Methicillin-resistant Staphylococcus aureus (MRSA) isolated from raw milk and nasal swabs of small ruminants in Abeokuta, Nigeria.

Methicillin-resistant Staphylococcus aureus (MRSA), an important widespread cause of severe infection in both humans and animals, is a significant pathogen of public health concern. This study examined the presence of MRSA in 400 samples comprising 200 raw milks (145 from goat and 55 from sheep) and 200 nasal swabs (145 from goats and 55 from sheep) collected from ten different locations in Abeokuta, Nigeria. Samples were examined using standard bacteriological methods for the isolation and identification of Staphylococcus aureus and culture on oxacillin (6 µg/ml) and cefoxitin (2 µg/ml) selective media for the detection of MRSA. Suspected MRSA isolates were confirmed by latex agglutination test for the detection of penicillin-binding protein 2a (PBP2a). Antibiotic susceptibility testing was determined by Kirby Bauer disc diffusion method. Staphylococcus aureus was detected in 72 (18%) of 400 samples of which 52 (13%) were confirmed as MRSA. Methicillin-resistant S. aureus was detected in raw milk (37 of 200; 18.5%) and nasal swab (15 of 200; 7.5%). There was no significance difference (p > 0.05) in the prevalence of MRSA in sheep (37.7%) and goat (23.4%). The MRSA isolates showed resistance to ampicillin (100%), cloxacillin (100%), sulphamethoxazole-trimethoprim (100%), amoxicillin-clavulanate (84.6%), ceftriaxone (75%), cefuroxime (69.2%), erythromycin (65.4%), streptomycin (38.5%), ciprofloxacin (23.1%), pefloxacin (21.2%) and gentamicin (17.3%). The presence of multidrug-resistant MRSA in small ruminants reared in Abeokuta metropolis may be due to regular use of antibiotics and unhygienic practices by farmers. This in turn constitutes a potential public health risk to the owners, consumers of small ruminant products and the general populace.

Antimicrobial resistance in bacteria isolated from pigs with respiratory clinical signs in Brazil / Resistência a antimicrobianos em bactérias isoladas de suínos com sinais clínicos respiratórios no Brasil

Antimicrobial resistance is a current and important issue to public health, and it is usually associated with the indiscriminate use of antimicrobials in animal production. This study aimed to evaluate the antimicrobial susceptibility profile in bacterial isolates from pigs with clinical respiratory signs in Brazil. One hundred sixty bacterial strains isolated from pigs from 51 pig farms in Brazil were studied. In vitro disk-diffusion method was employed using 14 antimicrobial agents: amoxicillin, penicillin, ceftiofur, ciprofloxacin, enrofloxacin, chlortetracycline, doxycycline, oxytetracycline, tetracycline, erythromycin, tilmicosin, florfenicol, lincomycin, and sulfadiazine/trimethoprim. The majority of isolates were resistant to at least one antimicrobial agent (98.75%; 158/160), while 31.25% (50/160) of the strains were multidrug resistant. Streptococcus suis and Bordetella bronchiseptica were the pathogens that showed higher resistance levels. Haemophilus parasuis showed high resistance levels to sulfadiazine/trimethoprim (9/18=50%). We observed that isolates from the midwestern and southern regions exhibited four times greater chance of being multidrug resistant than the isolates from the southeastern region studied. Overall, the results of the present study showed a great level of resistance to lincomycin, erythromycin, sulfadiazine/trimethoprim, and tetracycline among bacterial respiratory pathogens isolated from pigs in Brazil. The high levels of antimicrobial resistance in swine respiratory bacterial pathogens highlight the need for the proper use of antimicrobials in Brazilian pig farms.(AU)

A resistência antimicrobiana é uma questão atual e muito importante para a saúde pública, geralmente associada ao uso indiscriminado de antimicrobianos na produção animal. Diante disso, foi investigado o perfil de sensibilidade-antimicrobiana em isolados bacterianos de suínos com sinais clínicos respiratórios no Brasil. Foram estudadas 96 isolados provenientes de 51 granjas de suínos do Brasil. O método de disco-difusão foi empregado usando 14 antimicrobianos: amoxicilina, penicilina, ceftiofur, ciprofloxacina, enrofloxacina, clortetraciclina, doxiciclina, oxitetraciclina, tetraciclina, eritromicina, tilmicosina, florfenicol, lincomicina e sulfadiazina/trimetoprim. Streptococcus suis e Bordetella bronchiseptica foram os patógenos que apresentaram maiores níveis de resistência. Haemophilus parasuis apresentou altos níveis de resistência à sulfadiazina/trimetoprim (9/18=50%). Observou-se que isolados das regiões Centro-Oeste e Sul apresentaram quatro vezes mais chance de serem multirresistentes do que os isolados da região Sudeste. A maioria foi resistente a pelo menos um agente antimicrobiano (98,75%; 158/160) e 31,25% (50/160) das estirpes isoladas eram multirresistentes. No geral, os resultados do presente estudo mostraram grande nível de resistência à lincomicina, eritromicina, sulfadiazina/trimetoprim e tetraciclina entre patógenos respiratórios bacterianos isolados de suínos no Brasil. Os altos níveis de resistência antimicrobiana em patógenos bacterianos respiratórios em suínos reforçam a necessidade do uso criterioso de antimicrobianos na suinocultura brasileira.(AU)

Intra- and inter-laboratory agreement of the disc diffusion assay for assessing antimicrobial susceptibility of porcine Escherichia coli.

