Diversity and epidemiological profile of Pseudomonas aeruginosa from drinking water in Brazil genotyped using multi-locus sequence typing.

Pseudomonas aeruginosa is a Gram-negative bacillus associated with waterborne diseases. The objective of this study was to determine whether particular P. aeruginosa sequence types (STs) were associated with drinking water contamination in Brazil. This was achieved by searching the Pseudomonas PubMLST database, which contains the records for 8358 strains collected between 1938 and 2023. The majority (97.2%) had the complete 7-loci multilocus sequence typing profile and were assigned to 3486 STs. After eBURST (an algorithm used to infer patterns of evolutionary descent among clusters), 1219 groups with single-locus variant and 575 groups with double-locus variant were formed. Brazil was the South American country with the most isolates (n = 219, 58.24%), and the Simpson's index was 0.9392. Of the 219 Brazilian isolates, eight were isolated in water and identified as STs 252, 1417, 1605, 2502, 2620, 3078, and 3312. ST252, 1417, and 3078 have already been isolated from clinical cases worldwide. Furthermore, ST1605 and 2620, after the eBURST, they were grouped in the same clonal complex as STs involved in human infections. In conclusion, P. aeruginosa STs involved in human infections were found in bottled drinking water commercialized in Brazil, revealing that these types of drinking waters can be a vehicle of contamination.

Tolerance of foodborne Acinetobacter spp. to sanitizer agents.

INTRODUCTION: The bacteria Acinetobacter spp. are extremely relevant in clinical settings. Recently, they have emerged as potential food-borne opportunistic pathogens. Their ability to form biofilms contributes to antibiotic resistance by generating an environment that facilitates the acquisition and transfer of resistance genes. Studies on the tolerance of Acinetobacter spp. from food sources to sanitizers used in the food industry and homes are necessary to help mitigate food contamination by these microorganisms. METHODOLOGY: Isolates from ready-to-eat salads (n = 11) and raw goat milk (n = 4) were evaluated for their tolerance to sodium hypochlorite (NaClO), quaternary ammonium compound/biguanide (QAC/BG), and peracetic acid (PAA). The Food and Drug Administration (FDA) recommends that the concentration of these sanitizers in food-processing equipment and utensils and other food-contact articles should not exceed 200 parts per million (ppm). RESULTS: The minimum inhibitory (MIC) and bactericidal (MBC) concentrations of NaClO were above 312.5 ppm for all isolates tested and ≥ 2,500 ppm for four isolates from salads. Only three isolates from salads and four isolates from goat milk were inhibited by an MIC lower than 200 ppm of PAA. QAC/BG presented the lowest MIC and MCB values (9.37/6.25  ppm for all isolates tested), suggesting that it is the most effective agent against the isolates used in this study. CONCLUSIONS: Our results demonstrate that Acinetobacter spp. isolates from food can be tolerant to the recommended concentrations of NaClO and PAA, which highlights the health risks to consumers.

Antibiotic resistance profile and detection of degradative enzymes by Enterobacteriaceae isolated from raw goat milk.

INTRODUCTION: Enterobacteriaceae are often reported as a typical bacterial population in raw milk from any mammalian origin. The frequent concern with bacteria, especially those related to this group of microorganisms, is their increasing resistance to antibiotics and the emergence of enzymes that degrade them. This study aimed to characterize isolates of Enterobacteriaceae from raw goat milk to expose associated safety problems and possible technological challenges. METHODS: Isolates from 21 raw goat milk samples purchased in the State of Rio de Janeiro, Brazil, were identified by mass spectrometry, after isolation on Violet Red Bile Glucose agar. The isolates were subjected to evaluation of proteolytic, lipolytic, hemolytic, and biofilm producing activities. Furthermore, resistance profiles and production capacity of enzymes that degrade antimicrobials were evaluated. RESULTS: Almost half of the 59 isolates (48%) belonged to the Enterobacter genus, with a significant prevalence of the Serratia (20%) and Klebsiella (11%) genera. The majority showed biofilm-producing activity (90%), while the activity of degradative enzymes was observed in approximately 20%. Few isolates were found with a profile of resistance to antimicrobials, with only one isolate of Klebsiella variicola being classified as multidrug-resistant. However, chromogenic culture media showed high production of extended-spectrum beta-lactamases and carbapenemases (54% and 46%, respectively), as a presumptive identification. CONCLUSIONS: A considerable degree of virulence was observed in the Enterobacteriaceae isolates, as well as the potential for undesirable technological damage. The characterization and identification of the isolates contributes to the improvement of the risk monitoring process of goat's milk.

