Carlos Alonso-Calleja

Antibiotic-Resistant Bacteria: A Challenge for the Food Industry

Antibiotic-resistant bacteria were first described in the 1940s, but whereas new antibiotics were being discovered at a steady rate, the consequences of this phenomenon were slow to be appreciated. At present, the paucity of new antimicrobials coming into the market has led to the problem of antibiotic resistance fast escalating into a global health crisis. Although the selective pressure exerted by the use of antibiotics (particularly overuse or misuse) has been deemed the major factor in the emergence of bacterial resistance to these antimicrobials, concerns about the role of the food industry have been growing in recent years and have been raised at both national and international levels. The selective pressure exerted by the use of antibiotics (primary production) and biocides (e.g., disinfectants, food and feed preservatives, or decontaminants) is the main driving force behind the selection and spread of antimicrobial resistance throughout the food chain. Genetically modified (GM) crops with antibiotic resistance marker genes, microorganisms added intentionally to the food chain (probiotic or technological) with potentially transferable antimicrobial resistance genes, and food processing technologies used at sub-lethal doses (e.g., alternative non-thermal treatments) are also issues for concern. This paper presents the main trends in antibiotic resistance and antibiotic development in recent decades, as well as their economic and health consequences, current knowledge concerning the generation, dissemination, and mechanisms of antibacterial resistance, progress to date on the possible routes for emergence of resistance throughout the food chain and the role of foods as a vehicle for antibiotic-resistant bacteria. The main approaches to prevention and control of the development, selection, and spread of antibacterial resistance in the food industry are also addressed.

Microbiological quality of retail chicken by-products in Spain

The microbiological quality of retail chicken parts (legs, wings and giblets) and processed chicken products (hamburgers and sausages) in Spain was investigated. Mean counts (log(10) cfu/g) ranged from 5.56 to 7.28, 5.96 to 7.87, 3.49 to 5.42, 2.60 to 4.33 and 2.47 to 3.48 for mesophiles, psychrotrophs, coliforms, Escherichia coli and Staphylococcus aureus, respectively. Chicken parts were generally regarded as being of unacceptable quality, since psychrotrophs, E. coli and S. aureus counts were higher than the maximum limits established in the guidelines for poultry meat. On the basis of Spanish Microbiological Standards, 80% of the samples of hamburgers and sausages were also regarded as being of unacceptable quality. The main reason for the lack of acceptability was excessive counts of mesophiles. The results in this study indicate that chicken by-products (especially those made with ground meat) could represent notable hazards to humans and are a cause of public health concern.

Occurrence of Listeria species in retail poultry meat and comparison of a cultural/immunoassay for their detection

The prevalence of Listeria spp. on the skin of a hundred fresh chicken carcasses purchased from 20 retail stores in León was investigated using the routine test procedure recommended by the United States Department of Agriculture (USDA). PALCAM and Modified Oxford agar (MOX) were used for isolation. Listeria spp., Listeria monocytogenes, L. innocua, L. welshimeri, L. grayi and L. ivanovii were present in 95%, 32%, 66%, 7%, 4% and 2% of the samples, respectively. Next, an immunoassay test (Listeria Rapid Test; Oxoid, Unipath) and the routine test procedure (USDA) were compared for their ability to detect Listeria spp. on 40 chicken carcasses. When we used MOX for the isolations, the sensitivity of the immunoassay test was 100% and the specificity 85.7%. When we used PALCAM medium the sensitivity and specificity of the test was 94.29% and 80%, respectively.

Prevalence and antimicrobial resistance of Salmonella serotypes isolated from poultry in Spain: comparison between 1993 and 2006.