Reliable assessment of the susceptibility of animal bacterial pathogens to antimicrobials is of paramount importance in the fight against antimicrobial resistance. This work aims to estimate the repeatability (intra-laboratory agreement) and reproducibility (inter-laboratory agreement) of the disc diffusion assay in veterinary laboratories to understand further if the assay has a role in the surveillance of antimicrobial resistance in animals. Seven major veterinary laboratories from all States in Australia participated, and each tested the same panel of isolates five times at three to four-week intervals, against six antimicrobial agents using Clinical and Laboratory Standards Institute protocols. The panel consisted of twenty different isolates from porcine Escherichia coli from clinical cases and a single reference strain (ATCC 25922). Laboratories were blinded to the identity of the isolates, replicates, and to each other. In total, 4200 inhibition zone diameters (mm) were collected, and analysed descriptively, graphically, and with linear mixed models. Regardless of the laboratories and isolate/antimicrobial combinations, the overall very major error rate (proportion of isolates classified as susceptible when actual status is resistant) was 1.6%; the major error rate (proportion of isolates classified as resistant when actual status is susceptible) was 1.6%; and the 'minor error' rate (proportion of isolates with intermediate susceptibility that measure fully susceptible or resistant or vice versa) was 2.4%. The variation between repeated measurements ranged between 4.4-7.2 mm depending on the antimicrobial agent assessed. The reproducibility was always more variable than the repeatability, which suggested some laboratory effects. The repeatability coefficient of disc diffusion was lowest for tetracycline (4.4 mm, 95% CI: 3.8-5.0 mm) and ampicillin (4.6 mm, 95% CI: 4.2-5.2 mm) and highest for trimethoprim-sulfamethoxazole (6.6 mm, 95% CI: 5.9-7.4 mm). The reproducibility coefficient of disc diffusion was lowest for gentamicin (5.4, 95% CI: 4.0-7.2) and highest for trimethoprim-sulfamethoxazole (7.2 mm, 95%CI: 4.5-11.7 mm). The precision of the disc diffusion assay was deemed satisfactory for use in a national surveillance program for clinical porcine E. coli isolates. However, measurement variation of the disc diffusion assay is of concern for isolates with marginal susceptibility or resistance due to increased risk of misclassification.

Occurrence of genes encoding aminoglycoside-modifying enzymes in Escherichia coli isolates from chicken meat.

1. The aim of the experiment was to determine the occurrence of genes encoding aminoglycoside-modifying enzymes (AMEs) in Escherichia coli isolates recovered from chicken meat.2. Antibiotic sensitivity was tested using the disc diffusion test. AMEs and virulence profile were determined by PCR/sequencing.3. Out of 195 meat samples collected, 185 (95%) isolates were identified as E. coli. Disc diffusion showed a resistance value of 22% (n = 42) for at least one of the antibiotic aminoglycosides (AGs) tested (tobramycin, gentamycin, amikacin and kanamycin). PCR screening showed the presence of three classes of AMEs, namely, aac(3)-II (12%), aac(6')-Ib (7%) and aac(2')-Ia (5%). Eight of the 42 isolates were positive for the stx1 and sxt2 genes and were defined as Shiga toxin-producing E coli., while the eae gene was positive in one strain. Among the 42 isolates, group A was the predominant phylogenetic identified (76%), followed by group D (21%). One isolate belonged to subgroup B23.4. The results suggested that chicken meat could be an important reservoir of AMEs, and pose a potential risk by dissemination of resistance to humans through the food chain.

First description of Streptococcus lutetiensis from a diseased cat.

This paper describes for the first time the isolation of Streptococcus lutetiensis in a cat with intestinal lymphoma. The Streptococcus bovis group has undergone significant taxonomic changes over the past two decades and, in 2002, Poyart et al. described two distinct novel species within the genus Streptococcus: Streptococcus lutetiensis and Streptococcus pasteurianus. The bovis group streptococci include commensal species and subspecies or opportunistic pathogens of humans and animals. The cat was referred to the Veterinary Teaching Hospital, University of Bologna for chronic diarrhoea associated with fresh blood. A diagnosis of intestinal lymphoma was advanced. S. lutetiensis was accidentally isolated from the faeces of the cat and identified through MALDI-TOF and 16s rRNA sequencing. The Kirby-Bauer test revealed that the isolate was resistant to enrofloxacin, erythromycin, clindamycin, marbofloxacin and tetracycline. The detection of S. lutetiensis in cat faeces might suggest that it could be a normal inhabitant of cat intestinal tract or that it could be involved in the manifestation of intestinal diseases. Since bacteria belonging to the S. bovis group are considered emerging pathogens, additional research is required to evaluate the role of S. lutetiensis in cats and its role in the transmission of antimicrobial resistance. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study the isolation of Streptococcus lutetiensis from a cat with intestinal lymphoma was described for the first time. An antimicrobial susceptibility test performed by means of the disc diffusion method revealed that the isolate was resistant to enrofloxacin, erythromycin, clindamycin, marbofloxacin and tetracycline. Nowadays the ecological or pathogenetic role of S. lutetiensis in the gut of animals remains unclear but, even if its role as commensal bacterium was confirmed, the presence of multi-resistant S. lutetiensis in cat gut could favour the transmission of antimicrobial resistance to other bacteria.

