Fermentation of whey-derived matrices by Kluyveromyces marxianus: alcoholic beverage development from whey and fruit juice mixes.

This research paper addresses the hypotheses that Kluyveromyces marxianus can be cultured with good alcohol production on different whey-derived matrices, and that the fermented product can be used in order to develop alcoholic beverages with acceptable sensory characteristics by mixtures with yeast-fermented fruit-based matrices. Growth and fermentative characteristics of Kluyveromyces marxianus LFIQK1 in different whey-derived matrices were explored by culturing (24 h, 30°C) on reconstituted whey, demineralized whey, heat-treated whey and milk permeate media. High lactose consumption, ethanol production and yield were observed. Reconstituted whey matrix was selected for mixing with orange or strawberry juices fermented using Saccharomyces cerevisiae to obtain alcoholic beverages (W-OR and W-ST, respectively). Consumer evaluation of beverages was performed using acceptability and Check-All-That-Apply (CATA) questions. Good acceptance was observed, significantly higher for W-ST than for W-OR. CATA questions gave information about organoleptic characteristics of beverages. Penalty analysis showed W-R and W-ST were positively associated with smooth/refreshing and fruity/natural, respectively. Liking was represented, accordingly with penalty analysis, by natural/refreshing. A novel alternative for utilization of whey and whey-related matrices by alcoholic beverages production with natural ingredients is presented.

Inhibitory action of antibiotics on Kluyveromyces marxianus.

A bioassay containing Kluyveromyces marxianus in microtiter plates was used to determine the inhibitory action of 28 antibiotics (aminoglycosides, beta-lactams, macrolides, quinolones, tetracyclines and sulfonamides) against this yeast in whey. For this purpose, the dose-response curve for each antibiotic was constructed using 16 replicates of 12 different concentrations of the antibiotic. The plates were incubated at 40°C until the negative samples exhibited their indicator (5-7h). Subsequently, the absorbances of the yeast cells in each plate were measured by the turbidimetric method (λ=600nm) and the logistic regression model was applied. The concentrations causing 10% (IC10) and 50% (IC50) of growth inhibition of the yeast were calculated. The results allowed to conclude that whey contaminated with cephalosporins, quinolones and tetracyclines at levels close to the Maximum Residue Limits inhibits the growth of K. marxianus. Therefore, previous inactivation treatments should be implemented in order to re-use this contaminated whey by fermentation with K. marxianus.

Similarities of Geobacillus bacteria based on their profiles of antimicrobial susceptibility in milk samples.

The genus Geobacillus is composed of thermophilic bacteria that exhibit diverse biotechnological potentialities. Specifically, Geobacillus stearothermophilus is included as a test bacterium in commercial microbiological inhibition methods, although it exhibits limited sensitivity to aminoglycosides, macrolides, and quinolones. Therefore, this article evaluates the antibiotic susceptibility profiles of five test bacteria (G. stearothermophilus subsp. calidolactis C953, Geobacillus thermocatenulatus LMG 19007, Geobacillus thermoleovorans LMG 9823, Geobacillus kaustophilus DSM 7263 and Geobacillus vulcani 13174). For that purpose, the minimum inhibitory concentrations (MICs) of 21 antibiotics were determined in milk samples for five test bacteria using the radial diffusion microbiological inhibition method. Subsequently, the similarities between bacteria and antibiotics were analyzed using cluster analysis. The dendrogram of this multivariate analysis shows an association between a group formed by G. thermocatenulatus and G. stearothermophilus and another by G. thermoleovorans, G. kaustophilus and G. vulcani. Finally, future microbiological methods could be developed in microtiter plates using G. thermocatenulatus as test bacterium, as it exhibits similar sensitivities to G. stearothermophilus. Conversely, G. vulcani, G. thermoleovorans and G. kaustophilus show higher MICs than G. thermocatenulatus.

Transfer of antibiotics from goat's milk to rennet curd and whey fractions during cheese-making.