A total of 226 chicken samples (carcasses, legs, wings, necks and breasts) were obtained (73 in 1993 and 153 in 2006) from 10 retail outlets in North-Western Spain and screened for the presence of Salmonella. Isolates were subjected to serotyping, phage typing (Salmonella Enteritidis and Salmonella Typhimurium) and antimicrobial susceptibility testing (15 antimicrobials; disk diffusion method). Salmonella was detected in 40 (55%) samples in 1993 and 19 (12.4%) in 2006 (P<0.001). The serotypes (S. Enteritidis, Salmonella Poona, Salmonella Infantis, Salmonella Newport and S. Typhimurium) and phage types (1, 4, 14b and 35 in the case of S. Enteritidis and 193 for S. Typhimurium) detected are among the main types responsible for human salmonellosis in Spain. All strains were multi-resistant (resistant to 3-13 antimicrobials). The average number of resistances per strain increased (P<0.05) from 3.98 in 1993 to 5.00 in 2006. An increase in the incidence of resistance was observed between 1993 and 2006 for cephalothin (P<0.01), enrofloxacin (P<0.001) and tetracycline (P<0.01). The decreases in the prevalence of Salmonella between 1993 and 2006 suggest that the mandatory measures introduced over the last decade in the European Union to reduce the incidence of Salmonella in poultry have apparently been successful. However, the increase in antibiotic resistance rates is of concern and constitutes a threat to public health. Because the data in this study demonstrated that chicken in North-Western Spain is a potential source of antibiotic-resistant Salmonella strains, the need of consumer education on good sanitary practices is highlighted.

Sampling Methods for Microbiological Analysis of Red Meat and Poultry Carcasses

Microbiological analysis of carcasses at slaughterhouses is required in the European Union for evaluating the hygienic performance of carcass production processes as required for effective hazard analysis critical control point implementation. The European Union microbial performance standards refer exclusively to the excision method, even though swabbing using the wet/dry technique is also permitted when correlation between both destructive and nondestructive methods can be established. For practical and economic reasons, the swab technique is the most extensively used carcass surface-sampling method. The main characteristics, advantages, and limitations of the common excision and swabbing methods are described here.

Occurrence of salmonellae in retail chicken carcasses and their products in Spain.

This study examined the incidence of Salmonella in Spanish poultry products. Samples included chicken carcasses, chicken parts (wings, legs and giblets-livers and hearts) and processed chicken products (red sausages, white sausages and hamburgers). The average detection rate was 49%, with the highest (55%) in chicken carcasses (skin) and the lowest (20%) in hamburgers. The chicken carcasses purchased in supermarkets were more contaminated (75%) than those from poulterers shops (25%). Salmonella Enteritidis, S. Poona, S. Paratyphi B and S. Worthington were isolated in 34.3%, 11.4%, 2.8% and 1.4% of the samples, respectively. One (1.4%) red sausage sample harboured two serotypes (S. Enteritidis and S. Worthington). This fact emphasizes the usefulness of subtyping several Salmonella isolates from the same sample in epidemiological studies.

Discrimination of Enterobacterial Repetitive Intergenic Consensus PCR Types of Campylobacter coli and Campylobacter jejuni by Fourier Transform Infrared Spectroscopy

ABSTRACT Fourier transform infrared spectroscopy (FT-IR) has been used together with pattern recognition methodology to study isolates belonging to the species Campylobacter coli and Campylobacter jejuni and to compare FT-IR typing schemes with established genomic profiles based on enterobacterial repetitive intergenic consensus PCR (ERIC-PCR). Seventeen isolates were cultivated under standardized conditions for 2, 3, and 4 days to study variability and improve reproducibility. ERIC-PCR profiles and FT-IR spectra were obtained from strains belonging to the species Campylobacter coli and C. jejuni, normalized, and explored by hierarchical clustering and stepwise discriminant analysis. Strains could be differentiated by using mainly the first-derivative FT-IR spectral range, 1,200 to 900 cm−1 (described as the carbohydrate region). The reproducibility index varied depending on the ages of the cultures and on the spectral ranges investigated. Classification obtained by FT-IR spectroscopy provided valuable taxonomic information and was mostly in agreement with data from the genotypic method, ERIC-PCR. The classification functions obtained from the discriminant analysis allowed the identification of 98.72% of isolates from the validation set. FT-IR can serve as a valuable tool in the classification, identification, and typing of thermophilic Campylobacter isolates, and a number of types can be differentiated by means of FT-IR spectroscopy.