From food to hospital: we need to talk about Acinetobacter spp.

Some species of the genus Acinetobacter are admittedly important hospital pathogens. Additionally, various animal and plant foods have been linked to the presence of Acinetobacter, including resistant strains. However, due to isolation difficulties and the lack of official standard methods, there is a dearth of work and epidemiological data on foodborne diseases caused by this microorganism. Considering that Acinetobacter spp. may represent a serious public health problem, especially because of their resistance to carbapenems and colistin, and because of the fact that these pathogens may transfer resistance genes to other bacteria, studies are needed to evaluate the pathogenicity of both food and clinical isolates and to search for them using control strategies, such as the adoption of more efficient disinfection measures and use of antimicrobial substances (AMS). In contrast, AMS production by strains of the genus Acinetobacter has already been described, and its potential for application against other Gram-negative food or clinical pathogens, reveals a new field to be explored.

Characterization of Acinetobacter spp. from raw goat Milk / Caracterização de Acinetobacter spp. isolados de leite caprino cru

ABSTRACT: Goat's milk has been suggested as an alternative to cow's milk, being a better digestible and hypoallergenic option. However, the presence of contaminating bacteria may significantly affect the safety of the product. In this research, we reported the isolation and characterization of Acinetobacter spp. isolates from raw goat milk samples purchased in the state of Rio de Janeiro, Brazil. Twenty-one samples were analyzed and ten isolates of Acinetobacter spp. were obtained. Six were identified as A. guillouiae, three as A. ursingii, and one as A. bereziniae. These isolates were characterized and eight showed proteolytic activity, seven showed lipolytic activity, and five isolates were able to produce both enzymes. None of the isolates was biofilm producer. However, when the production of antibiotic resistance enzymes KPC and ESBL were investigated, all isolates presented ESBL-positive phenotype, while eight (80%) were KPC-positive. This research, therefore, demonstrated that raw goat's milk can also be a source of Acinetobacter spp., which can produce important thermostable deteriorating enzymes and may play a role of reservoirs of antibiotic resistance genes.

RESUMO: O leite de cabra tem sido sugerido como alternativa ao leite de vaca, sendo uma opção com melhor digestibilidade e menor alergenicidade. No entanto, a presença de bactérias contaminantes pode afetar significativamente a segurança do produto. Neste trabalho, relatamos o isolamento e caracterização de Acinetobacter spp. isolados de amostras de leite cru de cabra adquiridas no Estado do Rio de Janeiro, Brasil. Vinte e uma amostras foram analisadas e dez isolados de Acinetobacter spp. foram obtidos, sendo seis identificados como A. guillouiae, três como A. ursingii e um como A. bereziniae. Estes isolados foram caracterizados e oito apresentaram atividade proteolítica, sete mostraram atividade lipolítica e cinco foram capazes de produzir ambas as enzimas. Nenhum dos isolados foi produtor de biofilme. No entanto, todos os isolados apresentaram fenótipo ESBL-positivo, enquanto oito (80%) foram positivos para KPC. Este trabalho demonstra, portanto, que o leite de cabra cru também pode ser uma fonte de Acinetobacter spp., que podem produzir enzimas deteriorantes termoestáveis e desempenhar, também, um papel de reservatórios de genes de resistência a antibióticos.

Pathogen killing pathogen: antimicrobial substance from Acinetobacter active against foodborne pathogens.