The transfer of 35 antibiotics from milk to curd and whey was evaluated. Cheeses were produced at laboratory scale, from antibiotic-free goat's milk spiked with different antibiotic concentrations between 0.25 and 4 times the Maximum Residue Limits established in milk. Drug concentrations in milk, curd and whey were analysed by UHPLC-HRMS. Results indicated that most antibiotics were mainly transferred from milk to whey (up to 85.9%), with retention percentages in the curd lower than 50%, except for ceftiofur (59.7%) and dicloxacillin (52.8%). In most cases, drug distribution was unaffected by the antibiotic concentration in milk and correlated significantly to the drug lipophilicity (Log P) for ß-lactams (R2 = 0.54) and sulfonamides (R2 = 0.62). When drug ionization was considered (Log D), improved correlation coefficients were obtained for macrolides (R2 = 0.98). However, other factors besides the drug solubility should be considered to explain and predict the partitioning of antibiotics during cheese-making.

Toxicity of fluoroquinolones on the cladoceran Daphnia magna.

This study evaluates the acute and chronic toxicological effects of six fluoroquinolones on the mortality and growth of Daphnia magna. The NOECs calculated with the multivariate Probit regression model for the chronic study were 56 µg/L ciprofloxacin, 63 µg/L enrofloxacin, 78 µg/L levofloxacin, 85 µg/L marbofloxacin, 69 µg/L norfloxacin, and 141 µg/L ofloxacin. The risk quotients were determined using the measure environmental concentrations reported in water sources from different countries. The risks were low and moderate in water samples from rivers and lakes, although concentrations of ciprofloxacin, norfloxacin, and ofloxacin reported in some countries can cause toxicological damage to D. magna. In addition, urban wastewater and hospital wastewater samples constitute a threat to D. magna (high and moderate risks), requiring the treatment of these wastewater. PRACTITIONER POINTS: The NOECs calculated with the multivariate Probit model for the six fluoroquinolonas are between 56 µg/L ciprofloxacin and 141 µg/L ofloxacin. The levels of ciprofloxacin, norfloxacin, and ofloxacin in urban wastewater and hospital wastewater produce moderate and high risks for D. magna. Water and river samples from some countries containing ciprofloxacin, norlfoxacin, and ofloxacin present high risks for D. magna.

Sistema microbiológico de cinco bioensayos para la detección de residuos de antibióticos / Five-assay microbiological system for the screeningf antibiotic residues

Abstract A novel microbiological system in microtiter plates consisting of five bioassays is presented for the detection and classification of antibiotic residues in milk. The bioassays were optimized for the detection of beta-lactams (Bioassay B: Geobacillus stearothermophilus), macrolides (Bioassay M: Bacillus megaterium with fusidic acid), tetracyclines (Bioassay T: B. megaterium with chloramphenicol), quinolones (Bioassay Q: Bacillus licheniformis) and sulfamides (Bioassay QS: B. licheniformis with trimethoprim) at levels near the maximum residue limits (MRL). The response of each bioassay was interpreted visually (positive or negative) after 4-5.5h of incubation. The system detects and classifies beta-lactams (5 pg/l of amoxicillin, 4 pg/l of ampicillin, 36 pg/l of cloxacillin, 22 pg/l of amoxicillin, 3 pg/l of penicillin, 114 pg/l of cephalexin, 89pg/l of cefoperazone and 116 pg/l of ceftiofur), tetracyclines (98 pg/l of chlortetracycline, 92 pg/l of oxytetracycline and 88 pg/l of tetracycline), macrolides (33 pg/l of erythromycin, 44 pg/l of tilmicosin and 50 pg/l of tylosin), sulfonamides (76 pg/l of sulfadiazine, 85 pg/l of sulfadimethoxine, 77 pg/l of sulfamethoxazole and 87pg/l of sulfathiazole) and quinolones (94 pg/l of ciprofloxacin, 98 pg/l of enrofloxacin and 79 pg/l marbofloxacin). In addition, the specificity values were high for B, T, Q (99.4%), M (98.8%) and QS (98.1%) bioassays. The control of antibiotics through this system can contribute to improving the quality and safety of dairy products.

