Brazilian Journal of Microbiology, Volume: 42, Issue: 1, Published: 2011
  • Microbial deterioration of vacuum-packaged chilled beef cuts and techniques for microbiota detection and characterization: a review Review

    Hernández-Macedo, Maria Lucila; Barancelli, Giovana Verginia; Contreras-Castillo, Carmen Josefina

    Abstract in English:

    Gas production from microbial deterioration in vacuum-packs of chilled meat leads to pack distension, which is commonly referred as blown pack. This phenomenon is attributed to some psychrophilic and psychrotrophic Clostridium species, as well as Enterobacteria. The ability of these microorganisms to grow at refrigeration temperatures makes the control by the meat industry a challenge. This type of deterioration has been reported in many countries including some plants in the Midwestern and Southeastern regions of Brazil. In addition to causing economic losses, spoilage negatively impacts the commercial product brand, thereby impairing the meat industry. In the case of strict anaerobes species they are difficult to grow and isolate using culture methods in conventional microbiology laboratories. Furthermore, conventional culture methods are sometimes not capable of distinguishing species or genera. DNA-based molecular methods are alternative strategies for detecting viable and non-cultivable microorganisms and strict anaerobic microorganisms that are difficult to cultivate. Here, we review the microorganisms and mechanisms involved in the deterioration of vacuum-packaged chilled meat and address the use of molecular methods for detecting specific strict anaerobic microorganisms and microbial communities in meat samples.
  • Beauveria bassiana: quercetinase production and genetic diversity Environmental Microbiology

    Costa, Eula Maria de M. B; Pimenta, Fabiana Cristina; Luz, Christian; Oliveira, Valéria de; Oliveira, Marília; Bueno, Elda; Petrofeza, Silvana

    Abstract in English:

    Beauveria bassiana genetic diversity and ability to synthesize quercetin 2,3-dioxygenase (quercetinase) were analyzed. B. bassiana isolates, obtained from Brazilian soil samples, produced quercetinase after induction using 0.5 g/L quercetin. B. bassiana ATCC 7159 (29.6 nmol/mL/min) and isolate IP 11 (27.5 nmol/ml/min) showed the best performances and IP 3a (9.5 nmol/mL/min) presented the lowest level of quercetinase activity in the culture supernatant. A high level of polymorphism was detected by random amplified polymorphic DNA (RAPD) analysis. The use of internal-transcribed-spacer ribosomal region restriction fragment length polymorphism (ITS-RFLP) did not reveal characteristic markers to differentiate isolates. However, the ITS1-5.8S-ITS2 region sequence analysis provided more information on polymorphism among the isolates, allowing them to be clustered by relative similarity into three large groups. Correlation was tested according to the Person's correlation. Data of our studies showed, that lower associations among groups, level of quercetinase production, or geographical origin could be observed. This study presents the production of a novel biocatalyst by B. bassiana and suggests the possible industrial application of this fungal species in large-scale biotechnological manufacture of quercetinase.
  • Characterization of a novel lipase from Bacillus sp. isolated from tannery wastes Environmental Microbiology

    Ghori, M. I; Iqbal, M. J; Hameed, A

    Abstract in English:

    Kinetics of a lipase isolated from Bacillus sp. was studied. The enzyme showed maximum activity at pH 9 and temperature 60ºC. The Michaelis constant (K M 0.31 µM) obtained from three different plots i.e., Lineweaver-Burk, Hanes-Wolf and Hofstee, was found to be lower than already reported lipases that confirmed higher affinity of the enzyme for its substrate p-NPL (p-nitrophenyl laurate). Vmax of the enzyme was found to be 7.6 µM/mL/min. Energy of activation calculated from Arrhenius plot was found to be 20.607 kJmol-1. Activation enthalpy (ΔH*) had negative trend and the value for the hydrolysis of p-NPL by the enzyme at optimum temperature was -2.748 kJmol-1 . Activation entropy (ΔS*) and free energy of activation (ΔG*) of the enzyme were found to be 1.468 Jmol-1K-1 and -3.237 kJmol-1, respectively at optimum temperature. Low value of Q10 (0.04788) shows high catalytic activity of the enzyme. Mn2+, Fe2+ and Mg2+ enhanced the lipase activity whereas Cu2+, Na+ and Co2+ inhibited the enzyme activity. However, the enzyme activity was not affected significantly by K+ ions. EDTA and SDS also significantly inhibited the lipase activity. Activity of the enzyme was increased in n-hexane while decreased with increase in concentration of acetone, chloroform, ethanol and isopropanol.
  • Quantitative changes in the biochemical composition of lignocellulosic residues during the vegetative growth of Lentinula edodes Environmental Microbiology

    Gaitán-Hernández, Rigoberto; Esqueda, Martín; Gutiérrez, Aldo; Beltrán-García, Miguel

    Abstract in English:

    The chemical changes in barley-straw (BS), wheat-straw (WS) and vineyard-pruning (VP) substrates were determined during colonization of Lentinula edodes mycelia (during primordium development) in solid state fermentation. Primordia appeared 39-50 days after inoculation. VP appeared to promote early sporophore initiation. The concentration of hemicellulose in BS and VP decreased gradually from 25.5% to 15.6% and from 15.8% to 12.3%, respectively. However in WS, hemicellulose decreased from 27.2% to 9.5%. Lignin broke down continuously in BS and WS, with 31.8% and 34.4% degradation, respectively; higher than that of cellulose. During the pinning stage, the C:N ratio decreased in VP and BS, but not in WS. On all substrates the phenols decreased notably throughout the first week of mycelial growth. The time elapsed (days) to pinning was positively correlated with cellulose content (r=0.89), total sugar (r=0.85) and inversely correlated to lignin (r=-1.00) and phenol content (r=-0.55).
  • Succession and growth strategy of a spring microbial community from kezhou sinter in China Environmental Microbiology

    Yang, Hongmei; Lou, Kai

    Abstract in English:

    The succession and growth strategy of a spring microbial community under earthquake action were investigated. The majority of pre-earthquake isolates belonged to the Gammaproteobacteria, including two numerically dominant Stenotrophomonas sp. RB25 and Acinetobacter sp. RB11 (r-strategists). The predominant post-earthquake isolates were Alphaproteobacteria, with Rhizobium sp. RA42 (K-strategists) being dominant among these organisms.
  • Evaluating most probable number method to count and isolate viable methylotrophs Environmental Microbiology