Review: Trisodium Phosphate (TSP) Treatment for Decontamination of Poultry

Use of trisodium phosphate (TSP) treatment for reducing levels of bacteria in poultry is discussed with reference to: health and economic consequences of poultry contamination, causes or routes of contamination, possibilities for reduction of microbial loads, mechanisms of action of TSP, sensory properties and quality of TSP-treated poultry, antimicrobial effectiveness, influences on shelf-life, and worldwide authorization of this process, with special reference to the situation in the European Union. A summary of the main results of microbial reductions on poultry following TSP treatment is shown for Salmonella, coliforms/Escherichia coli, Enterobacteriaceae, Campylobacter, Pseudomonas, total counts, Listeria, Staphylococcus aureus and Lactobacillus. The main results on microbial reductions assessed in foodstuffs other than poultry (beef, fruit, fish and shellfish) are also shown. En este artículo se discute el uso del tratamiento con fosfato trisódico (TSP) para reducir la contaminación bacteriana de la carne de ave con referencia a: consecuencias económicas y para la salud de la contaminación de la carne de ave, causas o rutas de contaminación, posibilidades de reducir la carga microbiana, mecanismos de acción del TSP, características organolépticas y calidad de la carne de ave tratada con TSP, eficacia antimicrobiana y factores que la afectan, influencia de este tratamiento en el tiempo de vida útil, y autorización de este proceso en el mundo, con especial referencia a la situación dentro de la Unión Europea. Se han resumido los resultados de las reducciones microbianas obtenidas en carne de ave tratada con TSP para Salmonella, coliformes/Escherichia coli, Enterobacteriaceae, Campylobacter, Pseudomonas, microorganismos totales, Listeria, Staphylococcus aureus y Lactobacillus. También se han contemplado los principales resultados obtenidos en otros alimentos distintos de la carne de ave (carne de vacuno, fruta, pescado y marisco).

Effectiveness of trisodium phosphate, acidified sodium chlorite, citric acid, and peroxyacids against pathogenic bacteria on poultry during refrigerated storage.

The effects of dipping treatments (15 min) in potable water or in solutions (wt/vol) of 12% trisodium phosphate (TSP), 1,200 ppm acidified sodium chlorite (ASC), 2% citric acid (CA), and 220 ppm peroxyacids (PA) on inoculated pathogenic bacteria (Listeria monocytogenes, Staphylococcus aureus, Bacillus cereus, Salmonella Enteritidis, Escherichia coli, and Yersinia enterocolitica) and skin pH were investigated throughout storage of chicken legs (days 0, 1, 3, and 5) at 3 +/- 1 degrees C. All chemical solutions reduced microbial populations (P < 0.001) as compared with the control (untreated) samples. Similar bacterial loads (P > 0.05) were observed on water-dipped and control legs. Type of treatment, microbial group, and sampling day influenced microbial counts (P < 0.001). Average reductions with regard to control samples were 0.28 to 2.41 log CFU/g with TSP, 0.33 to 3.15 log CFU/g with ASC, 0.82 to 1.97 log CFU/g with CA, and 0.07 to 0.96 log CFU/g with PA. Average reductions were lower (P < 0.001) for gram-positive (0.96 log CFU/g) than for gram-negative (1.33 log CFU/g) bacteria. CA and ASC were the most effective antimicrobial compounds against gram-positive and gram-negative bacteria, respectively. TSP was the second most effective compound for both bacterial groups. Average microbial reductions per gram of skin were 0.87 log CFU/g with TSP, 0.86 log CFU/g with ASC, 1.39 log CFU/g with CA, and 0.74 log CFU/g with PA for gram-positive bacteria, and 1.28 log CFU/g with TSP, 2.03 log CFU/g with ASC, 1.23 log CFU/g with CA, and 0.78 log CFU/g with PA for gram-negative bacteria. With only a few exceptions, microbial reductions in TSP- and ASC-treated samples decreased and those in samples treated with CA increased throughout storage. Samples treated with TSP and samples dipped in CA and ASC had the highest and lowest pH values, respectively, after treatment. The pH of the treated legs tended to return to normal (6.3 to 6.6) during storage. However, at the end of storage, the pH of legs treated with TSP remained higher and that of legs treated with CA remained lower than normal.

Prevalence of Salmonella enterica serovars and genovars from chicken carcasses in slaughterhouses in Spain

Aims: To determine the prevalence of Salmonella enterica serovars in chicken carcasses in slaughterhouses in Spain and to examine genotypic relations among these serovars.