Pathogenic potential and the role of clones and plasmids in beta-lactamase-producing E. coli from chicken faeces in Vietnam.

BACKGROUND: Antimicrobial resistance (AMR) in food-producing animals is a global public health issue. This study investigated AMR and virulence profiles of E. coli isolated from healthy chickens in Vietnam. E. coli were isolated from fecal samples collected in five chicken farms located in the provinces of Hoa Binh, Thai Nguyen and Bac Giang in the North of Vietnam. These isolates were examined by disk diffusion for their AMR, PCR for virulence and AMR genes, pulsed-field gel electrophoresis for relatedness between blaCMY-2/blaCTX-M-positive isolates, electroporation for transferability of blaCMY-2 or blaCTX-M genes and sequencing for mutations responsible for ciprofloxacin resistance. RESULTS: Up to 99% of indicator isolates were multidrug resistant. Resistance to third-generation cephalosporins (3GC) was encoded by both blaCTX-M and blaCMY-2 genes; blaCTX-M genes being of genotypes blaCTX-M-1, - 14, - 15, - 17, - 57 and - 87, whereas ciprofloxacin resistance was due to mutations in the gyrA and parC genes. Some isolates originating from farms located in different provinces of Vietnam were found to be closely related, suggesting they may have been disseminated from a same source of contamination. Plasmids may also have played a role in the diffusion of 3GC-resistance as the blaCMY-2 gene was located on plasmids A/C and I1, and the blaCTX-M gene variants were carried by I1, FIB, R and HI1. Plasmids carrying the blaCMY-2/blaCTX-M genes also co-transferred resistance to other antimicrobials. In addition, isolates potentially capable of infecting humans, of which some produced blaCMY-2/blaCTX-M, were identified in this study. CONCLUSIONS: Both clones and plasmids could be involved in the dissemination of 3GC-resistant E. coli within and between chicken farms in Vietnam. These results demonstrate the necessity to monitor AMR and control antimicrobial use in poultry in Vietnam.

Ceftaroline resistance in Staphylococcus pseudintermedius gene mecA carriers / Resistência à ceftarolina em Staphylococcus pseudintermedius portadores do gene mecA

Infections caused by methicillin-resistant Staphylococcus aureus (MRSA) being a constant concern, ceftaroline fosamil has been recently approved as a new cephalosporin, active against MRSA, for use in humans; only rare cases of resistance have been reported till date. There is no report of resistance to ceftaroline in Staphylococcus pseudintermedius, which is the main bacterium causing dermatitis and otitis in dogs. To evaluate staphylococcal resistance to ceftaroline, 35 isolates of methicillin-resistant S. pseudintermedius (MRSP), carrying the mecA gene, from 26 dogs with folliculitis and nine dogs with external otitis, underwent disk diffusion test with cefoxitin, oxacillin, and ceftaroline. Tests with cefoxitin and oxacillin showed > 90% sensitivity in methicillin resistance detection. In the disk diffusion test, 97.14% (34/35) were resistant to cefoxitin, 94.29% (33/35) to oxacillin, and 31.43% (11/35) to ceftaroline. Of the ceftaroline-resistant strains, 27.27% (3/11) were obtained from the ears of dogs while the rest (8/11) were from the skin. The current report is the first description of MRSP resistance to ceftaroline.(AU)

Infecções causadas por Staphylococcus aureus resistente à meticilina (MRSA) são uma preocupação médica constante. A ceftarolina fosamila é uma nova cefalosporina ativa contra Staphylococcus aureus resistente à meticilina recentemente aprovada para uso em humanos e raros casos de resistência relatados até agora. Não há relatos de resistência à ceftarolina em Staphylococcus pseudintermedius, principal bactéria causadora de dermatite e otite em cães. Com o objetivo de avaliar a resistência estafilocócica à ceftarolina, 35 amostras de S. pseudintermedius resistentes à meticilina (MRSP), portadoras do gene mecA, provenientes de 26 cães com foliculite e 9 com otite externa foram submetidos ao teste de disco-difusão com cefoxitina, oxacilina e ceftarolina. Os testes realizados com cefoxitina e oxacilina mostraram mais de 90% de sensibilidade na detecção da resistência à meticilina em ambas. No teste da disco-difusão, 97,14% (1/35) foram resistentes à cefoxitina, 94,29% (3/35) à oxacilina e 31,43% (11/35) à ceftarolina. Das cepas resistentes às ceftarolina, 27,27 (3/11) foram provenientes de ouvido de cães e as demais (8/11), provenientes da pele, sendo essa primeira descrição de resistência de MRSP à ceftarolina na literatura atual.(AU)

Multidrug-resistant Salmonellae isolated in Japanese quails reared in Abeokuta, Nigeria.

