Tinea corporis by Microsporum audouinii in a female chimpanzee (Pan troglodytes) from Guinea-Bissau: A case report.

We report a Microsporum audouinii infection in a female juvenile chimpanzee (Pan troglodytes) presenting generalized dermatitis compatible with dermatophytosis. Dermatophyte was identified by macro- and microscopic characterization of skin and scales cultures in Mycosel Agar. The topical treatment applied was effective, having the potential for dermatophytosis treatment in immunocompetent primates.

Phenotypic and Molecular Characterization of Salmonella 1,4,[5],12:i:- R-Type ASSuT Isolates from Humans, Animals, and Environment in Portugal, 2006-2011.

The increase in prevalence of Salmonella 1,4,[5],12:i:- related infections over the last few years has been considered a public health issue in many European countries, especially as this serovar may be associated with tetraresistance to ampicillin, streptomycin, sulfonamides, and tetracyclines (R-type ASSuT). Salmonella 1,4,[5],12:i:- isolates (n = 187) obtained by the Portuguese National Laboratory from different sources, including human clinical cases (n = 170), veterinary (n = 10), environmental (n = 6), and food samples (n = 1), were collected from 15 districts between 2006 and 2011. All isolates were serotyped using the slide agglutination method and results were confirmed by multiplex PCR for the monophasic variant. From the confirmed Salmonella 1,4,[5],12:i:-, R-type ASSuT isolates were selected by disc diffusion and minimal inhibitory concentration (MIC) determination for further characterization by pulsed-field gel electrophoresis restriction with XbaI, virulence genes determination by PCR, additional antimicrobial resistance profiling by disc diffusion, and epidemiological distribution evaluation. Out of the 187 serotyped isolates, 133 were confirmed as Salmonella 1,4,[5],12:i:- with a R-type ASSuT occurrence of 61.7%. Distribution among Portuguese districts showed a higher percentage of reported cases in coastal areas, in particular, in Porto (24.8%), Setúbal (13.5%), and Aveiro (12.8%), probably due to the higher population density. Clonality analysis revealed a high diversity of pulsotypes with the majority of human salmonellosis cases being attributed to sporadic events. All isolates harbored 14 out of the 18 virulence genes evaluated and 87.8% of the isolates showed all the resistance genes frequently associated with the European clone, blaTEM+sul2+straA-straB+tetB+. This study shows that Salmonella 1,4,[5],12:i:- resistant isolates are widely distributed in Portugal. This may be related to a selective advantage offered by R-type ASSuT profile, the presence of multiple virulent features, including the ability to form biofilms, which along with a high diversity of pulsotypes may be responsible for the dissemination through the country.

Biofilm formation by Salmonella enterica serovar 1,4,[5],12:i:- Portuguese isolates: a phenotypic, genotypic, and socio-geographic analysis.

Biofilm-forming ability is well established as an important virulence factor. However, there are no studies available regarding biofilm formation of Salmonella Typhimurium 1,4,[5],12:i:-, the new pandemic serovar in Europe. To address this problem, biofilm expression by Salmonella 1,4,[5],12:i:- was evaluated using 133 isolates from clinical, environmental and animal origins, collected in Portugal from 2006 to 2011. Biofilm detection was performed by phenotypic and genotypic methods, such growth characterization in agar and broth medium, optical density determination by microtiter assays and direct observation by fluorescent in situ hybridization. Biofilm-related genes adrA, csgD and gcpA were detected by PCR. A socio-geographic characterization of strains as biofilm producers was also performed. Results showed that biofilm formation in monophasic Salmonella is widely distributed in Portuguese isolates and could be one of the reasons for its dissemination in this country. Biofilm expression varies between locations, showing that isolates from some regions like Lisboa or Ponta Delgada have an increased ability to persist in the environment due to an enhanced biofilm production. Biofilm formation also varies between risk groups, with a higher prevalence in isolates from salmonellosis infections in women. Therefore, the analysis of the socio-geographic distribution of biofilm-forming bacteria should be considered for the establishment of more adequate regulatory measures or therapeutics regimens, especially important due to the continuous increase of infections caused by antimicrobial resistant microorganisms.

Salmonella spp. in Pet Reptiles in Portugal: Prevalence and Chlorhexidine Gluconate Antimicrobial Efficacy.

