic non-susceptibility to meropenen was attributed to the gene blaPOM-1. A total of 192 different genes encoding for quorum sensing system, antiphagocytosis, iron uptake, efflux pump, endotoxin and toxin, adherence, and secretion systems were detected by means of Virulence Factor Database (VFDB). Pseudomonas otitidis-pan genome was built using Roary-rapid large-scale prokaryote pan genome analysis using the present strain (K_25) and other two P. otitidis genomes (PAM-1, DSM 17224) publicly available at the NCBI. The core genome analysis of the two human strains resulted in similar percentages.Discussion: Carbapenems are critically important drugs for human health and bacterial strains resistant to these antimicrobials pose a public health problem. The blaPOM-1 gene harbored by the Pseudomonas otitidis K_25 strain encodes a metallo-beta-lactamase (MBL) conferring resistance to carbapenems. Pseudomonas otitidis was the first confirmed pathogenic Pseudomonas species expressing MBL constitutively in the absence of inducible beta-lactamase genes. Furthermore, the several virulence genes associated with the capacity of the P. otitidis K_25 to colonize, evade the immune system and cause lesions in the human host confirm this strain as a potential opportunistic pathogen contaminating foodstuff. These reinforce the need to address antimicrobial resistance in a One Health perspective, in which resistant bacteria and resistance determinants circulate among environment, animals and humans.

Quantification and Multidrug Resistance Profiles of Vancomycin-Resistant Enterococci Isolated from Two Wastewater Treatment Plants in the Same Municipality.

Wastewater treatment plants (WWTPs) are points of control for the environmental dissemination of antimicrobial resistant bacteria. Vancomycin-resistant enterococci (VRE) were used as indicators of antimicrobial resistance (AMR) in two WWTPs (biologically aerated filter (BAF) and conventional activated sludge (CAS)) in the same municipality. The removal and abundance of enterococci and VRE as well as the species and antimicrobial resistance profiles of VRE were assessed. Enterococci and VRE from the primary and final effluents were enumerated. Results were assessed from an ecological context. VRE was not selected for by either WWTP but the BAF system outperformed the CAS system for the removal of enterococci/VRE. Enterococcus faecalis (n = 151), E. faecium (n = 94) and E. casseliflavus/E. gallinarum (n = 59) were the dominant VRE species isolated. A decrease in levofloxacin resistance in enterococci was observed in the BAF WWTP. An increase in nitrofurantoin resistant (p < 0.001) and a decrease in quinupristin/dalfopristin (p = 0.003) and streptomycin (p = 0.022) resistant enterococci were observed in the CAS WWTP, corresponding to a shift of VRE from E. faecalis to E. faecium. Wastewater treatment processes can be managed to limit the dissemination of antimicrobial resistance determinants into the surrounding environment.

Antimicrobial resistance in Chile and The One Health paradigm: Dealing with threats to human and veterinary health resulting from antimicrobial use in salmon aquaculture and the clinic.

The emergence and dissemination of antimicrobial-resistant bacteria (ARB) is currently seen as one of the major threats to human and animal public health. Veterinary use of antimicrobials in both developing and developed countries is many-fold greater than their use in human medicine and is an important determinant in selection of ARB. In light of the recently outlined National Plan Against Antimicrobial Resistance in Chile, our findings on antimicrobial use in salmon aquaculture and their impact on the environment and human health are highly relevant. Ninety-five percent of tetracyclines, phenicols and quinolones imported into Chile between 1998 and 2015 were for veterinary use, mostly in salmon aquaculture. Excessive use of antimicrobials at aquaculture sites was associated with antimicrobial residues in marine sediments 8 km distant and the presence of resistant marine bacteria harboring easily transmissible resistance genes, in mobile genetic elements, to these same antimicrobials. Moreover, quinolone and integron resistance genes in human pathogens isolated from patients in coastal regions adjacent to aquaculture sites were identical to genes isolated from regional marine bacteria, consistent with genetic communication between bacteria in these different environments. Passage of antimicrobials into the marine environment can potentially diminish environmental diversity, contaminate wild fish for human consumption, and facilitate the appearance of harmful algal blooms and resistant zoonotic and human pathogens. Our findings suggest that changes in aquaculture in Chile that prevent fish infections and decrease antimicrobial usage will prove a determining factor in preventing human and animal infections with multiply-resistant ARB in accord with the modern paradigm of One Health.

Aeromonas Diversity and Antimicrobial Susceptibility in Freshwater-An Attempt to Set Generic Epidemiological Cut-Off Values.