Salmonellosis is a major bacterial disease causing huge economic losses in the poultry industry worldwide. This study was carried out to determine the period prevalence and antimicrobial susceptibility of Salmonella enterica in Japanese quails in Abeokuta, Nigeria. Four hundred cloacal swabs of quail birds were collected from 4 locations within Abeokuta. Salmonella was isolated from the samples using conventional methods for selective isolation of Salmonella and biochemical identification. Isolates were confirmed by polymerase chain reaction assays for the amplification and detection of Salmonella-associated virulence genes (invA and stn) using specific primers. Antimicrobial susceptibility testing was done using the Kirby-Bauer disk diffusion method. In all, Salmonella was isolated from 14 (3.5%) cloacal swabs. All 14 isolates possessed invA and stn genes. The Salmonella isolates showed resistance to tetracycline (100%), doxycycline (100%), ampicillin (100%), sulphamethoxazole (92.9%), nalidixic acid (85.8%), ceftazidime (78.6%), neomycin (64.3%), streptomycin (50%) and gentamycin (28.6%) but all the isolates were susceptible to ciprofloxacin. The isolates were resistant to at least three antimicrobials indicating multidrug resistance. The results concluded that Japanese quails harbour multidrug-resistant Salmonella which could be transmitted to humans through consumption of contaminated food or by direct and indirect contact with the carrier birds. Antimicrobial resistance could be due to overdependence on antimicrobials. Ciprofloxacin could be considered in the treatment of zoonotic Salmonellosis in humans.

Evaluation of direct Etest for antimicrobial susceptibility testing of bacteria isolated from synovial fluid of horses using enrichment bottles.

This study evaluated the Etest for direct antimicrobial susceptibility testing (AST) of bacteria from equine synovial specimens, incubated in BACTEC enrichment bottles. Ninety-four culture-positive broths were inoculated onto agar to directly determine the minimum inhibitory concentrations (MICs) of 13 antimicrobials, using the Etest (direct Etest). Results were compared with those obtained with the agar dilution reference method, the standard Etest, and the disc diffusion method, after subculture and standardisation of the inoculum. For categorical comparison of AST results, MICs were translated into susceptibility categories, using clinical breakpoints. The direct Etest predicted categorical susceptibility/resistance of bacteria from equine synovial fluid with acceptable accuracy (overall categorical agreement, 91%) and was more reliable than the disc diffusion test. The direct Etest was less accurate than the standard Etest for generating MICs ± 1 log dilution relative to the reference method (overall essential agreement, 69% vs. 89%). As the Etest generated a high percentage of inaccuracies with trimethoprim and sulfadiazine, these were less suitable antimicrobial agents for inclusion. In conclusion, the direct Etest reliably predicted the susceptibility of isolates from equine synovial fluid for the tested antimicrobials, except for trimethoprim and sulfadiazine. Since it did not require subculture and preparation of a standardised inoculum, direct Etest results were available at least 24 h earlier than with other methods, which could facilitate the diagnosis of synovial infections. However, when accuracy is prioritised over speed for MIC determination, the standard Etest is preferred over the direct Etest.

Proposal for agar disk diffusion interpretive criteria for susceptibility testing of bovine mastitis pathogens using cefoperazone 30µg disks.

Cefoperazone is a third generation cephalosporin which is commonly used for bovine mastitis therapy. Bacterial pathogens involved in bovine mastitis are frequently tested for their susceptibility to cefoperazone. So far, the cefoperazone susceptibility testing using 30µg disks has been hampered by the lack of quality control (QC) ranges as well as the lack of interpretive criteria. In 2014, QC ranges for 30 µg cefoperazone disks have been established for Staphylococcus aureus ATCC® 25923 and Escherichia coli ATCC® 25922. As a next step, interpretive criteria for the susceptibility testing of bovine mastitis pathogens should be developed. For this, 637 bovine mastitis pathogens (including 112 S. aureus, 121 coagulase-negative staphylococci (CoNS), 103 E. coli, 101 Streptococcus agalactiae, 100 Streptococcus dysgalactiae and 100 Streptococcus uberis) were investigated by agar disk diffusion according to the document Vet01-A4 of the Clinical and Laboratory Standards Institute (CLSI) using 30µg cefoperazone disks and the results were compared to the corresponding MIC values as determined by broth microdilution also according to the aforementioned CLSI document. Based on the results obtained and taking into account the achievable milk concentration of cefoperazone after regular dosing, the following interpretive criteria were proposed as a guidance for mastitis diagnostic laboratories: for staphylococci and E. coli ≥23mm (susceptible), 18-22mm (intermediate) and ≤17mm (resistant) and for streptococci ≥18mm (susceptible), and ≤17mm (non-susceptible). These proposed interpretive criteria shall contribute to a harmonization of cefoperazone susceptibility testing of bovine mastitis pathogens.