INTRODUCTION: Antimicrobial substances (AMS) produced by bacteria may reduce or prevent the growth of pathogenic and spoilage microorganisms in food. In this study, 16 isolates of Acinetobacter baumannii/calcoaceticus (ABC) complex, previously obtained from reconstituted infant milk formula (IMF) samples and the preparation and distribution utensils from the nursery of a public hospital, were used to screen for AMS production. METHODOLOGY: Antimicrobial substance production and spectrum of activity assays were performed by agar-spot assay. Optimization of growth conditions for AMS production was also evaluated. RESULTS: Three (17.6%) isolates, namely JE3, JE4, and JE6, produced AMS against the principal indicator strain Salmonella enterica subsp. enterica serotype Typhi ATCC 19214. JE6 was also able to inhibit strains of Klebsiella pneumoniae, Proteus vulgaris, and Bacillus cereus, a Gram-positive bacteria. Remarkably, JE6 was able to inhibit all the tested resistant and multidrug-resistant (MDR) strains of the ABC complex and Shigella dysenteriae associated with IMF and utensils, indicating a potentially valuable application. AMS produced by JE6 does not appear to be affected by proteolytic enzymes and the producer strain showed specific immunity to its own AMS. CONCLUSION: This study highlights AMS produced by Acinetobacter with applications against MDR spoilage and foodborne pathogens - some of them, infectious disease causing agents - which, to our knowledge, has not been previously described.

First Report of Human Gastroenteritis Caused by Escherichia coli O157:NM in Brazil.

In September 2005, the Sanitary Surveillance Service of Rio de Janeiro (SSS/RJ), Brazil, investigated a case of gastroenteritis involving a 13-year-old teenager hospitalized because of bloody diarrhea and severe abdominal pain. Owing to the severity of the symptoms, an epidemiological investigation was conducted in two states of Brazil. Escherichia coli O157:NM was isolated from stools and from a tomato and cheese salad prepared at the school canteen where the teenager attended. This is the first report of a human case of gastroenteritis related to E. coli O157:NM infection in Brazil.

Acinetobacter: an underrated foodborne pathogen?

The increasing prevalence of foodborne diseases observed in developing countries has been linked to a rise in the consumption of raw foods. However, unlike the classical pathogens that are commonly implicated in foodborne illnesses, members of the genus Acinetobacter are rarely associated with diarrheal disease, probably because of the difficulty in isolating these Gram-negative bacteria from food sources. Nevertheless, several species of Acinetobacter, especially A. baumannii, possess many of the characteristics associated with successful pathogens and exhibit a prodigious ability to acquire the multiple-drug resistance (MDR) phenotype. In this mini-review, we summarize the epidemiological data relating to MDR Acinetobacter and consider evidence suggesting that contaminated dairy products, along with raw fruit and vegetables, constitute extra-hospital reservoirs of this underrated pathogen, and may represent an increased risk to immunocompromised individuals and young children in healthcare settings.

Avaliação do crescimento de Salmonella enterica em fórmulas lácteas infantis sob diferentes condições de preparo e armazenamento / Evaluation of the growth of Salmonella enterica in infant milk formulas under different preparation and storage conditions