    Kashyap, S

    Abstract in English:

    Nine different receptacles were tested with the MPN method to determine which receptacle was most reliable and economical for MPN counts. Results showed that 96 well PCR plate were the best vessels for this type of analysis and facilitated the isolation of viable Methylotrophs.
  • Culturable fungal diversity of shrimp Litopenaeus vannamei boone from breeding farms in Brazil Environmental Microbiology

    Silva, Lidiane Roberta Cruz da; Souza, Odacy Camilo de; Fernandes, Maria José dos Santos; Lima, Débora Maria Massa; Coelho, Rosalie Reed Rodrigues; Souza-Motta, Cristina Maria

    Abstract in English:

    Litopenaeus vannamei, which is the most common shrimp species cultivated in the northeast of Brazil, is very susceptible to microbial diseases, and this consequently affects productivity. There are reports of bacteria, viruses and protozoa in these shrimp, but not fungi. This study aims to isolate and identify fungi present in shrimp Litopenaeus vannamei, and in their nursery waters, at two breeding farms in Brazil. The pathogenic potential of the isolates was assessed through the qualitative detection of proteases and aflatoxin B production. The 146 isolated fungi comprised 46 species. Aspergillus, Penicillium and Furarium were the three most relevant genera and Aspergillus flavus was the predominant species with a total of 33 isolates. Most of the isolated species are known as potentially pathogenic to humans and other animals. Eighteen isolates of A. flavus and two of A. parasiticus were able to produce aflatoxin B and 33 out of the 46 species produced protease, indicating that these fungi may also become pathogenic to shrimp and their consumers.
  • Seasonality and host preference of arbuscular mycorrhizal fungi of five plant species in the inner Mongolia steppe, China Environmental Microbiology

    Su, Yuan-Ying; Sun, Xin; Guo, Liang-Dong

    Abstract in English:

    The seasonal change and host preference of arbuscular mycorrhizal (AM) colonization and community composition of five common plant species Agropyron cristatum, Anemarrhena asphodeloides, Cleistogenes squarrosa, Leymus chinensis, and Stipa grandis in the Inner Mongolia steppe were investigated. The AM root length colonization rates were different among the five plant species and were generally high in early (May and June) and late (September) growth seasons and low in August. A total of 18 AM fungal species representing five genera were isolated from rhizosphere soils of the five plant species, and most AM fungi had not host specificity, except that Acaulospora sp., Glomus constrictum, G. diaphanum and Glomus sp. showed a certain degree of host preference. Glomus albidum, G. etunicatum and G. geosporum were the dominant species and showed various sporulation patterns in the five plants during the growth seasons. The AM fungal spore densities and species richness increased from May to September and decreased in October and were different in the same month in the five plants. Multivariate analyses revealed that season and host significantly co-affected the AM fungal spore density, species richness, and Shannon-Wiener diversity index, and the season had higher influence than the host.
  • Evaluation of copper resistant bacteria from vineyard soils and mining waste for copper biosorption Environmental Microbiology

    Andreazza, R; Pieniz, S; Okeke, B.C; Camargo, F.A.O

    Abstract in English:

    Vineyard soils are frequently polluted with high concentrations of copper due application of copper sulfate in order to control fungal diseases. Bioremediation is an efficient process for the treatment of contaminated sites. Efficient copper sorption bacteria can be used for bioremoval of copper from contaminated sites. In this study, a total of 106 copper resistant bacteria were examined for resistance to copper toxicity and biosorption of copper. Eighty isolates (45 from vineyard Mollisol, 35 from Inceptisol) were obtained from EMBRAPA (Empresa Brasileira de Pesquisa Agropecuária) experimental station, Bento Gonçalves, RS, Brazil (29º09'53.92''S and 51º31'39.40''W) and 26 were obtained from copper mining waste from Caçapava do Sul, RS, Brazil (30º29'43.48''S and 53'32'37.87W). Based on resistance to copper toxicity and biosorption, 15 isolates were identified by 16S rRNA gene sequencing. Maximal copper resistance and biosorption at high copper concentration were observed with isolate N2 which removed 80 mg L-1 in 24 h. Contrarily isolate N11 (Bacillus pumilus) displayed the highest specific copper biosorption (121.82 mg/L/OD unit in 24 h). GenBank MEGABLAST analysis revealed that isolate N2 is 99% similar to Staphylococcus pasteuri. Results indicate that several of our isolates have potential use for bioremediation treatment of vineyards soils and mining waste contaminated with high copper concentration.
  • Xylanase and cellulase activities during anaerobic decomposition of three aquatic acrophytes Environmental Microbiology

    Nunes, Maíra F; Cunha-Santino, Marcela B. da; Bianchini Jr, Irineu

    Abstract in English:

    Enzymatic activity during decomposition is extremely important to hydrolyze molecules that are assimilated by microorganisms. During aquatic macrophytes decomposition, enzymes act mainly in the breakdown of lignocellulolytic matrix fibers (i.e. cellulose, hemicellulose and lignin) that encompass the refractory fraction from organic matter. Considering the importance of enzymatic activities role in decomposition processes, this study aimed to describe the temporal changes of xylanase and cellulose activities during anaerobic decomposition of Ricciocarpus natans (freely-floating), Oxycaryum cubense (emergent) and Cabomba furcata (submersed). The aquatic macrophytes were collected in Óleo Lagoon, Luiz Antonio, São Paulo, Brazil and bioassays were accomplished. Decomposition chambers from each species (n = 10) were set up with dried macrophyte fragments and filtered Óleo Lagoon water. The chambers were incubated at 22.5ºC, in the dark and under anaerobic conditions. Enzymatic activities and remaining organic matter were measured periodically during 90 days. The temporal variation of enzymes showed that C. furcata presented the highest decay and the highest maximum enzyme production. Xylanase production was higher than cellulase production for the decomposition of the three aquatic macrophytes species.
  • Chromobacterium sp. from the tropics: detection and diversity of phytase activity Environmental Microbiology

    Costa, Patrícia S; Nascimento, Andréa M.A; Lima-Bittencourt, Cláudia I; Chartone-Souza, Edmar; Santos, Fabrício R; Vilas-Boas, Adlane

    Abstract in English:

    Phytases are a group of enzymes that catalyze phytic acid hydrolysis with release of phosphorus (P). The ability of Chromobacterium sp. to produce phytase was detected in 115 out of 118 candidate bacteria isolated from different Brazilian biomas. This is the first report revealing the genus Chromobacterium as phytase producer.
  • Zygomycetes from herbivore dung in the ecological reserve of Dois Irmãos, Northeast Brazil Environmental Microbiology