The importance of the role of environment in the dissemination of antimicrobial resistant bacteria is now well recognized. Thus, bacterial indicators to monitor the phenomena are required. The Aeromonas genus is autochthonous in the aquatic environment and easy to detect in any water type, such as freshwater, or wastewater. These microorganisms are also causing infections in humans and animals (including fish). Furthermore, as Aeromonas spp. is able to acquire antimicrobial resistance mechanisms, it is candidate for indicator bacteria to follow antimicrobial resistance dissemination in aquatic environments. Unfortunately, to date, interpretation criteria for Aeromonas spp. for antimicrobial susceptibility tests are scarce in the literature. No epidemiological cut-off values for Aeromonas are currently available at EUCAST to interpret Minimum Inhibitory Concentrations (MIC). The only interpretation criteria available are clinical breakpoints from CLSI that are adapted from Enterobacteriaceae. Based on the results of MIC distributions obtained for a collection of environmental isolates of Aeromonas, this study aimed at proposing tentative epidemiological cut-off values (COWT) for Aeromonas spp. assessing whether the genus is an acceptable level of definition. Thus, 233 isolates collected from 16 rivers were identified at species level using Maldi-Tof (Bruker). Eleven different species were identified, the most abundant were A. bestiarum (n = 54), A. salmonicida (n = 45), A. sobria (n = 41), and A. eucrenophila (n = 37). 96-well micro-plates containing different concentrations of 15 antimicrobials, namely cefotaxime, ceftazidime, chloramphenicol, colistin, enrofloxacin, erythromycin, florfenicol, flumequine, gentamicin, nalidixic acid, oxolinic acid, streptomycin, temocillin, tetracycline, and trimethoprim-sulfamethoxazole, were prepared. The broth micro-dilution method was used to determine the antimicrobial susceptibility of each isolate. The estimation of COWT values was satisfactory obtained at genus level for all antimicrobials except cefotaxime and erythromycin. This first step is an invitation for other research teams to increase the amount of antimicrobial resistance data collected. Then, robustness of our proposed provisional generic epidemiological cut-off values could be assessed by testing antimicrobial susceptibility of various Aeromonas collections.

Prevalence of Nontyphoidal Salmonella and Salmonella Strains with Conjugative Antimicrobial-Resistant Serovars Contaminating Animal Feed in Texas

The objective of this study was to characterize 365 nontyphoidal Salmonella enterica isolates from animal feed. Among the 365 isolates, 78 serovars were identified. Twenty-four isolates (7.0%) were recovered from three of six medicated feed types. Three of these isolates derived from the medicated feed, Salmonella Newport, Salmonella Typhimurium var. O 5- (Copenhagen), and Salmonella Lexington var. 15+ (Manila), displayed antimicrobial resistance. Susceptibility testing revealed that only 3.0% (12) of the 365 isolates displayed resistance to any of the antimicrobial agents. These 12 isolates were recovered from unmedicated dry beef feed (n = 3), medicated dry beef feed (n = 3), cabbage culls (n = 2), animal protein products (n = 2), dry dairy cattle feed (n = 1), and fish meal (n = 1). Only Salmonella Newport and Salmonella Typhimurium var. O 5- (Copenhagen) were multidrug resistant. Both isolates possessed the IncA/C replicon and the blaCMY-2 gene associated with cephalosporin resistance. Plasmid replicons were amplified from 4 of 12 resistant isolates. Plasmids (40 kb) were Salmonella Montevideo and Salmonella Kentucky. Conjugation experiments were done using 7 of the 12 resistant isolates as donors. Only Salmonella Montevideo, possessing a plasmid and amplifying IncN, produced transconjugants. Transconjugants displayed the same antimicrobial resistance profile as did the donor isolate. Three isolates that amplified replicons corresponding to IncA/C or IncHI2 did not produce transconjugants at 30 or 37°C. The results of this study suggest that the prevalence of antimicrobial-resistant Salmonella contaminating animal feed is low in Texas. However, Salmonella was more prevalent in feed by-products; fish meal had the highest prevalence (84%) followed by animal protein products (48%). Ten of the 35 feed types had no Salmonella contamination. Further investigation is needed to understand the possible role of specific feed types in the dissemination of antimicrobial resistant bacteria.

Exploratory Spatial Mapping of the Occurrence of Antimicrobial Resistance in E. coli in the Community.

The use of antimicrobials over the past six decades has been associated with the emergence and dissemination of antimicrobial-resistant bacteria. To explore local geographical patterns in the occurrence of acquired antimicrobial resistance (AMR), AMR of E. coli causing urinary tract infections (UTI) in the community in the West of Ireland was mapped. All adult patients consulting with a suspected UTI in 22 general practices in the West of Ireland over a nine-month study period were requested to supply a urine sample. Those with a laboratory confirmed E. coli infection were included (n = 752) in the study. Antimicrobial susceptibility testing was performed by standardized disc diffusion. Patient addresses were geocoded. The diameters of the zone of inhibition of growth for trimethoprim (5 μg) and ciprofloxacin (5 μg) for the relevant isolate was mapped against the patient address using ArcGIS software. A series of maps illustrating spatial distribution of AMR in the West of Ireland were generated. The spatial data demonstrated a higher proportion of isolates with AMR from urban areas. Some rural areas also showed high levels of resistant E. coli. Our study is the first to demonstrate the feasibility of using a geographical information system (GIS) platform for routine visual geographical analysis of AMR data in Ireland. Routine presentation of AMR data in this format may be valuable in understanding AMR trends at a local level.