Resumen Se presenta un novedoso sistema microbiológico en placas de microtitulación compuesto por 5 bioensayos para la detección y clasificación de residuos de antibióticos en leche. Los bioensayos fueron optimizados para la detección de betalactámicos (bioensayo B: Geobacillus stearothermophilus), macrólidos (bioensayo M: Bacillus megaterium con ácido fusídico), tetraciclinas (bioensayo T: Bacillus megaterium con cloranfenicol), quinolonas (bioensayo Q: Bacillus licheniformis) y sulfamidas (bioensayo QS: Bacillus licheniformis con trimetoprima), a niveles cercanos a los límites máximos de residuos (LMR). La respuesta de cada bioensayo se interpretó visualmente (positiva o negativa) después de 4 a 5,5 h de incubación. El sistema detecta y clasifica betalactámicos (5 pg/l de amoxicilina, 4 pg/l de ampicilina, 36 pg/l de cloxacilina, 22 pg/l de amoxicilina, 3 pg/l de penicilina, 114 pg/l de cefalexina, 89 pg/l de cefoperazona y 116 pg/l de ceftiofur), tetraciclinas (98 pg/l de clortetraciclina, 92 pg/l de oxitetraciclina y 88 pg/l de tetraciclina), macrólidos (33 pg/l de eritromicina, 44 pg/l de tilmi-cosina y 50 pg/l de tilosina), sulfamidas (76 pg/l de sulfadiacina, 85 pg/l de sulfadimetoxina, 77 pg/l de sulfametoxazol y 87 pg/l de sulfatiazol) y quinolonas (94 pg/l de ciprofloxacina, 98 pg/l de enrofloxacina y 79pg/l de marbofloxacina). Además, los valores de especificidad fueron altos para los bioensayos B, T, Q (99,4%), M (98,8%) y QS (98,1%). El control de residuos de antibióticos mediante este sistema puede contribuir a mejorar la calidad e inocuidad de los productos lácteos.

Five-assay microbiological system for the screening of antibiotic residues.

A novel microbiological system in microtiter plates consisting of five bioassays is presented for the detection and classification of antibiotic residues in milk. The bioassays were optimized for the detection of beta-lactams (Bioassay B: Geobacillus stearothermophilus), macrolides (Bioassay M: Bacillus megaterium with fusidic acid), tetracyclines (Bioassay T: B. megaterium with chloramphenicol), quinolones (Bioassay Q: Bacillus licheniformis) and sulfamides (Bioassay QS: B. licheniformis with trimethoprim) at levels near the maximum residue limits (MRL). The response of each bioassay was interpreted visually (positive or negative) after 4-5.5h of incubation. The system detects and classifies beta-lactams (5µg/l of amoxicillin, 4µg/l of ampicillin, 36µg/l of cloxacillin, 22µg/l of amoxicillin, 3µg/l of penicillin, 114µg/l of cephalexin, 89µg/l of cefoperazone and 116µg/l of ceftiofur), tetracyclines (98µg/l of chlortetracycline, 92µg/l of oxytetracycline and 88µg/l of tetracycline), macrolides (33µg/l of erythromycin, 44µg/l of tilmicosin and 50µg/l of tylosin), sulfonamides (76µg/l of sulfadiazine, 85µg/l of sulfadimethoxine, 77µg/l of sulfamethoxazole and 87µg/l of sulfathiazole) and quinolones (94µg/l of ciprofloxacin, 98µg/l of enrofloxacin and 79µg/l marbofloxacin). In addition, the specificity values were high for B, T, Q (99.4%), M (98.8%) and QS (98.1%) bioassays. The control of antibiotics through this system can contribute to improving the quality and safety of dairy products.

Effect of thermal treatment of whey contaminated with antibiotics on the growth of Kluyveromyces marxianus.