Detecção e avaliação do perfil de sensibilidade antimicrobiana de enterobactérias isoladas de periquitos cara-suja (Pyrrhura griseipectus) em cativeiro / Detection and evaluation of the antimicrobial sensibility profile of enterobacteria isolated from captive Grey-breasted parakeet (Pyrrhura griseipectus)

This study was conducted to detect enterobacteria in captive Grey-breasted parakeet from a commercial breeding located in the state of Ceara. 25 individual cloacal swabs were collected and this material was subjected to microbiological processing to obtain the bacterial isolation, identification, and evaluation of the sensitivity profile to antimicrobials. The bacterial isolation in ten samples was determined, belonging to the genera Enterobacter, Hafnia, Serratia, Escherichia, Pantoea and Klebsiella, in which these last three corresponded to the bacteria most isolated, with respective percentages of 28.6%, 21.4%, and 21.4%. As for the sensitivity profile, antibiotics to which the bacteria showed more resistance were trimethoprim-sulfamethoxazole (57%), tetracycline (21%), and azithromycin (43%). Thus, it was shown that the parakeets were carriers of Enterobacteriaceae with resistance characteristics.(AU)

Uso de antimicrobiano nanoparticulado para o tratamento da mastite subclínica de ovelhas de corte no período seco / Use of nanoparticulated antimicrobians to treat subclinical mastitis of ewes during the dry period

A inflamação da glândula mamária é uma das principais causas de prejuízo na ovinocultura. Este estudo teve como objetivo investigar as taxas de cura do tratamento da mastite subclínica após infusão intramamária de princípio ativo antimicrobiano no momento da secagem, em formulações convencional e nanoparticulada. Os rebanhos estavam localizados em São Carlos, São Paulo, Brasil. Analisou-se um total de 584 glândulas mamárias de 307 ovelhas de aptidão para produção de carne. Triagem prévia dos casos subclínicos de mastite foi efetuada por meio do California Mastitis Test (CMT) e/ou da contagem de células somáticas (CCS). Análises microbiológicas foram realizadas para confirmação da etiologia infecciosa. As glândulas mamárias com mastite subclínica foram distribuídas em três grupos: G1 (Controle; glândulas mamárias que não receberam tratamento antimicrobiano); G2 (glândulas mamárias em que foi administrado 100 mg de cloxacilina benzatina em estrutura convencional) e G3 (glândulas mamárias em que foi administrado 50 mg de cloxacilina benzatina em estrutura nanoencapsulada). O tratamento aplicado ao G3 mostrou-se mais eficiente (P=0,047) na cura de glândulas mamárias com mastite subclínica. O uso da cloxacilina nanoencapsulada no momento da secagem de ovelhas de corte auxilia no controle da mastite subclínica infecciosa e reduz os prejuízos consequentes.(AU)

Inflammation of the mammary gland is one of the main causes of losses in sheep-rearing. This study aimed to investigate the cure rates from treating subclinical mastitis after intramammary infusion of active antimicrobial agents as conventional formulations or as nanoparticles, at the time when the ewes are being dried off. A total of 584 mammary glands in 307 ewes in meat-producing herds located in São Carlos, São Paulo, Brazil, were analyzed. Prescreening of subclinical mastitis cases was done using the California mastitis test (CMT) and/or the somatic cell count (SCC). Microbiological analyses were performed to confirm the infectious etiology. The mammary glands with subclinical mastitis were distributed into three groups: G1 (control; mammary glands that did not receive any antimicrobial treatment); G2 (mammary glands to which 100mg of benzathine cloxacillin in conventional form were administered); and G3 (mammary glands to which 50mg of benzathine cloxacillin in nanoparticulate form were administered). The treatment applied to G3 was more efficient (P=0.047) in curing mammary glands with subclinical mastitis. Use of cloxacillin nanoparticles at the time when the ewes are being dried off helps to control infectious subclinical mastitis and reduces consequential losses among meat-producing herds.(AU)

Cloacal enterobacteria isolated from captive roadside hawks (Rupornis magnirostris, GMELIN, 1788) and their antimicrobial susceptibility profile / Enterobactérias isoladas da cloaca de Gaviões-carijós (Rupornis magnirostris, GMELIN, 1788) cativos e seu perfil de susceptibilidade a antimicrobianos

Knowledge of the enterobacteria present in the roadside hawk can bring about an understanding of infectious diseases that can affect this bird, as well as other animals and/or humans, while also adding information of great ecological importance. Thus, the aim of the study was to determine the enterobacteria present in the cloaca of captive roadside hawks and antimicrobial susceptibility profile. Initially, cloacal samples from nine specimens were collected with the aid of swabs. Samples were placed in petri dishes with MacConkey agar, Hektoen agar, EMB agar and Salmonella-Shigella (SS) agar and incubated for 24 h at 35C. After incubation, the microorganisms were submitted to biochemical testing to confirm the presence of enterobacteria. Thereafter, the susceptibility profile of bacteria to antimicrobial agents was evaluated by a disk diffusion test according to the Clinical and Laboratory Standards Institute (CLSI). Escherichia coli (77.8%), Klebsiella oxytoca (11.1%), Klebsiella pneumoniae (11.1%) and Salmonella spp. (55.6%) were isolated from the collected samples. Among the isolates, some bacteria showed resistance to up to three antimicrobial agents. This study has brought greater insight about the enterobacteria present in the roadside hawk (Rupornis magnirostris), exhibiting a significant percentage of enterobacteria important to public health; also, it showed the occurrence of strains with resistance profile to antimicrobial agents...