    Santiago, André Luiz Cabral Monteiro de Azevedo; Trufem, Sandra Farto Botelho; Malosso, Elaine; Santos, Paulo Jorge Parreira dos; Cavalcanti, Maria Auxiliadora de Queiroz

    Abstract in English:

    Thirty-eight taxa of Zygomycetes distributed in 15 genera were recorded from tapir (Tapirus terrestris), camel (Camelus bactrianus), horse (Equus caballus), deer (Cervus elaphus), agouti (Dasyprocta aguti), donkey (Equus asinus), llama (Llama glama) and waterbuck (Kobus ellipsiprymnus) dung collected at the Reserva Ecológica de Dois Irmãos located in Recife, State of Pernambuco, Northeast Brazil. The samples were collected on a monthly basis from June 2005 to May 2006, taken to the laboratory and incubated in moist chambers. Higher number of taxa was observed in the excrements of tapir, followed by deer and donkey. The highest number of species was detected for Mucor, followed by Pilobolus. Statistical analyses showed significant differences in richness of Zygomycetes taxa between the herbivore dung types. Differences of species composition, however, were weak. Seasonality influenced the Zygomycetes species composition but not its richness. Variations in taxa composition between ruminants and non-ruminants dung were non significant.
  • Characterization and comparison of Serratia marcescens isolated from edible cactus and from silkworm for virulence potential and chitosan susceptibility Environmental Microbiology

    Li, Bin; Yu, Rongrong; Liu, Baoping; Tang, Qiaomei; Zhang, Guoqing; Wang, Yanli; Xie, Guanlin; Sun, Guochang

    Abstract in English:

    Representative strains of Serratia marcescens from an edible cactus plant and silkworms were characterized and a comparison based on their cellular fatty acid composition, 16S rRNA and groE gene sequence analysis as well as silkworm virulence and chitosan susceptibility was carried out. Results from this study indicate that there are no significant differences between the phenotypic and molecular characterization, virulence and chitosan susceptibility of the S. marcescens strains from the cactus plant and silkworms. Silkworms inoculated with S. marcescens from either plant or silkworm resulted in nearly 100% mortality. Chitosan solution exhibited strong antibacterial activity against S. marcescens. This activity increased with the increase of chitosan concentration and incubation time regardless of the strain source. Also, the results indicate that the plant associated S. marcescens maybe plays a possible role in the contamination of humans and animals, in particular silkworms, while chitosan showed a potential to control the contamination caused by S. marcescens.
  • The nematicidal effect of some bacterial biofertilizers on Meloidogyne incognita in sandy soil Environmental Microbiology

    El-Hadad, M.E; Mustafa, M.I; Selim, Sh.M; El-Tayeb, T.S; Mahgoob, A.E.A; Aziz, Norhan H. Abdel

    Abstract in English:

    In a greenhouse experiment, the nematicidal effect of some bacterial biofertilizers including the nitrogen fixing bacteria (NFB) Paenibacillus polymyxa (four strains), the phosphate solubilizing bacteria (PSB) Bacillus megaterium (three strains) and the potassium solubilizing bacteria (KSB) B. circulans (three strains) were evaluated individually on tomato plants infested with the root-knot nematode Meloidogyne incognita in potted sandy soil. Comparing with the uninoculated nematode-infested control, the inoculation with P. polymyxa NFB7, B. megaterium PSB2 and B. circulans KSB2, increased the counts of total bacteria and total bacterial spores in plants potted soil from 1.2 to 2.6 folds estimated 60 days post-inoculation. Consequently, the inoculation with P. polymyxa NFB7 increased significantly the shoot length (cm), number of leaves / plant, shoot dry weight (g) / plant and root dry weight (g) / plant by 32.6 %, 30.8 %, 70.3 % and 14.2 %, respectively. Generally, the majority treatments significantly reduced the nematode multiplication which was more obvious after 60 days of inoculation. Among the applied strains, P. polymyxa NFB7, B. megaterium PSB2 and B. circulans KSB2 inoculations resulted in the highest reduction in nematode population comparing with the uninoculated nematode-infested control. They recorded the highest reduction in numbers of hatched juveniles/root by 95.8 %, females/root by 63.75 % and juveniles/1kg soil by 57.8 %. These results indicated that these bacterial biofertilizers are promising double purpose microorganisms for mobilizing of soil nutrients (nitrogen, phosphate and potassium) and for the biological control of M. incognita.
  • Structural interaction between GFP-labeled diazotrophic endophytic bacterium Herbaspirillum seropedicae RAM10 and pineapple plantlets 'Vitória' Environmental Microbiology

    Baldotto, Lílian Estrela Borges; Olivares, Fábio Lopes; Bressan-Smith, Ricardo

    Abstract in English:

    The events involved in the structural interaction between the diazotrophic endophytic bacterium Herbaspirillum seropedicae, strain RAM10, labeled with green fluorescent protein, and pineapple plantlets 'Vitória' were evaluated by means of bright-field and fluorescence microscopy, combined with scanning electron microscopy for 28 days after inoculation. After 6 hours of inoculation, H. seropedicae was already adhered to the roots, colonizing mainly root hair surface and bases, followed by epidermal cell wall junctions. Bacteria adherence in the initial periods occurred mainly in the form of solitary cells and small aggregates with pleomorphic cells. Bacteria infection of root tissue occurred through the cavities caused by the disruption of epidermal cells during the emergence of lateral roots and the endophytic establishment by the colonization of intercellular spaces of the cortical parenchyma. Moreover, within 1 day after inoculation the bacteria were colonizing the shoots. In this region, the preferred sites of epiphytic colonization were epidermal cell wall junctions, peltate scutiform trichomes and non-glandular trichomes. Subsequently, the bacteria occupied the outer periclinal walls of epidermal cells and stomata. The penetration into the shoot occurred passively through stoma aperture followed by the endophytic establishment on the substomatal chambers and spread to the intercellular spaces of spongy chlorenchyma. After 21 days of inoculation, bacterial biofilm were seen at the root hair base and on epidermal cell wall surface of root and leaf, also confirming the epiphytic nature of H. seropedicae.
  • The occurrence of Aeromonas in drinking water, tap water and the Porsuk River Environmental Microbiology