The objective of the studies reported in this research communication was to investigate the use of whey contaminated with antibiotics such as cephalosporins, quinolones and tetracyclines as a nutrient medium for the growth of Kluyveromyces marxianus with particular attention to the effect of thermal treatment used to overcome the inhibitory effects of antibiotic concentrations close to the Maximum Residue Limits. The heat treatments at 120 °C for 40 min, 120 °C for 83 min, and 120 °C for 91 min caused total inactivation of cephalosporins, tetracyclines and quinolone residues in whey respectively.

Albendazole residues in goat's milk: Interferences in microbial inhibitor tests used to detect antibiotics in milk.

Albendazole (ABZ) residues in goat's milk and their effect on the response of microbial inhibitor tests used for screening antibiotics were evaluated. A total of 18 Murciano-Granadina goats were treated with ABZ and individually milked once a day over a 7-day period. ABZ quantification was performed by high performance liquid chromatography. The ABZ parent drug was not detected. The maximum concentration of its metabolites (ABZ sulfoxide, ABZ sulfone, and ABZ 2-aminosulfone) was reached on the 1st day post treatment (260.0 ± 70.1 µg/kg, 112.8 ± 28.7 µg/kg, 152.0 ± 23.6 µg/kg, respectively), decreasing to lower than the maximum residue limit (MRL, 100 µg/kg) on the 3rd day post treatment. Milk samples were also analyzed by microbial tests [Brilliant Black Reduction Test (BRT) MRL, Delvotest SP-NT MCS and Eclipse 100], and only one positive result was found for Delvotest SP-NT MCS and Eclipse 100. However, a high occurrence of positive outcomes was obtained for BRT MRL during 6 days post treatment, whereas ABZ residues were not detected from the 4th day post administration, suggesting that factors other than the antiparasitic agent might affect the microbial test response.

Interferences on microbial inhibitor tests related to ivermectin treatment in lactating dairy goats.

This Research Communication reports interferences related to the administration of ivermectin in lactating dairy goats on the response of microbial tests for screening antibiotics in milk. Twenty-eight Murciano-Granadina goats, naturally infested with Sarcoptes scabiei var. caprae, were treated with a subcutaneous injection of ivermectin (200 µg/kg b.w.). To prevent re-infestation, a second dose was applied 7 d later. Individual milk samples were collected, daily, up to 15 d post-treatment. Milk samples were analysed by microbial inhibitor tests (BRT MRL, Delvotest SP-NT MCS and Eclipse 100) and ivermectin residues were quantified by HPLC. A large number of positive results were obtained for all microbial tests, especially on the first day after treatment (BRT MRL = 46·4%; Delvotest SP-NT MCS = 14·3%; and Eclipse 100 = 17·8%). However, the highest concentration of drug residues in milk (24·3 ng/ml) was detected on the tenth day after treatment, when positive outcomes were relatively lower (BRT MRL = 17·8%; Delvotest SP-NT MCS = 10·7%; and Eclipse 100 = 7·4%). Results herein suggest that factors related to the ivermectin treatment other than drug residues in milk, or alterations produced by the parasitic disease itself affecting the immune response of animals, could be the cause of false-positive results in microbial tests. It can be concluded that the application of ivermectin in dairy goats infested with sarcoptes mange during lactation produces persistent drug residues in milk, and could also cause false-positive results in microbial inhibitor tests for screening antibiotics.

Novel bioassay using Bacillus megaterium to detect tetracycline in milk.

Tetracyclines are used for the prevention and control of dairy cattle diseases. Residues of these drugs can be excreted into milk. Thus, the aim of this study was to develop a microbiological method using Bacillus megaterium to detect tetracyclines (chlortetracycline, oxytetracycline and tetracycline) in milk. In order to approximate the limits of detection of the bioassay to the Maximum Residue Limit (100µg/l) for milk tetracycline, different concentrations of chloramphenicol (0, 1000, 1500 and 2000µg/l) were tested. The detection limits calculated were similar to the Maximum Residue Limits when a bioassay using B. megaterium ATCC 9885 spores (2.8×10(8)spores/ml) and chloramphenicol (2000µg/l) was utilized. This bioassay detects 105µg/l of chlortetracycline, 100µg/l of oxytetracycline and 134µg/l of tetracycline in 5h. Therefore, this method is suitable to be incorporated into a microbiological multi-residue system for the identification of tetracyclines in milk.