O conhecimento das enterobactérias presentes em Gavião-carijó pode trazer uma compreensão sobre as doenças infecciosas que podem acometer essa ave, como também outros animais e/ou humanos, além de trazer mais informações sobre essa espécie de grande importância ecológica. Desta forma, o objetivo do estudo foi determinar as enterobactérias presentes na cloaca de Gaviões-carijós cativos e seu perfil de susceptibilidade a antimicrobianos. Inicialmente, foram coletadas amostras cloacais de nove espécimes com o auxílio de swabs. As amostras foram plaqueadas em ágar MacConkey, ágar Hektoen, ágar EMB e ágar SS e incubadas por 24 h a 35C. Após incubação, as colônias foram submetidas às provas bioquímicas para confirmação da presença de enterobactérias. Posteriormente, o perfil de susceptibilidade das bactérias frente a agentes antimicrobianos foi avaliado através do teste da difusão em disco de acordo com o Clinical and Laboratory Standards Institute (CLSI). Escherichia coli (77,8%), Klebsiella oxytoca (11,1%), Klebsiella pneumoniae (11,1%) e Salmonella spp. (55,6%) foram isoladas das amostras coletadas. Dentre as cepas isoladas, algumas apresentaram resistência a até três antimicrobianos. O presente estudo trouxe um maior conhecimento sobre as enterobactérias presentes no Gavião-carijó (Rupornis magnirostris), mostrando um percentual significativo de enterobactérias de importância na saúde pública, evidenciando também a ocorrência de cepas com perfil de resistência a agentes antimicrobianos...

Enumeration, identification and safety proprieties of lactic acid bacteria isolated from pork sausage / Enumeração, identificação e propriedades de segurança de bactérias ácido-lácticas isoladas de linguiça suína

Lactic Acid Bacteria (LAB) are indigenous microorganisms occurring in pork sausages. The utilization of selected autochthonous LAB may improve the safety of meat products. This study aims to enumerate and identify LAB in pork sausage and to characterize their safety properties, such as antimicrobial susceptibility and antibacterial activity. A total of 189 sealed packages of pork sausages were collected in seven municipalities (27 samples in each city) of Minas Gerais, Brazil. Microbiological analyses were performed to enumerate LAB. Two pre-selection criteria were applied to 567 isolates of LAB: catalase activity and tolerance to pH 2. A total of 32 strains of UFLA SAU were selected, characterized phenotypically and identified through 16S rDNA region sequencing. The susceptibility to antimicrobial and antibacterial activities of isolates was evaluated. The LAB count ranged from 3.079 to 8.987 log10 CFU/g. Lactobacillus plantarum and Lactobacillus paracasei were identified in the samples. UFLA SAU 11, 20, 34, 86, 131 and 258 showed a profile of susceptibility to four antimicrobials: erythromycin, ampicillin, chloramphenicol and gentamycin. In the antibacterial activity test, with exception of UFLA SAU 1, all other strains showed efficiency in inhibiting Escherichia coli, Salmonella Typhiand Listeria monocytogenes. In the statistical analysis there was interaction among strains of Lactobacillus against the pathogens tested. L. monocytogenes (P=0.05) was more sensitive to Lactobacillus strains and the highest inhibitory activity against this pathogen was achieved by strains UFLA SAU 135, 226, 238 and 258. Thus, UFLA SAU 11, 20, 34, 86, 131, 135, 226, 238 and 258 possess safety characteristics for application in meat products.

Bactérias ácido-lácticas (BAL) são microrganismos indígenas em linguiças. A utilização de selecionadas BAL autóctones pode melhorar a segurança dos produtos cárneos. Este estudo objetivou enumerar e identificar BAL em linguiças suínas e caracterizar suas propriedades de segurança, como a susceptibilidade antimicrobiana e a atividade antibacteriana. Um total de 189 embalagens fechadas de linguiça suína foi adquirido em sete municípios (27 amostras em cada cidade) de Minas Gerais, Brasil. Análises microbiológicas para a enumeração de BAL foram realizadas. Dois critérios de pré-seleção foram aplicados para os 567 isolados de BAL: atividade catalase e tolerância ao pH 2. Um total de 32 estirpes UFLA SAU foi selecionado, caracterizado fenotipicamente e identificado por meio do sequenciamento da região 16S rDNA. A susceptibilidade a antimicrobianos e a atividade antimicrobiana dos isolados foram avaliadas. Nas linguiças, a contagem de BAL variou de 3,079 a 8,987log10 UFC/g. Lactobacillus plantarum e Lactobacillus paracasei foram identificados nas amostras. UFLA SAU 11, 20, 34, 86, 131 e 258 apresentaram um perfil de suscetibilidade a quatro antimicrobianos: eritromicina, ampicilina, cloranfenicol e gentamicina. No teste de atividade antibacteriana, com exceção da UFLA SAU 1, todas as outras estirpes mostraram eficiência em inibir Escherichia coli, Salmonella Typhi e Listeria monocytogenes. Na análise estatística, houve interação entre estirpes de Lactobacillus contra os patógenos testados. L. monocytogenes (P=0,05) foi mais sensível às estirpes de Lactobacillus, e a maior atividade inibitória contra este patógeno foi apresentada por estirpes UFLA SAU 135, 226, 238 e 258. Assim, estirpes UFLA SAU 11, 20, 34, 86, 131, 135, 226, 238 e 258 possuem características de segurança para aplicação em produtos cárneos.