Salmonella enterica é um dos mais importantes patógenos associados a fórmulas lácteas infantis (FLI). Neste trabalho, foi avaliada a sobrevivência e crescimento de uma estirpe padrão de Salmonella enterica sorotipo Typhi em duas FLI reconstituídas (F1 e F2) sob diferentes condições de preparo e armazenamento similares às utilizadas em lactários hospitalares. Para ambas as FLI inoculadas e incubadas à temperatura ambiente, observou-se um aumento de 3,3 log na população de S. enterica logo nas primeiras 24 horas de incubação e um aumento superior a 4,5 log ao término de 72 horas. A 4ºC, nas primeiras 48 h de incubação, houve leve aumento populacional (< 1,0 log) e, após 72 horas, o crescimento foi similar ao observado sob temperatura ambiente. Em relação ao aquecimento em banho-maria, verificou-se que a 60ºC/ 5 min houve uma redução de um pouco mais de 1 log UFC/ml, entretanto, a 60ºC/ 10 min, a queda observada foi de 2,8 log em F1 e de 2,3 log em F2. Já a 70ºC/ 5 min, ocorreu redução de cerca de 3 log UFC/ml para F1 e F2, enquanto que por 10 min, essa redução foi cerca de 1 log maior. O aquecimento em forno de micro-ondas mostrou ser a forma mais rápida e eficiente de redução da população de Salmonella nas FLI, uma vez que não foi detectada contagem celular após nenhum dos tratamentos utilizados. Os resultados sugerem que FLI contaminadas durante a etapa de preparo podem apresentar um crescimento bacteriano mesmo sob temperatura de refrigeração se mantidos por tempo prolongado e que determinados métodos de tratamento térmico não são suficientes para inibição completa de S. enterica.

Salmonella enterica is one of the most important pathogens associated with infant milk formulas (IMFs). In this study, the survival and growth of an S. enterica serovar Typhi strain in two reconstituted IMFs (F1 and F2) under different conditions of preparation and storage, similar to those used in hospital lactaries, was evaluated. For both IMFs inoculated at room temperature, there was 3.3 log increase in the population of S. enterica by the first 24 h of incubation and > 4.5 log increase at the end of 72 h. At 4ºC, in the first 48 h of incubation, there was a slight increase in population (< 1.0 log) and after 72 h, growth was similar to that observed at room temperature. A heated water bath was used to test the effect of thermal treatment methods on bacterial viability. At 60ºC for 5 min, there was a reduction of slightly more than 1 log CFU/mL; however, at 60ºC for 10 min, the observed decrease was 2.8 log for F1 and 2.3 log for F2. At 70ºC for 5 min, there was a reduction of approximately 3 log CFU/ml for both IMFs and for 10 min, this reduction was approximately 1 log higher. Heating in a microwave oven was the most efficient way of reducing populations of Salmonella in the IMFs, as bacterial cell counts were not detected after any of the treatments used. Our results suggest that IMF contamination during the preparation step may support bacterial growth even under refrigeration if kept for a prolonged time and that some thermal treatment methods are not sufficient for the complete inhibition of S. enterica.

Antibiotic resistance versus antimicrobial substances production by gram-negative foodborne pathogens isolated from minas frescal cheese: heads or tails?

In this study, 15 Gram-negative isolates from Minas Frescal cheese sold in commercial establishments in Rio de Janeiro, Brazil, were able to produce antimicrobial substances (AMSs). Seven, four, two, one, and one isolates identified as Yersinia, Acinetobacter, Enterobacter, Escherichia, and Hafnia genera, respectively, were considered potentially pathogenic. All 15 AMS(+) isolates were resistant to at least 1 antibiotic; however, 7 strains presented resistance to at least 3 antibiotics from different classes, exhibiting multiresistance profiles. The strains were also subjected to plasmid profile analysis. All isolates presented different plasmid forms with most ranging in size from 1 to 10 kb. Activity against various pathogens associated with food was tested and all 15 AMS(+) showed the same activity spectrum, inhibiting all Escherichia coli and Salmonella strains that were tested. Although restricted, the action spectrum of AMS-producing strains is extremely relevant to the food industry because Gram-negative bacteria such as E. coli and Salmonella spp. are most often associated with foodborne illnesses. The findings of this study reveal that even AMS produced by pathogens can have potential applications against other foodborne pathogens.