Microbiological bioassay using Bacillus pumilus to detect tetracycline in milk.

The tetracyclines (TCs) are widely used in the treatment of several diseases of cattle and their residues may be present in milk. To control these residues it is necessary to have available inexpensive screening methods, user-friendly and capable of analysing a high number of samples. The purpose of this study was to design a bioassay of microbiological inhibition in microtiter plates with spores of Bacillus pumilus to detect TCs at concentrations corresponding to the Maximum Residue Limits (MRLs). Several complementary experiments were performed to design the bioassay. In the first study, we determined the concentration of spores that produce a change in the bioassay's relative absorbance in a short time period. Subsequently, we assessed the concentration of chloramphenicol required to decrease the detection limit (DL) of TCs at MRLs levels. Thereafter, specificity, DL and cross-specificity of the bioassay were estimated. The most appropriate microbiological inhibition assay had a B. pumilus concentration of 1.6 × 10(9) spores/ml, fortified with 2500 µg chloramphenicol/l (CAP) in Mueller Hinton culture medium using brilliant black and toluidine blue as redox indicator. This bioassay detected 117 µg chlortetracycline/l, 142 µg oxytetracycline/l and 105 µg tetracycline/l by means of a change in the indicator's colour in a period of 5 h. The method showed good specificity (97.9%) which decreased slightly (93.3%) in milk samples with high somatic cell counts (>250,000 cells/ml). Furthermore, other antimicrobials studied (except neomycin) must be present in milk at high concentrations (from >5 to >100 MRLs) to produce positive results in this assay, indicating a low cross specificity.

Detection of antibiotics in goat's milk: effect of detergents on the response of microbial inhibitor tests.

The aim of the study was to evaluate the interference of acid and alkaline detergents employed in the cleaning of milking equipment of caprine dairy farms on the performance of microbial tests used in antibiotic control (BRT MRL, Delvotest MCS, and Eclipse 100). Eight concentrations of commercial detergents, five acid (0-0.25%) and five alkaline (0-1%) were add to antimicrobial-free goat's milk to evaluate the detergent effect on the response of microbial inhibitor tests. To evaluate the effect of detergents on the detection capability of microbial tests two detergents at 0.5 ml/l (one acid and one basic) and eight concentrations of four ß-lactam antibiotics (ampicillin, amoxicillin, cloxacillin and benzylpenicillin) were used. Milk without detergents was used as control. The spiked samples were analysed twelve times by three microbial tests. The results showed that the presence of acid detergents did not affect the response of microbial tests for any of the concentrations tested. However, at concentrations equal to or greater than 2 ml/l alkaline detergents positive results were found in microbial tests (16.7-100%). The detection limits of the screening tests for penicillins were not modified substantially by the presence of detergents. In general, the presence of acid and alkaline detergents in goat's milk did not produce a great interference in the microbial tests, only high concentrations of detergents could cause non-compliant results, but these concentrations are difficult to find in practice if proper cleaning procedures are applied in goat dairy farms.

Effect of storage and preservation of milk samples on the response of microbial inhibitor tests.

The aim of this study was to evaluate the effect of storage time (4 °C) on milk samples and the use of azidiol as preservative on the results of microbial inhibitor tests used to detect antimicrobials in milk. For this purpose, 16 milk bulk samples divided into two aliquots, preservative-free and with azidiol, spiked with 12 concentrations of amoxicillin, ampicillin, penicillin G and oxytetracycline, were used. The milk samples were analysed using the BRT MRL, Delvotest MCS Accelerator and Eclipse 100 at 0, 24, 48 and 72 h stored at 4 °C. The logistic regression model was applied to study the effect of storage time (ST), preservative (P) and their interaction (ST×P). At a concentration equivalent to the Detection Limit (DL), the positive results of microbial inhibitor tests do not remain stable during storage time. These results are more reproducible if samples are stored with a preservative. At Maximum Residue Limits (MRLs) concentration microbial inhibitor tests can detect penicillin for up to 72 h of storage. For oxytetracycline, the BRT MRL and Delvotest MCS tests presents DL exceeding the MRL, therefore they are not sensitivity to this tetracycline (0% positive results). By contrast Eclipse 100, whose DL is lower than the MRL, gave 100% positive results during the 72 h storage period. It can be concluded that it would be convenient to store milk samples with azidiol at 4 °C and to carry out analyses within the first 48 h since milk sampling.