    Kivanc, Merih; Yilmaz, Meral; Demir, Filiz

    Abstract in English:

    The occurrence of Aeromonas spp. in the Porsuk River, public drinking water and tap water in the City of Eskisehir (Turkey) was monitored. Fresh water samples were collected from several sampling sites during a period of one year. Total 102 typical colonies of Aeromonas spp. were submitted to biochemical tests for species differentiation and of 60 isolates were confirmed by biochemical tests. Further identifications of isolates were carried out first with the VITEK system (BioMe˜rieux) and then selected isolates from different phenotypes (VITEK types) were identified using the DuPont Qualicon RiboPrinter® system. Aeromonas spp. was detected only in the samples from the Porsuk River. According to the results obtained with the VITEK system, our isolates were 13% Aeromonas hydrophila, 37% Aeromonas caviae, 35% Pseudomonas putida, and 15% Pseudomonas acidovorans. In addition Pseudomonas sp., Pseudomonas maltophila, Aeromonas salmonicida, Aeromonas hydrophila, and Aeromonas media species were determined using the RiboPrinter® system. The samples taken from the Porsuk River were found to contain very diverse Aeromonas populations that can pose a risk for the residents of the city. On the other hand, drinking water and tap water of the City are free from Aeromonas pathogens and seem to be reliable water sources for the community.
  • Molecular identification of methanogenic archaea from Surti buffaloes (bubalus bubalis), reveals more hydrogenotrophic methanogens phylotypes Environmental Microbiology

    Singh, K.M; Pandya, P.R; Parnerkar, S; Tripathi, A.K; Rank, D.N; Kothari, R.K; Joshi, C.G

    Abstract in English:

    Methane emissions from ruminant livestock are considered to be one of the more potent forms of greenhouses gases contributing to global warming. Many strategies to reduce emissions are targeting the methanogens that inhabit the rumen, but such an approach can only be successful if it targets all the major groups of ruminant methanogens. Therefore, a thorough knowledge of the diversity of these microbes in breeds of buffaloes, as well as in response to geographical location and different diets, is required. Therefore, molecular diversity of rumen methanogens in Surti buffaloes was investigated using 16S rRNA gene libraries prepared from pooled rumen contents from three Surti buffaloes. A total of 171 clones were identified revealing 23 different sequences (phylotypes). Of these 23 sequences, twelve sequences (12 OTUs, 83 clones) and 10 sequences (10 OTUs, 83 clones) were similar to methanogens belonging to the orders Methanomicrobiales and Methanobacteriales, and the remaining 1 phylotype (5 clones) were similar to Methanosarcina barkeri. These unique sequences clustered within a distinct and strongly supported phylogenetic group. Further studies and effective strategies can be made to inhibit the growth of Methanomicrobiales and Methanobacteriales phylotypes to reduce the methane emission from rumen and thus help in preventing global warming.
  • Characterization of cry2-type genes of Bacillus thuringiensis strains from soil-isolated of Sichuan basin, China Environmental Microbiology

    Liang, Hongxia; Liu, Yao; Zhu, Jun; Guan, Peng; Li, Shuangcheng; Wang, Shiquan; Zheng, Aiping; Liu, Huainian; Li, Ping

    Abstract in English:

    Sichuan basin, situated in the west of China, is the fourth biggest basin in China. In order to describe a systematic study of the cry2-type genes resources from Bacillus thuringiensis strains of Sichuan basin, a total of 791 Bacillus thuringiensis strains have been screened from 2650 soil samples in different ecological regions. The method of PCR-restriction fragment length polymorphism (PCR-RFLP) was used to identify the type of cry2 genes. The results showed that 322 Bacillus thuringiensis strains harbored cry2-type genes and four different RFLP patterns were found. The combination of cry2Aa/cry2Ab genes was the most frequent (90.4%), followed by cry2Aa (6.8%) and cry2Ab alone (2.5%), and only one novel type of cry2 gene was cloned from one isolate (JF19-2). The full-length of this novel gene was obtained by the method of thermal asymmetric interlaced PCR (Tail-PCR), which was designated as cry2Ag1 (GenBank No. ACH91610) by the Bt Pesticide Crystal Protein Nomenclature Committee. In addition, the result of scanning electron microscopic (SEM) observation showed that these strains had erose, spherical, bipyramidal, and square crystal. And the results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that these strains harbored about one to three major proteins. These strains exhibited a wide range of insecticidal spectrum toxic to Aedes aegypti (Diptera) and Pieris rapae Linnaeus, 1758 (Lepidoptera). Particularly, JF19-2 contained cry2Ag gene had the highest insecticidal activity. All these researches mentioned above revealed the diversity and particularity of cry2-type gene resources from Bacillus thuringiensis strains in Sichuan basin.
  • Comparative analysis of two component signal transduction systems of the Lactobacillus acidophilus group Food Microbiology

    Cui, Yanhua; Qu, Xiaojun

    Abstract in English:

    The Lactobacillus acidophilus group is a phylogenetically distinct group of closely related lactobacilli. Members of this group are considered to have probiotic properties and occupy different environmental niches. Bacteria generally sense and respond to environmental changes through two component systems (TCSs) which consist of a histidine protein kinase (HPK) and its cognate response regulator (RR). With the use of in silico techniques, the five completely sequenced L. acidophilus group genomes were scanned in order to predict TCSs. Five to nine putative TCSs encoding genes were detected in individual genomes of the L. acidophilus group. The L. acidophilus group HPKs and RRs were classified into subfamilies using the Grebe and Stock classification method. Putative TCSs were analyzed with respect to conserved domains to predict biological functions. Putative biological functions were predicted for the L. acidophilus group HPKs and RRs by comparing them with those of other microorganisms. Some of TCSs were putatively involved in a wide variety of functions which are related with probiotic ability, including tolerance to acid and bile, production of antimicrobial peptides, resistibility to the glycopeptide antibiotic vancomycin, and oxidative condition.
  • Characterization of Lactobacillus from Algerian goat's milk based on phenotypic, 16S rDNA sequencing and their technological properties Food Microbiology

    Marroki, Ahmed; Zúñiga, Manuel; Kihal, Mabrouk; Pérez-Martínez, Gaspar

    Abstract in English:

    Nineteen strains of Lactobacillus isolated from goat's milk from farms in north-west of Algeria were characterized. Isolates were identified by phenotypic, physiological and genotypic methods and some of their important technological properties were studied. Phenotypic characterization was carried out by studying physiological, morphological characteristics and carbohydrate fermentation patterns using API 50 CHL system. Isolates were also characterized by partial 16S rDNA sequencing. Results obtained with phenotypic methods were correlated with the genotypic characterization and 13 isolates were identified as L. plantarum, two isolates as L. rhamnosus and one isolate as L. fermentum. Three isolates identified as L. plantarum by phenotypic characterization were found to be L. pentosus by the genotypic method. A large diversity in technological properties (acid production in skim milk, exopolysaccharide production, aminopeptidase activity, antibacterial activity and antibiotic susceptibility) was observed. Based on these results, two strains of L. plantarum (LbMS16 and LbMS21) and one strain of L. rhamnosus (LbMF25) have been tentatively selected for use as starter cultures in the manufacture of artisanal fermented dairy products in Algeria.
  • Patulin accumulation in apples during storage by Penicillium expansum and Penicillium griseofulvum strains Food Microbiology

    Welke, Juliane Elisa; Hoeltz, Michele; Dottori, Horacio Alberto; Noll, Isa Beatriz

    Abstract in English:

    A part of apples destined to juice production is generally of poor quality. Apples from cold storage or recently harvest (ground harvested or low quality apples) are stored under ambient conditions until they are processed. Since Penicillium expansum and P. griseofulvum are the principal fungal species isolated from stored apples in Brazil, the objective of this study was to investigate the ability of these strains to produce patulin in apples and report the consequences of this type of storage in loss of quality. The toxin was quantified using thin layer chromatography and charge-coupled device camera (TLC-CCD). The rate and quantities that P. expansum and P. griseofulvum can grow and produce patulin are highly dependent on the fungal strain and time. Lesion diameter resulted to be independent of the strain considered. The maximum period of time which apples were kept at cold storage (4 ºC) without patulin accumulation was 27 days. When these apples were kept at 25 ºC during 3 days, both factors lesion diameter and patulin production increased significantly. These results confirm that time in which apples are taken out from cold storage room before juice production is critical in order to prevent patulin accumulation.
  • Comparative analysis of cultural isolation and PCR based assay for detection of Campylobacter jejuni in food and faecal samples Food Microbiology

    Singh, Harkanwaldeep; Rathore, R.S; Singh, Satparkash; Cheema, Pawanjit Singh

    Abstract in English:

    In the present study, the efficacy of polymerase chain reaction (PCR) based on mapA gene of C. jejuni was tested for detection of Campylobacter jejuni in naturally infected as well as spiked faecal and food samples of human and animal origin. Simultaneously, all the samples were subjected to the cultural isolation of organism and biochemical characterization. The positive samples resulted in the amplification of a DNA fragment of size ~589 bp in PCR assay whereas the absence of such amplicon in DNA extracted from E. coli, Listeria, Salmonella and Staphylococcus confirmed the specificity of the primers. Of randomly collected 143 faecal samples comprising human diarrheic stools (43), cattle diarrheic faeces (48) and poultry faecal swabs (52) only 4, 3 and 8, respectively, could be detected by isolation whereas 6, 3 and 10, respectively, were found positive by PCR. However, among food samples viz. beef (30), milk (35), cheese (30), only one beef sample was detected both by culture as well as PCR. Additionally, PCR was found to be more sensitive for C. jejuni detection in spiked faecal and food samples (96.1% each) as relative to culture isolation which could detect the organism in 86.7% and 80% samples, respectively. The results depicted the superior efficacy of PCR for rapid screening of samples owing to its high sensitivity, specificity and automation potential.
  • Antimicrobial activity of essential oils against Vancomycin-Resistant enterococci (VRE) and Escherichia coli O157:H7 in feta soft cheese and minced beef meat Food Microbiology

    Selim, Samy

    Abstract in English:

    Eleven essential oils (EOs) were evaluated for their antibacterial properties, against Vancomycin-Resistant Enterococci (VRE) and E. coli O157:H7. EOs were introduced into Brain Heart Infusion agar (BHI) (15ml) at a concentration of 0.25 to 2% (vol/vol) to determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) for each pathogen evaluated. Results showed that the most active essential oils against bacteria tested were thyme oil, with MIC90 and MBC90 for the VRA strains of 0.25% and 0.5%, respectively. Eucalyptus, juniper and clove oils were the least potent agent, with MIC90 and MBC90 of 2%. Furthermore, the inhibitory effect of these EO were evaluated against VRE and E. coli O157:H7, experimentally inoculated (10³ cfu/g) in Feta soft cheese and minced beef meat, which was mixed with different concentrations (0.1%, 0.5% and 1%) of the EO and stored at 7 ºC for 14 days. Out of eucalyptus, juniper, mint, rosemary, sage, clove and thyme oils tested against target bacteria sage and thyme showed the best results. Clove and mint did not show any effect on VRE and E. coli O157:H7 in both kinds of studied foods. The addition of thyme oil at concentrations of 0.5 and 1% caused best significant reduction in the growth rate of VRE and E. coli O157:H7 in cheese and meat at 7 ºC. It is concluded that selected plant EOs can act as potent inhibitors of both microorganisms in a food product. The results revealed the potential of thyme oil as a natural preservative in feta soft cheese and minced beef meat against VRE and E. coli O157:H7 contamination.
  • Antioxidant activity of Agaricus brasiliensis basidiocarps on different maturation phases Food Microbiology

    Mourão, Francielly; Umeo, Suzana Harue; Takemura, Orlando Seiko; Linde, Giani Andrea; Colauto, Nelson Barros

    Abstract in English:

    Different maturation phases of basidiocarp could affect the bioactivity and concentration of some active substances. A. brasiliensis Wasser et al.(A. blazei Murrill) has shown antitumor activity that could be related to the antioxidant activity. However there is no information of the best basidiocarp maturation phase for extracting antioxidant substances in order to determine the moment of harvesting in mushroom cultivation. The objective of this work was to evaluate the antioxidant activity of A. brasiliensis strains on different basidiocarp maturation phases. The best condition for extraction of A. brasiliensis antioxidants is with methanol as solvent at 60 ºC for 60 min. Strains with closed basidiocarp have higher antioxidant activity than with opened basidiocarp. Antioxidant activity varies in each strain. It was concluded that A. brasiliensis is a natural source of antioxidant compounds. Also there is higher antioxidant activity in closed than opened caps and consequently higher functional activity. It reinforces the synergic action among different A. brasiliensis compounds as a functional food and the importance of further investigation for isolation and characterization of antioxidant substances of A. brasiliensis. It also determines the best harvest period in order to obtain the highest antioxidant activity from basidiocarp.
  • Growth of Byssochlamys nivea in pineapple juice under the effect of water activity and ascospore age Food Microbiology