Desenvolvimento de material de referência para microbiologia de alimentos contendo Listeria monocytogenes em matriz queijo / Development of reference material for food microbiology containing Listeria monocytogenes in matrix

A NBR ISO/IEC 17025:2005, uma das normas mais utilizadas em laboratórios de ensaio, descreve como um de seus critérios para a garantia da qualidade dos resultados analíticos a participação periódica dos laboratórios em ensaios de proficiência (EP). Os analitos, utilizados nos EP são materiais de referência (MR) provenientes de um mesmo lote, e devem apresentar características de homogeneidade e estabilidade. O objetivo deste estudo foi produzir um MR qualitativo destinado ao ensaio de pesquisa de Listeria monocytogenes em matriz queijo pela técnica de liofilização. Para a produção do MR, foi utilizado como matriz o queijo Minas frescal (QMF) ultrafiltrado. A matriz foi distribuída em frascos, contaminada com a bactéria alvo e submetida à liofilização, tendo a sacarose como crioprotetor. O MR produzido foi considerado homogêneo e estável na temperatura de ≤-70°C durante todo o período estudado (10 meses). O material apresentou estabilidade a 4, 25, 30 e 35°C por quatro dias e a -20°C por 48 dias, e os resultados estatísticos indicam tendência à estabilidade. Conclui-se que o material apresentou todos os requisitos necessários de um MR de qualidade e poderia ser transportado aos laboratórios participantes de um EP à temperatura máxima de 35°C por até quatro dias, uma vez que os resultados indicaram a manutenção da concentração celular nesse período. Esse foi o primeiro trabalho a descrever uma metodologia de produção de MR contendo L. monocytogenes em matriz queijo.

The standard most used in testing laboratories, ISO/IEC 17025:2005, describes the participation of laboratories in periodic proficiency testing (PT), as a criteria for quality assurance of analytical results. The analyses used in PT are reference materials (MR) from the same lot, and must have characteristics of homogeneity and stability. This study aimed to produce a qualitative RM for detection of Listeria monocytogenes assays in cheese matrix. For the production of RM, Minas Frescal cheese (MFQ) was used as matrix. The matrix was distributed in flasks, contaminated with the target bacteria and submitted to freeze-drying. Sucrose was used as cryoprotector. The RM produced was considered homogeneous and stable at -70°C during the entire period of study (10 months). The material showed stability at 4, 25, 30 and 35°C for 4 days and at -20°C the RM showed stability for 48 days, and the statistical results indicate a tendency to maintain stability. It was concluded that the material showed all the requirements of an RM quality and could be transported to the laboratory participants of a PT at 35°C up to 4 days, since the results indicate the maintenance of cell concentration during this period. This is the first study to describe a methodology for producing MR containing L. monocytogenes in cheese matrix.

Genes involved in immunity to and secretion of aureocin A53, an atypical class II bacteriocin produced by Staphylococcus aureus A53.

Aureocin A53 is an antimicrobial peptide produced by Staphylococcus aureus A53. The genetic determinants involved in aureocin A53 production and immunity to its action are organized in at least four transcriptional units encoded by the 10.4-kb plasmid pRJ9. One transcriptional unit carries only the bacteriocin structural gene, aucA. No immunity gene is found downstream of aucA, as part of the same transcriptional unit. Further downstream of aucA is found an operon which contains the three genes aucEFG, whose products seem to associate to form a dedicated ABC transporter. When aucEFG were expressed in RN4220, an aureocin A53-sensitive S. aureus strain, this strain became partially resistant to the bacteriocin. A gene disruption mutant in aucE was defective in aureocin A53 externalization and more sensitive to aureocin A53 than the wild-type strain, showing that aucEFG are involved in immunity to aureocin A53 by active extrusion of the bacteriocin. Full resistance to aureocin A53 was exhibited by transformants carrying, besides aucEFG, the operon formed by two genes, aucIB and aucIA, located between aucA and aucEFG and carried in the opposite strand. AucIA and AucIB share similarities with hypothetical proteins not found in the gene clusters of other bacteriocins. A gene disruption mutant in orf8, located upstream of aucA and whose product exhibits about 50% similarity to a number of hypothetical membrane proteins found in many Gram-positive bacteria, was strongly affected in aureocin A53 externalization but resistant to aureocin A53, suggesting that Orf8 is also involved in aureocin A53 secretion.