Assessing the risk of pesticide environmental impact in several Argentinian cropping systems with a fuzzy expert indicator.

BACKGROUND: The introduction of transgenic soybean (Glycine max, L.) varieties resistant to glyphosate (GR soybeans) has rapidly expanded in Argentina, increasing pesticide use where only grasslands were previously cultivated. The authors compared an estimate of environmental risk for different crops and active ingredients using the IPEST index, which is based on a fuzzy-logic expert system. For IPEST calculations, four modules are defined, one reflecting the rate of application, the other three reflecting the risk for groundwater, surface water and air. The input variables are pesticide properties, site-specific conditions and characteristics of the pesticide application. The expert system calculates the value of modules according to the degree of membership of the input variables to the fuzzy subsets F (favourable) and U (unfavourable), and they can be aggregated following sets of decision rules. IPEST integrated values of >or= 7 reflect low environmental risk, and values of < 7 reflect high risk. RESULTS: Alfalfa, soybean and wheat showed IPEST values over 7 (low risk), while maize had the lowest IPEST values (high risk). Comparing active ingredients applied in annual and perennial crops, atrazine and acetochlor gave the highest risks of environmental contamination, and they are mainly used in maize. Groundwater was the most affected compartment. CONCLUSIONS: Fuzzy logic provided an easy tool combining different environmental components with pesticide properties to give a simple and accessible risk assessment. These findings provide information about active ingredients that should be replaced in order to protect water and air from pesticide contamination.

Evaluation of a Microbiological Multi-Residue System on the detection of antibacterial substances in ewe milk.

To protect both, public health and the dairy industry, from the presence of antibiotic residues in milk, control programmes have been established, which include the needed screening tests. This work focuses on the application of a Microbiological Multi-Residue System in ewe milk, a method based on the use of six different plates, each seeded with one of the following bacteria: Geobacillus stearothermophilus var. calidolactis (beta-lactams), Bacillus subtilis at pH 8.0 (aminoglycosides), Kocuria rhizophila (macrolides), Escherichia coli (quinolones), B. cereus (tetracyclines) and B. subtilis at pH 7.0 (sulphonamides), respectively. Twenty-three antimicrobial substances were analysed and a logistic regression was established for each substance assayed to relate the antibiotic concentration and the zone of microbial growth inhibition. Great linearity in the response was observed (regression coefficients of over 0.97). This fact suggests the possibility of establishing a decision level of antibiotic concentrations near to the Maximum Residue Limits (MRL). Zones of inhibition were suggested as proposed action levels for the different antimicrobial groups (diameters of inhibition of 18 mm for the aminoglycoside, beta-lactam and sulphonamide plates; 19 mm for the tetracycline plate, 21 mm for the macrolide plate, and 24 mm for the quinolone plate). Specificity and cross-reactivity were also assayed.

Influence of environmental factors on airborne fungi in houses of Santa Fe City, Argentina.