    Zimmermann, M; Miorelli, S; Massaguer, P.R; Aragão, G.M.F

    Abstract in English:

    The study of thermal resistant mould, including Byssochlamys nivea, is of extreme importance since it has been associated with fruit and fruit products. The aim of this work is to analyze the influence of water activity (a w) and ascospore age (I) on the growth of Byssochlamys nivea in pineapple juice. Mold growth was carried out under different conditions of water activity (a w) (0.99, 0.96, 0.95, 0.93, 0.90) and ascospore age (I) (30, 51, 60, 69, 90 days). Growth parameters as length of adaptation phase (λ), maximum specific growth rate (µmax) and maximum diameter reached by the colony (λ) were obtained through the fit of the Modified Gompertz model to experimental data (measuring radial colony diameter). Statistica 6.0 was used for statistical analyses (significance level α = 0.05). The results obtained clearly showed that water activity is statistically significant and that it influences all growth parameters, while ascospore age does not have any statistically significant influence on growth parameters. Also, these data showed that by increasing a w from 0.90 to 0.99, the λ value substantially decreased, while µmax and λ values rose. The data contributed for the understanding of the behavior of B. nivea in pineapple juice. Therefore, it provided mathematical models that can well predict growth parameters, also helping on microbiological control and products' shelf life determination.
  • Detection of quinolone-resistance mutations in Salmonella spp. strains of epidemic and poultry origin Food Microbiology

    Souza, Roberta Barreiros de; Magnani, Marciane; Ferrari, Rafaela Gomes; Kottwitz, Luciana Bill Mikito; Sartori, Daniele; Tognim, Maria Cristina Bronharo; Oliveira, Tereza Cristina R. M. de

    Abstract in English:

    Mutations into codons Aspartate-87 (62%) and Serine-83 (38%) in QRDR of gyrA were identified in 105 Salmonella strains resistant to nalidixic acid (94 epidemic and 11 of poultry origin). The results show a high incidence of mutations associated to quinolone resistance but suggest association with others mechanisms of resistance.
  • The potential inhibitory effect of cuminum cyminum, ziziphora clinopodioides and nigella sativa essential oils on the growth of Aspergillus fumigatus and Aspergillus flavus Medical Microbiology

    Khosravi, A.R; Minooeianhaghighi, M.H; Shokri, H; Emami, S.A; Alavi, S.M; Asili, J

    Abstract in English:

    The goals of this study were to evaluate the effectiveness of Cuminum cyminum, Ziziphora clinopodioides and Nigella sativa essential oils to inhibit the growth of Aspergillus fumigatus and A. flavus and to evoke ultrastructural changes. The fungi were cultured into RPMI 1640 media in the presence of oils at concentrations of 8, 6, 5, 4, 3, 2, 1.5, 1.25, 1, 0.75 and 0.5 mg/ml in broth microdilution and 2, 1.5, 1 and 0.5 mg/ml in broth macrodilution methods with shaking for 48 h at 28ºC. Conidial and mycelial samples exposed to 0.25, 0.5, 1, 1.5 and 2 mg essential oils/ml for 5 days in 2% yeast extract granulated plus 15% Saccharose media were processed for transmission electron microscopy (TEM). Based on broth dilution methods, C. cyminum and to a lesser extent Z. clinopodioides oils exhibited the strongest activity against A. fumigatus and A. flavus with MIC90 ranging from 0.25 to 1.5 mg/ml, while the oil from N. sativa exhibited relatively moderate activity against two above fungi with MIC90 ranging from 1.5 to 2 mg/ml. The main changes observed by TEM were in the cell wall, plasma membrane and membranous organelles; in particular, in the nuclei and mitochondria. These modifications in fungal structure were associated with the interference of the essential oils with the enzymes responsible for cell wall synthesis, which disturbed normal growth. Moreover, the essential oils caused high vacuolation of the cytoplasm, detachment of fibrillar layer of cell wall, plasma membrane disruption and disorganization of the nuclear and mitochondrial structures. Aspergillus fumigatus and A. flavus growth inhibition induced by these oils were found to be well-correlated with subsequent morphological changes of the fungi exposed to different fungistatic concentrations of the oils. Our results show the anti-Aspergillus activities of C. cyminum, Z. clinopodioides and N. sativa essential oils, which strengthens the potential use of these substances as anti-mould in the future.
  • Correlation between API 50 CH and multiplex polymerase chain reaction for the identification of vaginal lactobacilli in isolates Medical Microbiology

    Brolazo, Eliane Melo; Leite, Domingos Silva; Tiba, Monique Ribeiro; Villarroel, Marina; Marconi, Camila; Simoes, Jose Antonio

    Abstract in English:

    Identification of Lactobacillus sp. strains by phenotypic methods may lead to doubtful results possibly interfering in the reliability of the epidemiological and probiotics studies. Therefore this study aimed to determine the best methodology for the identification of the large diversity of lactobacilli species found in the vagina by comparing two techniques, one based on their biochemical profile and other employing molecular biology. A carbohydrate fermentation test (API 50 CH) was compared with multiplex polymerase chain reaction (PCR) for the identification of species of vaginal lactobacilli from 135 healthy women. The kappa index was used to evaluate agreement between the methods. Using the molecular technique, L. crispatus (32.6%), L. jensenii (25%) and L. gasseri (20.6%) were the most frequent species. However, using the biochemical technique, the most frequent species were: L. acidophilus (34.8%), L. crispatus (27.2%) and L. fermentum (13%). Although L. acidophilus was the most frequent specie found by biochemical tests, no strain of this microorganism was detected by PCR. Agreement between the methods was low for identification of all the most common species. Although rates of L. crispatus detected were similar using both methods (32.6% and 27.2%), agreement between them was relatively low (kappa = 0.52). Conclusions: Our results confirmed the limitation of the biochemical method and the applicability of a previously published molecular method (Multiplex PCR) for the identification of lactobacilli in the vaginal tract, focusing on further necessity of its improvement for also targeting L. vaginalis and L. iners.
  • Growth inhibition and morphological alterations of Trichophyton rubrum induced by essential oil from Cymbopogon winterianus Jowitt ex Bor Medical Microbiology