Substâncias antimicrobianas produzidas por bacillus spp. isolados de frutas / Antimicrobial substances produced by bacillus spp. isolated from fruit

O objetivo deste trabalho consistiu na investigação da produção de substâncias antimicrobianas por estirpes de Bacillus, isoladas de frutas, e verificar o potencial dessas substâncias na inibição de patógenos, associados à deterioração ou à transmissão de doenças por alimentos, como fungos e bactérias Gram-negativas ou Gram-positivas. Dez estirpes submetidas à coloração de Gram que mostraram ser bacilos foram testadas quanto à produção de substâncias antimicrobianas, utilizando-se como indicadoras diferentes bactérias Gram-positivas e Gram-negativas. Três estirpes, denominadas de Ame3, Mam1 e Pes1 apresentaram os maiores espectros de ação contra bactérias Gram-positivas, sugerindo que as substâncias produzidas por essas estirpes possam apresentar potencial de aplicação como biopreservativo de alimentos.Dez estirpes submetidas à coloração de Gram que mostraram ser bacilos foram testadas quanto à produção de substâncias antimicrobianas, utilizando-se como indicadoras diferentes bactérias Gram-positivas e Gram-negativas. Três estirpes, denominadas de Ame3, Mam1 e Pes1 apresentaram os maiores espectros de ação contra bactérias Gram-positivas, sugerindo que as substâncias produzidas por essas estirpes possam apresentar potencial de aplicação como biopreservativo de alimentos. Dez estirpes submetidas à coloração de Gram que mostraram ser bacilos foram testadas quanto à produção de substâncias antimicrobianas, utilizando-se como indicadoras diferentes bactérias Gram-positivas e Gram-negativas. Três estirpes, denominadas de Ame3, Mam1 e Pes1 apresentaram os maiores espectros de ação contra bactérias Gram-positivas, sugerindo que as substâncias produzidas por essas estirpes possam apresentar potencial de aplicação como biopreservativo de alimentos.

Nukacin 3299, a lantibiotic produced by Staphylococcus simulans 3299 identical to nukacin ISK-1.

Nukacin 3299 (formerly designated simulancin 3299), produced by a Staphylococcus simulans strain involved in bovine mastitis in Brazil, is the first peptide bacteriocin described in this staphylococcal species. With the intent to elucidate some aspects of its biology, nukacin 3299 was purified and characterized. The mass of the purified bacteriocin was shown to be 2957.3 Da, and the peptide N-terminal amino acids (KKKSGVI) were identified by Edman degradation. The nukacin 3299 structural gene, nukA, was detected by PCR and DNA sequencing, showing that this bacteriocin is identical to nukacin ISK-1, a 27-amino acid type-A (II) lantibiotic produced by Staphylococcus warneri ISK-1, isolated from a "nukadoko", in Japan. The genes involved in nukacin 3299 biosynthesis are located on plasmid pRJ97 (>27 kb). They have an organization similar to that of the nukacin ISK-1 gene cluster, excepted for the presence of an IS257/431 element (791 bp) present between the orf1 and nukA genes of the nukacin 3299 gene cluster. The presence of this insertion sequence is expected to affect the expression of orf1, whose function is presently unknown. Nukacin 3299 proved to be sensitive to proteolytic enzymes and relatively stable at different temperatures and between pH 3.0-9.0. Nukacin 3299 exhibited activity towards staphylococcal strains involved in bovine mastitis, showing a potential application on mastitis control, a disease with great economic impact.

Aureocins 4185, bacteriocins produced by Staphylococcus aureus 4185: potential application in food preservation.