A fraction of human Salmonella infections is associated with direct contact with reptiles, yet the number of reptile-associated Salmonellosis cases are believed to be underestimated. Existing data on Salmonella spp. transmission by reptiles in Portugal is extremely scarce. The aim of the present work was to evaluate the prevalence of Salmonella spp. in pet reptiles (snakes, turtles, and lizards), as well as evaluate the isolates' antimicrobial resistance and virulence profiles, including their ability to form biofilm in the air-liquid interface. Additionally, the antimicrobial effect of chlorhexidine gluconate on the isolates was tested. Salmonella was isolated in 41% of the animals sampled and isolates revealed low levels of antimicrobial resistance. Hemolytic and lypolytic phenotypes were detected in all isolates. The majority (90.63%) of the Salmonella isolates were positive for the formation of pellicle in the air-liquid interface. Results indicate chlorhexidine gluconate is an effective antimicrobial agent, against the isolates in both their planktonic and biofilm forms, demonstrating a bactericidal effect in 84.37% of the Salmonella isolates. This study highlights the possible role of pet reptiles in the transmission of non-typhoidal Salmonella to humans, a serious and increasingly relevant route of exposure in the Salmonella public health framework.

Interaction of wild strains of Aspergilla with Aspergillus parasiticus ATCC15517 and aflatoxin production.

Aflatoxins are secondary metabolites produced by some competent mould strains of Aspergillus flavus, A. parasiticus and A. nomius. These compounds have been extensively studied with regards to their toxicity for animals and humans; they are able to induce liver cancer and may cause a wide range of adverse effects in living organisms. Aflatoxins are found as natural contaminants of food and feed; the main line of the strategy to control them is based on the prevention of the mould growth in raw vegetable or during its storage and monitoring of each crop batch. Mould growth is conditioned by many ecological factors, including biotic ones. Hazard characterization models for aflatoxins in crops must take into consideration biotic interactions between moulds and their potential effects on growth development. The aim of this work is to study the effect of the biotic interaction of 14 different wild strains of Aspergilla (different species), with a competent strain (Aspergillus parasiticus ATCC 15517) using an in vitro production model. The laboratory model used was a natural matrix (humidified cracked corn), on which each wild strain challenged the aflatoxin production of a producer strain. Cultures were incubated at 28 degrees C for 12 days and sampled at the 8(th) and 12(th). Aflatoxin detection and quantification was performed by HPLC using a procedure with a MRPL = 1 microg/kg. Results of those interactive cultures revealed both synergic and antagonistic effects on aflatoxin biosynthesis. Productivity increases were particularly evident on the 8(th) day of incubation with wild strains of A. flavipes (+ 70.4 %), A. versicolor (+ 54.9 %) and A. flavus 3 (+ 62.6 %). Antagonistic effects were found with A. niger (- 69.5%), A. fumigatus (- 47.6 %) and A. terreus (- 47.6 %) on the 12(th) day. The increased effects were more evident on the 8(th) of incubation and the decreases were more patent on the 12(th) day. Results show that the development of Aspergilla strains concomitantly with competent aflatoxin producing moulds has a significant influence on the natural biosynthesis pattern.

Antimicrobial resistance in Escherichia coli isolated in wastewater and sludge from poultry slaughterhouse wastewater plants.

The authors investigated the antimicrobial resistance of Escherichia coli isolates in 22 samples of crude inflow, treated effluent, and sludge collected at the wastewater treatment plants of eight poultry slaughterhouses in Portugal. A total of 549 E. coli strains were recovered and tested for resistance to 12 antimicrobial agents. Multidrug resistance was present in 55.7 percent of the isolates. Resistance to tetracycline, ampicillin, trimethoprim/sulfamethoxazole, streptomycin, and enrofloxacin was found in 80.7 percent, 56.5 percent, 47.5 percent, 39.2 percent, and 18.4 percent of the isolates, respectively. Resistance rates of E. coli to nearly all of the tested antibiotics were higher in the strains obtained from the six slaughterhouses that handled conventional broilers than in the two slaughterhouses that handled free-range broilers. Wastewater treatment resulted in an E. coli decrease of between 0.5 log and 3 log; nevertheless, an average of 5.2 x 10(5) CFUs/100 mL were present in the outflow of the plants. These data indicate that the use of antimicrobials in poultry production leads to the selection of a large pool of resistance genes and that wastewater treatment processes are unable to inactivate the bacteria and thus will result in dissemination of resistant E. coli into the environment.

Antimicrobial resistance in Enterococcus spp. and Escherichia coli isolated from poultry feed and feed ingredients.