    Pereira, Fillipe de Oliveira; Wanderley, Paulo Alves; Viana, Fernando Antônio Cavalcanti; Lima, Rita Baltazar de; Sousa, Frederico Barbosa de; Lima, Edeltrudes de Oliveira

    Abstract in English:

    Trichophyton rubrum is one of the most common fungi causer of dermatophytosis, mycosis that affect humans and animals around the world. Researches aiming new products with antifungal activity become necessary to overcome difficulties on treatment of these infections. Accordingly, this study aimed to investigate the antifungal activity of essential oil from Cymbopogon winterianus against the dermatophyte T. rubrum. The antifungal screening was performed by solid medium diffusion method with 16 T. rubrum strains, minimum inhibitory concentration (MIC) and minimum fungicide concentration (MFC) were determined using the microdilution method. The effects on mycelial dry weight and morphology were also observed. Screening showed essential oil in natura inhibited all the tested strains, with inhibition zones between 24-28 mm diameter. MIC50 and MIC90 values of the essential oil were 312 µg/mL for nearly all the essayed strains (93.75 %) while the MFC50 and MFC90 values were about eight times higher than MIC for all tested strains. All tested essential oil concentrations managed to inhibit strongly the mycelium development. Main morphological changes on the fungal strains observed under light microscopy, which were provided by the essential oil include loss of conidiation, alterations concerning form and pigmentation of hyphae. In the oil presence, colonies showed folds, cream color and slightly darker than the control, pigment production was absent on the reverse and with evident folds. It is concluded that C. winterianus essential oil showed activity against T. rubrum. Therefore, it could be known as potential antifungal compound especially for protection against dermatophytosis.
  • The prevalence of enterotoxigenic E. coli isolated from the stools of children aged 0-10 years with diarrhea in mid-anatolia region, Turkey Medical Microbiology

    Işeri, Latife; Apan, Teoman Zafer; Aksoy, Altan; Koç, Figen; Göçmen, Jülide Sedef; Nuristani, Diba

    Abstract in English:

    The stool samples from 245 patients with diarrhea were tested for heat labile toxin (LT) and heat stable toxins (ST) by passive latex agglutination and enzyme immunoassay methods respectively. Twelve (4.9%) enterotoxigenic E. Coli ETEC strains were isolated. Five strains (2%) expressed ST, and 7 (2.8%) expressed LT.
  • Prevalence, aetiology and antibiotic resistance profiles of coagulase negative staphylococci isolated in a teaching hospital Medical Microbiology

    Keim, Luiz S; Torres-Filho, Sylvio R; Silva, Patricia Vollú; Teixeira, Lenise A

    Abstract in English:

    In this paper we carried out a study about prevalence of the clinically significant coagulase negative staphylococcal (CNS) isolates found in an university hospital. Two hundred four CNS isolates from 191 patients obtained between the period of 1998 to 2002, were studied. About 27% (52/191) of the infection cases studied were confirmed as CNS-associated diseases. Blood stream infection (BSI) was the most frequent CNS associated-disease (25%; 13/52). The great majority of the BSI was verified in the Neonatal Intensive Care Unit (NICU). The analysis of the 52 patients medical history showed that 85% of the BSI was acquired in hospital. Most of the CNS nosocomial infections were associated with the use of indwelling medical devices. The incidence of methicillin-resistance among significant CNS isolates was 38%. In this study, a high percentage of exogenous contaminant was verified (60%), indicating that contamination of clinical specimens during sample collection is critical.
  • Antibiotic susceptibility profiles of Mycoplasma hominis and Ureaplasma urealyticum isolated during a population-based study concerning women infertility in northeast Romania Medical Microbiology

    Mihai, Mareş; Valentin, Năstasă; Bogdan, Doroftei; Carmen, Chifiriuc Mariana; Coralia, Bleotu; Demetra, Socolov

    Abstract in English:

    The study was carried out on 1068 infertile women under initial evaluation. For Mycoplasma hominis, the highest resistance rates were registered for ciprofloxacin (72.22%), followed by macrolides and ofloxacin. For Ureaplasma urealyticum, the ciprofloxacin resistance was also high (51.72%), while the resistance rates to other tested antibiotics were significantly lower.
  • Campylobacter insulaenigrae: first isolation report from South American sea lion (Otaria flavescens, (Shaw, 1800) Veterinary Microbiology

    González, Mario; Villanueva, Maria Paz; Debruyne, Lies; Vandamme, Peter; Fernández, Heriberto

    Abstract in English:

    Campylobacter insulaenigrae have been isolated from different pinnipeds but not from South American sea lion (Otaria flavescens). The aim of this work is to report the first isolation of C. insulaenigrae from South American sea lion (Otaria flavescens). The isolate, identified by its phenotypic and molecular characteristics, allow recognizing O. flavescens as a new host for C. insulaenigrae.
  • Behavior of Salmonella heidelberg and Salmonella enteritidis strains following broiler chick inoculation: evaluation of cecal morphometry, liver and cecum bacterial counts and fecal excretion patterns Veterinary Microbiology

    Borsoi, Anderlise; Santos, Luciana Ruschel do; Rodrigues, Laura Beatriz; Moraes, Hamilton Luiz de Souza; Salle, Carlos Tadeu Pippi; Nascimento, Vladimir Pinheiro do

    Abstract in English:

    Over the years, Salmonella Heidelberg (SH) has gained prominence in North America poultry production and in the poultry production of other countries. Salmonella Heidelberg has been isolated and reported from poultry and poultry products in Brazil since 1962, whereas Salmonella Enteritidis (SE) has only emerged as a serious problem in poultry and public health since 1993. These strains of Salmonella can cause intestinal problems in newly hatched chicks, and infection may persist until adulthood. Upon slaughter of chickens, Salmonella can contaminate carcasses, a condition that poses a threat to human health. The aim of this study was to compare the fecal excretion of Salmonella Enteritidis and Salmonella Heidelberg in newly hatched chicks (orally inoculated with 10(5)ufc/mL each) until 20 days of age. In addition, the ratio of cecal villus height:crypt depth (morphometry) and liver and cecum cell counts was analyzed in chicks ranging from 0 to 3 days of age and infected with these two Salmonella strains. One hundred seventeen chicks were separated into one of three experimental groups: a control group, an SE-infected group and an SH-infected group. Eight chicks per group were euthanized at 6, 12 and 72 hours post-inoculation (pi) to allow for Salmonella isolation from the liver and cecum and for the collection of the cecum for villi and crypt analysis. Other birds were allowed to mature to 20 days of age and cloacal swabs were taken at 2, 6, 13 and 20 days pi to compare the fecal excretion of inoculated strains. The Salmonella Enteritidis group had a higher number of cells excreted during the trial. Both strains were isolated from the liver and cecum by 6h pi. At 12h pi the Salmonella Heidelberg group had high cell counts in the cecum. No difference was found in liver cell counts. Both strains showed lower villus height:crypt depth ratio than the control group post-infection.
  • The use of ozonized oil in the treatment of dermatophitosis caused by Microsporum canis in rabbits Veterinary Microbiology