Antimicrobial susceptibility of Histophilus somni isolated from clinically affected cattle in Australia.

This study investigated antimicrobial resistance traits, clonal relationships and epidemiology of Histophilus somni isolated from clinically affected cattle in Queensland and New South Wales, Australia. Isolates (n = 53) were subjected to antimicrobial susceptibility testing against six antimicrobial agents (ceftiofur, enrofloxacin, florfenicol, tetracycline, tilmicosin and tulathromycin) using disc diffusion and minimum inhibitory concentration (MIC) assays. Clonal relationships were assessed using repetitive sequence PCR and descriptive epidemiological analysis was performed. The H. somni isolates appeared to be geographically clonal, with 27/53 (47%) isolates grouping in one cluster from one Australian state. On the basis of disc diffusion, 34/53 (64%) isolates were susceptible to all antimicrobial agents tested; there was intermediate susceptibility to tulathromycin in 12 isolates, tilmicosin in seven isolates and resistance to tilmicosin in one isolate. Using MIC, all but one isolate was susceptible to all antimicrobial agents tested; the non-susceptible isolate was resistant to tetracycline, but this MIC result could not be compared to disc diffusion, since there are no interpretative guidelines for disc diffusion for H. somni against tetracycline. In this study, there was little evidence of antimicrobial resistance in H. somni isolates from Australian cattle. Disc diffusion susceptibility testing results were comparable to MIC results for most antimicrobial agents tested; however, results for isolates with intermediate susceptibility or resistance to tilmicosin and tulathromycin on disc diffusion should be interpreted with caution in the absence of MIC results.

Antimicrobial resistance of F4+ Escherichia coli isolated from Swine in Italy.

Four-hundred and forty-two F4+ pathogenic Escherichia coli were isolated in a period of 10 years (2002-2011), from pigs that were suffering from diarrhoea belonging to Italian swine herds. The strains were analysed for their susceptibility to 12 antimicrobials using the disc diffusion method. During the study period, a statistically significant proportion of isolates resistant to enrofloxacin (14.5-89.3%), marbofloxacin (5.4-60.7%), flumequine (49.1-92.9%), danofloxacin (21.6-80%), florfenicol (9.8-64.3%), thiamphenicol (50-92%) and cefquinome (3.8-44%) was recorded. An increase in resistance (not statistically significant) to gentamicin (63.6-85.7%), apramycin (61.8-82.1%), trimethoprim-sulphamethoxazole (75-89.3%), tetracycline (97-100%) and erythromycin (92.4-100%) was also observed. Based on antimicrobial multiresistance, the strains were collected into three groups: I. resistant to 2-5 antimicrobials; II. resistant to 6-8 antimicrobials; III. resistant to 9-12 antimicrobials. The number of isolates belonging to the first group showed a statistically significant decrease (P < 0.05; R(2)  = 0.896; r = -0.9608), while the isolates belonging to the second and third groups showed a statistically significant increase in resistance (P < 0.05; R(2)  = 0.753; r = 0.8890 and P < 0.05; R(2)  = 0.727; r = 0.8701, respectively) over the period of study. The results of this study suggest the need for continued monitoring of the development of resistance.

Zebrafish fed on recombinant Artemia expressing epinecidin-1 exhibit increased survival and altered expression of immunomodulatory genes upon Vibrio vulnificus infection.

Artemia has been used extensively in aquaculture as fodder for larval fish, shrimp, and shellfish. Epinecidin-1, an antimicrobial peptide, was isolated from grouper (Epinephelus coioides) in 2005. Epinecidin-1 has been previously reported to possess antimicrobial activity against several Gram-positive and Gram-negative bacterial species, including Staphylococcus coagulase, Pseudomonas aeruginosa, Streptococcus pyogenes, and Vibrio vulnificus. In this study, we used electroporation to introduce plasmid DNA encoding a green fluorescent protein (EGFP)-epinecidin-1 fusion protein under the control of the cytomegalovirus (CMV) promoter into decapsulated Artemia cysts. Optimization of various properties (including cyst weight (0.2 g), plasmid concentration (50 µg/100 µl), and pulse voltage (150 V), length (10 ms), and number (2)) resulted in a hatching rate of 41.15%, a transfection efficiency of 49.81%, and a fluorescence intensity (A.U.) of 47.46. The expression of EGFP-epinecidin-1 was first detected by quantitative RT-PCR at 120 h post-electroporation, and protein was identified by Western blot at the same time. Furthermore, the EGFP-epinecidin-1 protein inhibited V. vulnificus (204) growth, as demonstrated by zone of inhibition studies. Zebrafish fed on transgenic Artemia expressing CMV-gfp-epi combined with commercial fodder were more resistant to infection by V. vulnificus (204): survival rate was enhanced by over 70% at 7, 14, and 21 days post-infection, and bacterial numbers in the liver and intestine were reduced. In addition, feeding of transgenic Artemia to zebrafish affected the immunomodulatory response to V. vulnificus (204) infection; expression of immune-responsive genes, including hepcidin and defbl2, was altered, as shown by qPCR. These findings suggest that feeding transgenic Artemia expressing CMV-gfp-epi to larval fish has antimicrobial effects, without the drawbacks of introducing drug residues or inducing bacterial drug resistance.