This study investigated concentration and types of airborne fungi spores of indoor air. Forty nine houses of Santa Fe city (Argentina) were examined during one year. This city is characterized by a warm climate with an annual mean temperature of 18.6 degrees C and a relative humidity of 74.6%. Based on similar characteristics, a group of representative houses were selected from both urban and suburban areas. The study began by evaluating the airborne fungal concentrations on environmental factors such as area (urban-suburban), season (winter-summer) and presence/absence of a convection gas-fired heating system during winter. Samples were taken with a Standard RCS centrifugal air sampler which operates on the principle of impact onto an agar media strip by centrifugal force. Strips were filled with malt extract agar containing chloramphenicol to inhibit bacterial growth. After incubation and identification, concentrations of airborne fungi were calculated as CFU/m(3). Indoor results showed the presence of thirteen dominant genera: Cladosporium (58.90%), Alternaria (8.68%), Epicoccum (5.74%), Fusarium (5.37%), Curvularia (3.50%), Acremonium (1.27%), Drechslera (1.26%), Penicillium (1.25%), Aspergillus (1.14%), Mucor (0.61%), Ulocladium (0.57%), Nigrospora (0.48%), Chrysosporium (0.42%) and yeast (3.74%), whose presence varied throughout the year. Multivariate Analyses of Variance were performed to study the influence of environmental factors on concentrations of fungal flora. The results obtained were significant for season (lambda=0.1225), area (lambda=0.6371) and for the presence of a convection gas-fired heating system during winter (lambda=0.4765). ANOVA test for the season showed the highest fungal levels (Geometric Mean) in the summer for Alternaria (181.97 CFU/m(3) vs. 17.38 CFU/m(3)), Fusarium (158.49 CFU/m(3) vs. 2.14 CFU/m(3)), Curvularia (66.07 CFU/m(3) vs. 1.62 CFU/m(3)), Acremonium (7.24 CFU/m(3) vs. 2.29 CFU/m(3)), Mucor (3.16 CFU/m(3) vs. 1.15 CFU/m(3)), Nigrospora (2.34 CFU/m(3) vs. 1.07 CFU/m(3)), Chrysosporium (2.73 CFU/m(3) vs. 1.23 CFU/m(3)). In winter, the highest levels (Geometric Mean) were for Penicillium (5.13 CFU/m(3) vs. 1.91 CFU/m(3)) and yeast (16.22 CFU/m(3) vs. 3.09 CFU/m(3)). As for the area, ANOVA showed the highest fungal levels (Geometric Mean) in suburban areas for Cladosporium (676.08 CFU/m(3) vs. 380.19 CFU/m(3)), Curvularia (6.76 CFU/m(3) vs. 4.27 CFU/m(3)) Ulocladium (3.31 CFU/m(3) vs. 1.20 CFU/m(3)) and yeast (18.62 CFU/m(3) vs. 4.90 CFU/m(3)), while Aspergillus (4.57 CFU/m(3) vs. 1.38 CFU/m(3)), showed the highest levels (Geometric Mean) in the urban area. On the other hand, only Cladosporium showed a higher level (Geometric Mean) in houses without convection gas-fired heating system during winter, compared to that corresponding to heated houses.

Accuracy of BRT and Delvotest microbial inhibition tests as affected by composition of ewe's milk.

The presence of drug residues in ewe's milk samples can be determined by microbial assays. The main limitation of these tests is the large number of false-positive results associated with them. False-positive results can be explained by the interaction of certain substances naturally existing in ewe's milk with the growth of the microorganism used in the test. In this study, milk chemical composition (fat, protein, lactose, total solids), somatic cell counts (SCCs), free fatty acid concentrations, and lactoperoxidase system components were determined in order to investigate their influence on the rate of false-positive results for the BRT and Delvotest microbiological inhibitor tests. Milk samples were obtained after morning milking of Manchega ewes at 15, 30, 45, 60, 75, 90, 105, 120, and 135 days after parturition. The animals did not receive any kind of treatment or medicated feed throughout the experiment. The false-positive rates for BRT and Delvotest were 3.75 and 2.4%, respectively. When the logistic regression model was applied, the percentages of total solids for positive samples were significantly different from those for negative samples (16.90 versus 18.42% for BRT, 16.05 versus 18.45% for Delvotest), while the SCC logarithmic transformation was significantly higher for the positive samples than for the negative samples (5.38 versus 5.11 log units for BRT, 5.32 versus 5.11 log units for Delvotest). Moreover, Delvotest-positive samples exhibited thiocyanate concentrations higher than those of Delvotest-negative samples (8.18 mg/liter versus 6.85 mg/liter). Further analyses are needed to confirm the possible presence of antimicrobial residues in this particular type of milk sample.