In the present study, the bacteriocins produced by Staphylococcus aureus 4185, a strain isolated from bovine mastitis, were purified and partially characterized. After purification by ammonium sulfate precipitation, cation-exchange chromatography, and five runs of high-performance liquid chromatography (HPLC), antimicrobial activity was recovered with 40% and 80% isopropanol, suggesting that more than one antimicrobial peptide, named aureocins 4185, is produced by S. aureus 4185. Mass spectrometry analyses revealed three peptides eluted with 40% isopropanol: peptide A (2,305.3 +/-1.5 Da), peptide B (2,327.3 +/-1.5 Da), and peptide C (3,005.5 +/-1.5 Da), and two peptides eluted with 80% isopropanol: peptide D (6,413.5 +/-1.5 Da) and peptide E (12,834.5 +/-1.5 Da). Although five peptides have been detected, only four small peptide sequences were obtained by matrix-assisted laser desorption/ionization time of flight (MALDI-TOF)/TOF mass spectrometry analyses: SLLEQFTGK (eluted with 40% isopropanol), ALLYDER, NNTSHNLPLGWFNVK, and NNLAQGTFNATK (eluted with 80% isopropanol). The sequences SLLEQFTGK and ALLYDER revealed identity with hypothetical peptides with unknown function. The sequences NNTSHNLPLGWFNVK and NNLAQGTFNATK showed similarity to a segment of a precursor of staphylococcal autolysins. The antimicrobial activity detected in the supernatant of strain 4185 proved to be resistant to heat treatment at 65°C; however, treatment at 80°C abolished completely its antimicrobial properties. The concentrated supernatant containing aureocins 4185 exhibited a strong bacteriolytic activity toward Micrococcus luteus ATCC 4698. Additionally, aureocins 4185 exhibited antagonistic activity against important foodborne pathogens, including Listeria monocytogenes, thus showing a potential application in food preservation.

Antimicrobial substances produced by coliform strains active against foodborne pathogens.

In the present study, 31 coliform strains were isolated from salad, cheese, and meat products sold in commercial establishments in Rio de Janeiro city, and were tested for antibiotic resistance and antimicrobial substance production. Thirteen strains (41.9%) were resistant to at least one antibiotic tested, among which one presented resistance to nine different antibiotics. Two strains (6.4%) exhibited inhibitory activity against the indicator strains, Escherichia coli LMIFRJ and Salmonella enterica I. The antimicrobial substances that they produced were sensitive to proteolytic enzymes, suggesting that they might be bacteriocins. The producer strains were identified as Klebsiella ozaenae and Raoultella terrigena. Although they had similar spectrums of action, the bacteriocins were shown to be different. Both of them were able to inhibit E. coli, Klebsiella, Enterobacter, and Salmonella strains, including antibiotic-resistant ones. Our results suggest that these bacteriocins, named klebicin K and raoultellin L, could have potential use against some foodborne pathogens.

Bacteriocin production by Staphylococcus aureus involved in bovine mastitis in Brazil.

In the present study, 257 Staphylococcus spp. strains were isolated from bovine mastitis cases in 56 different Brazilian dairy herds located in the southeast region of the country and tested for antimicrobial substance (AMS) production. Forty-six strains (17.9%) exhibited AMS production and their identification as Staphylococcus aureus was based on the presence of Gram-positive cocci and on positive results in tests for the ability to coagulate rabbit plasma, to ferment mannitol, and to produce acetoin. The AMS were characterized as bacteriocins (Bac) by their sensitivity to proteolytic enzymes. The Bac(+) strains were tested for resistance to 14 antimicrobial agents showing different profiles. Eighteen strains (39.0%) expressed a multiple antibiotic resistance phenotype. Forty-five strains exhibited at least one plasmid DNA. Cross-immunity analysis against strain S. aureus A70, which produces aureocin A70, amplification of the aurABCD operon (which encodes aureocin A70) or detection of this same operon by DNA/DNA hybridization revealed that 34 strains produce bacteriocins either identical or similar to aureocin A70. The remaining 12 Bac(+) strains produce antimicrobial peptides that seem to be distinct from the best characterized staphylococcal bacteriocins described thus far. The bacteriocin produced by strain 4185 may possess potential practical applications, since it was able to inhibit important pathogens such as Bacillus cereus, Listeria monocytogenes, and Staphylococcus spp. isolated from nosocomial infections.