First insights into antimicrobial resistance among faecal Escherichia coli isolates from small wild mammals in rural areas

Wild rodents can be carriers of antimicrobial resistant Escherichia coli. As rodents are known to be involved in the transmission of bacteria of human and animal health concern, they could likewise contribute to the dissemination of antimicrobial resistant bacteria in the environment. The aim of this study was therefore to get first insights into the antimicrobial resistance status among E. coli isolated from wild small mammals in rural areas. We tested 188 faecal isolates from eight rodent and one shrew species originating from Germany. Preselected resistant isolates were screened by minimal inhibitory concentration (MIC) testing or agar diffusion test and subsequent PCR analysis of resistance genes. The prevalence of antimicrobial resistant isolates was low with only 5.5% of the isolates exhibiting resistant phenotypes against at least one antimicrobial compound including beta-lactams, tetracyclines, aminoglycosides and sulfonamides. These results suggest a minor role of wild rodents from rural areas in the cycle of transmission and spread of antimicrobial resistant E. coli into the environment. Nevertheless E. coli with multiple antimicrobial resistances were significantly more often detected in wildlife rodents originating from areas with high livestock density suggesting a possible transmission from livestock to wild rodents.

Recent applications of liquid chromatography–mass spectrometry to residue analysis of antimicrobials in food of animal origin

Residual antimicrobials in food constitute a risk to human health. Although epidemiological data on the real magnitude of their adverse effects are very scarce, they indicate that food could be an important vehicle for evolution and dissemination of antimicrobial-resistant bacteria. Public health agencies in many countries rely on detection by mass spectrometry (MS) for unambiguous identification of residues of antimicrobial agents in animal food products for human consumption. The introduction of relatively inexpensive and robust liquid chromatography (LC)-MS systems has given a strong impulse to the development of confirmatory methods for the above medicines in foodstuffs. The initial part of this review, after a brief introduction into the field of antimicrobials, is dedicated to the most important EU regulations and directives for control of residues of these substances in animal products. The main attention in this review is on the sample-treatment and MS detection systems in use today for analysing the most important classes of antimicrobials in various biological matrices (milk, animal tissues, eggs, and honey). As evidenced by this review, reversed-phase LC combined with tandem MS, usually triple-quadrupole MS (QqQMS), is currently the preferred technique in most residue analysis of a single-class of antimicrobials. A recently emerging analytical strategy is that of developing methods for detecting a large variety of veterinary drugs belonging to different classes, including pesticides (multi-class residue analysis). To do this, simple and generic extraction and separation techniques applicable to a broad range of compounds differing in physical and chemical properties have been adopted. Such methods are still based mainly on LC-QqQMS. Emerging alternative MS detection systems are time-of-flight MS, which provides accurate mass of the analyte(s), or Q-linear ion trap (IT) MS that eliminates some limitations of ITMS(n).

Use of Antimicrobial Agents in United States Neonatal and Pediatric Intensive Care Patients

Antimicrobial use contributes to the development of emergence and dissemination of antimicrobial-resistant bacteria among intensive care unit (ICU) patients. There are few published data on antimicrobial use in neonatal (NICU) and pediatric ICU (PICU) patients. Personnel at 31 Pediatric Prevention Network hospitals participated in point prevalence surveys on August 4, 1999 (summer) and February 8, 2000 (winter). Data collected for all NICU and PICU inpatients included demographics, antimicrobials and indications for use and therapeutic interventions. Data were reported for 2647 patients in 29 NICUs (827 patients in summer; 753 in winter) and 35 PICUs (512 patients in summer; 555 in winter). PICU patients were more likely than NICU patients to be receiving antimicrobials on the survey date [758 of 1070 (70.8%) versus 684 of 1582 (43.2%), P < 0.0001]. NICU patients were receiving a higher median number of antimicrobials (2 versus 1, P < 0.0001). The most common agents among NICU patients were gentamicin, ampicillin and vancomycin; the most common agents among PICU patients were cefazolin, vancomycin and cefotaxime. Use of aminoglycosides, aminopenicillins and topical antibacterials was significantly more common in NICU patients; first, second and third generation cephalosporins, extended spectrum penicillins, sulfonamides, fluoroquinolones, antianaerobic agents, systemic antifungals and systemic antivirals were more common in PICU patients. This is the first U.S. national multicenter description of antimicrobial use in NICUs and PICUs and demonstrates the high prevalence of antimicrobial use among these patients. Assessment strategies targeting antimicrobial use in pediatrics are needed.