Poultry feed is at the start of the food safety chain in the "farm-to-fork" model, and might serve as a source of antimicrobial resistant bacteria present in poultry meat. Antimicrobial resistance was investigated in 1137 enterococci and 163 Escherichia coli strains recovered from 23 samples of commercial broiler feed and 66 samples of raw feeding materials taken over half a year timespan. Enumeration of enterococci and E. coli were also performed using traditional plating and fluorescent in situ hybridisation methods. Viable enterococci were detected in all feed samples and in 66% of samples of separate feed ingredients, while E. coli was present in 50% and 32% of feed and raw feeding materials, respectively. The median values (50th percentile) for plate and FISH counts for feeds were, respectively, 2.70 log CFU/g and 5.52 log cells/g for enterococci, and 0.15 log CFU/g and 6.00 log cells/g for E. coli. Among enterococci recovered from feed ingredients, resistance to rifampicin, erythromycin, nitrofurantoin, tetracycline, and ciprofloxacin was found in 59.8%, 21.6%, 21.2%, 18.0% and 6.9% of the isolates, respectively. A considerable proportion of the enterococci isolates obtained from broiler feed displayed resistance to tetracycline (69.1%), rifampicin (58.5%), erythromycin (52.9%) and nitrofurantoin (36.2%). Lower percentage of resistance was observed to chloramphenicol (4.6%), ciprofloxacin (3.9%), vancomycin (1.9%) and ampicillin (1.2%). Among E. coli recovered from feed ingredients and poultry feeds, resistance to ampicillin, tetracycline and streptomycin was found in 22.9%, 27.6% and 19.0% and in 22.4%, 41.4% and 17.0% of the isolates, respectively. These data show that feedstuffs and poultry feeds are extensively contaminated by resistant enterococci and, in a lesser extent, by E. coli, thus leading to their introduction in the farm environment.

Occurrence of aflatoxin B1 in dairy cow feed over 10 years in Portugal (1995-2004).

In Portugal, there is rather little information about the natural occurrence of aflatoxin in feedstuffs. The aim of this work was to report the results of screening the natural incidence of aflatoxin B1 (AFB1) in samples of cattle feed collected from seven dairy cow's farms from Portugal distributed in several locations of the country. One thousand and one samples were taken from 1995 to 2004. High performance liquid chromatography (HPLC) was used for separation, identification and quantification of the compound. Detection limit was 1 microg/kg. Aflatoxin B1 was detected in 374 (37.4%) of the samples. The incidence and mean content of AFB1 was generally low. Levels of aflatoxin B1 above the maximum limit established in Portugal (5 microg/kg) for dairy cattle feed samples were observed in 62 samples (6.2%) with levels ranging from 5.1 to 74 microg/kg. Out of those 62 samples, 3.7% had levels between 5.1 to 10 (mean 7.8); 1.8% had a contamination level of 10.1 to 20 (mean 12.0), and 0.7% exceeded 20.1 microg/kg (mean 50.4). On the last two years (2003-04) none of the samples exceeded the maximum permissible level of the toxin.

Antimicrobial resistance in Enterococcus spp. isolated in inflow, effluent and sludge from municipal sewage water treatment plants.

Antimicrobial resistance of enterococci was investigated in 42 samples of crude inflow, treated effluent and sludge collected in 14 municipal sewage treatment plants of Portugal. A total of 983 enterococci were recovered and tested, using the diffusion agar method, regarding their sensitivity to 10 different antimicrobial drugs. Multidrug resistance was present in 49.4% of the isolates. Only 3.3% and 0.6% of the investigated strains were resistant to ampicillin and vancomycin, respectively. Resistances found against rifampicin (51.5%), tetracycline (34.6%), erythromycin (24.8%) and nitrofurantoin (22.5%), are causes for substantial concern. Almost 14% of isolates were resistant to ciprofloxacin. Wastewater treatment resulted in enterococci decrease between 0.5 and 4log; nevertheless, more than 4.4 x 10(5)CFU/100ml were present in the outflow of the plants. Our data indicate that the use of antimicrobials had created a large pool of resistance genes and that sewage treatment processes are unable to avoid the dissemination of resistant enterococci into the environment.

Fecal contamination of wastewater treatment plants in Portugal.

Reutilization of effluents from wastewater treatment plants (WWTP) for non-potable applications is increasing due to the reduction of sustainable water resources. These products mostly come from municipal WWTP and also from slaughterhouses effluents. The microbiological certification of these products is mandatory before their discharge into the environment. This study evaluates if the treatment applied in WWTP to municipal waters or to poultry slaughterhouse effluents distributed over the Portuguese continental territory is efficient in reducing the microbiological risk associated with the reutilization of those wastewaters and sludges. Fecal indicators Escherichia coli and enterococci were evaluated in 42 and 24 wastewater samples from 14 municipal WWTP and 8 poultry slaughterhouse treatment plants, respectively, by the conventional culture method and a rapid Fluorescent in situ hybridization (FISH) technique. Bacterial enumeration in inflow water from most WWTP was rather high (generally >10(5) cells/ml), for both E. coli and Enterococcus spp., and the bacterial quantification by FISH was generally higher than enumeration by the conventional culture method. In both types of treatment plants studied, bacterial load from effluents and sludges was not statistically different from the inflows, indicating that the treatment applied seems to be equally unable to reduce the microbiological load of the effluents. These findings may jeopardize the safe reuse of treated wastewaters in agriculture and the quality of the water environment. Therefore, products like water, sewage sludge, and biosolids originated from the municipal and slaughterhouse WWTP studied should not be reutilized, and effluents treatment should be urgently reviewed.