Dietary supplementation of probiotics affects growth, immune response and disease resistance of Cyprinus carpio fry.

The effects of dietary Bacillus coagulans (MTCC 9872), Bacillus licheniformis (MTCC 6824) and Paenibacillus polymyxa (MTCC 122) supplementation on growth performance, non-specific immunity and protection against Aeromonas hydrophila infection were evaluated in common carp, Cyprinus carpio fry. Laboratory maintained B. coagulans, B. licheniformis and P. polymyxa were used to study antagonistic activity against fish pathogenic bacteria by agar well diffusion assay. Healthy fish fry were challenged by this bacterium for determination of its safety. Fish were fed for 80 days with control basal diet (B0) and experimental diets containing B. coagulans (B1), B. licheniformis (B2) and P. polymyxa (B3) at 10(9) CFU/g diet. Fish fry (mean weight 0.329 ± 0.01 g) were fed these diets and growth performance, various non-specific immune parameters and disease resistance study were conducted at 80 days post-feeding. The antagonism study showed inhibition zone against A. hydrophila and Vibrio harveyi. All the probiotic bacterial strains were harmless to fish fry as neither mortality nor morbidities were observed of the challenge. The growth-promoting influences of probiotic supplemented dietary treatments were observed with fish fry and the optimum survival, growth and feed utilization were obtained with P. polymyxa (B3) supplemented diet. Study of different non-specific innate immunological parameters viz. lysozyme activity, respiratory burst assay and myeloperoxidase content showed significant (p < 0.05) higher values in fish fry fed B3 diet at 10(9) CFU/g. The challenge test showed dietary supplementation of B. coagulans, B. licheniformis and P. polymyxa significantly (p < 0.05) enhanced the resistance of fish fry against bacterial challenge. These results collectively suggests that P. polymyxa is a potential probiotic species and can be used in aquaculture to improve growth, feed utilization, non-specific immune responses and disease resistance of fry common carp, C. carpio.

Antimicrobial resistance among Campylobacter spp. strains isolated from different poultry production systems at slaughterhouse level.

The aim of the current work was to evaluate the prevalence and antimicrobial susceptibility of Campylobacter spp. isolated from different chicken production systems at the slaughterhouse level. Chicken sampling at slaughterhouse was performed for cecum, carcass, and breast meat from flocks of organic (n = 6), extensive indoor (n = 14), and intensive production (n = 14), totaling 34 ceca pools, 64 neck skin pools, and 132 breasts, representing 96,386 chickens. A collection of 167 strains were identified as Campylobacter coli (n = 85) and Campylobacter jejuni (n = 82) and were tested for susceptibility to 11 antimicrobial agents by the disk diffusion method. The frequency of Campylobacter in chicken samples from different production systems was between 79 and 100%. Campylobacter isolated from all origins were resistant to the fluoroquinolones studied (80-98%). However, for ciprofloxacin and ofloxacin, the Campylobacter isolates from extensive indoor chicken were significantly (P < 0.05) less resistant (77 and 58%) than that from organic (97 and 91%) and intensive production (96 and 95%). A high probability of tetracycline resistance occurrence was also found for the Campylobacter spp. tested (58% for C. jejuni and 76% for C. coli). A more frequent profile of multidrug resistance was noticed for isolates from intensive and organic production than for extensive indoor production. These results reinforce the need of efficient strategy implementation to control and reduce Campylobacter in chickens at production and slaughter levels, and the necessity to reduce the use of antimicrobials in poultry sector.

Antimicrobial susceptibility and distribution of inhibition zone diameters of bovine mastitis pathogens in Flanders, Belgium.

In dairy farms, antimicrobial drugs are frequently used for treatment of (sub)clinical mastitis. Determining the antimicrobial susceptibility of mastitis pathogens is needed to come to a correct use of antimicrobials. Strains of Staphylococcus aureus (n=768), Streptococcus uberis (n=939), Streptococcus dysgalactiae (n=444), Escherichia coli (n=563), and Klebsiella species (n=59) originating from routine milk samples from (sub)clinical mastitis were subjected to the disk diffusion method. Disks contained representatives of frequently used antibiotics in dairy. A limited number of clinical breakpoints were available through CLSI, and showed that susceptibility of Staph. aureus, E. coli, and Klebsiella was moderate to high. For streptococcal species however, a large variation between the tested species and the different antimicrobials was observed. In a next step, wild type populations were described based on epidemiological cut off values (EUCAST). Because of the limited number of official cut off values, the data were observed as a mastitis subpopulation and self-generated cut off values were created and a putative wild type population was suggested. The need for accurate clinical breakpoints for veterinary pathogens is high. Despite the lack of these breakpoints, however, a population study can be performed based on the distribution of inhibition zone diameters on the condition that a large number of strains is tested.