    Daud, Fernanda Vasquez; Ueda, Suely Mitoi Ykko; Navarini, Alessandra; Mímica, Lycia Mara Jenné

    Abstract in English:

    The ozone is effective against most microorganisms due to its high oxidant power. Low concentrations and short-term contact are sufficient to inactivate bacteria, mold, yeast, parasites, seaweeds, protozoa and fungi. Microsporum canis is an important agent of dermatophitosis in human and animal. The aim of the current study was to assess the efficacy of ozonized oil over Microsporum canis in rabbits. Eighteen male New Zealand white rabbits, weight ranging from 2 to 3.2 kg were depilated in the cranial dorso-lateral and right caudal, and cranial and left caudal regions. The regions were inoculated with Microsporum canis, excepting the right caudal region, and were denominated TM, O, OM and M, respectively. After seven days, the treatment of lesions in TM began with 0.12g of terbinaphine 1% cream; in OM and O with 0.12g of ozonized oil; all animals were treated once a day for 28 days. Region M was not treated. Material was collected from those regions for cultivation in Sabouraud agar at day 28 of treatment. In the evolution of the treatment with terbinaphine, of 14 contaminated regions with Microsporum canis ten evolved to cure. With the ozonized oil, of 15 contaminations, four were cured. Clinically, that is, the macroscopic evaluation of lesions showed improvement in the TM and OM treated regions. We can conclude that there was statistical evidence of the protection action of the oil against the dermatophyte.
  • Hematology and productive performance of nile tilapia (Oreochromis niloticus) naturally infected with Flavobacterium columnare Veterinary Microbiology

    Sebastião, F. A; Nomura, D; Sakabe, R; Pilarski, F

    Abstract in English:

    Columnaris disease is one of the main causes of mortality in tilapia rearing and is responsible for large economic losses worldwide. Hematology is a tool that makes it possible to study organisms' physiological responses to pathogens. It may assist in making diagnoses and prognoses on diseases in fish populations. The hematological variables of nile tilapia were studied in specimens with a clinical diagnosis of columnaris disease and in specimens that were disease-free. The total erythrocyte count, hemoglobin rate, hematocrit percentage, mean corpuscular hemoglobin (MCH), mean corpuscular hemoglobin concentration (MCHC), mean corpuscular volume (MCV), organic defense blood cell percentages (leukocytes and thrombocytes) and hepatosomatic and splenosomatic index were determined. The results showed that there were changes in the erythrocytic series and in organic defense blood cells, in the fish infected with the bacterium, with reductions in erythrocytic variables and significant increases in the numbers of circulating lymphocytes and neutrophils.
  • Gastric Helicobacter spp. infection in captive neotropical Brazilian feline Veterinary Microbiology

    Camargo, Pedro Luiz de; Uenaka, Simone Akemi; Motta, Maitê Bette; Adania, Cristina Harumi; Yamasaki, Letícia; Alfieri, Amauri A; Bracarense, Ana Paula F. R. L

    Abstract in English:

    Ten captive neotropical Brazilian feline were submitted to gastroscopic examination and samples of gastric mucosa from fundus, corpus and pyloric antrum were evaluated for the presence of Helicobacter species. Warthin-Starry (WS) staining and PCR assay with species-specific primers and enzymatic cleavage were applied for bacterial detection and identification. Histological lesions were evaluated by haematoxylin and eosin staining. All animals showed normal gross aspect of gastric mucosa. Helicobacter heilmannii was confirmed in 100% of the samples by WS and PCR assay. Mild lymphocytic infiltrate in the lamina propria was observed in eight animals, mainly in the fundus region. Small lymphoid follicles were seen in three animals. No significant association between Helicobacter infection and histological findings was verified. These observations suggest that gastric Helicobacter spp. could be a commensal or a eventual pathogen to captive neotropical feline, and that procedures, way life, and stress level on the shelter apparently had no negative repercussion over the integrity of the stomach.
  • Paralytic rabies in swine Veterinary Microbiology

    Pessoa, Clarice Ricardo de Macedo; Silva, Maria Luana Cristiny Rodrigues; Gomes, Albério Antônio de Barros; Garcia, Andrea Isabel Estévez; Ito, Fumio Honma; Brandão, Paulo Eduardo; Riet-Correa, Franklin

    Abstract in English:

    Rabies transmitted by vampire bats was diagnosed in pigs with paralysis of the pelvic limbs. Diffuse nonsuppurative encephalomyelitis, affecting mainly the spinal cord, was observed histologically. Despite the various diagnosis of rabies in pigs this is the first report of clinical signs and pathology of rabies transmitted by vampire bats.
  • Diagnostic performance of an immunoassay for simultaneous detection of HCV core antigen and antibodies among haemodialysis patients Genetics And Molecular Microbiology

    El-Emshaty, Wafaa M; Raafat, Douaa; Elghannam, Doaa M; Saudy, Niveen; Eltoraby, Ehab E; Metwalli, Abd Elhameed A

    Abstract in English:

    Nosocomial transmission of HCV is a concern in haemodialysis (HD) units worldwide. Diagnosis of HCV infection among dialysis patients is currently based on the detection of anti HCV antibodies by ELISA, and is confirmed by HCV RNA. The average window period between HCV infection and seroconversion with new generations of HCV antibody tests remains approximately 70 days with more prolonged period among dialysis patients. In this study we assessed the diagnostic performance of an immunoassay designed for simultaneous detection of anti HCV antibodies and core antigen in one step in comparison to qualitative RT-PCR and anti HCV antibodies detection test among Egyptian haemodialysis patients. The studied patients were 39 chronic renal failure patients on maintenance haemodialysis. The results obtained in the present study revealed HCV infection of 56.4%. Combined Ag/Ab test detected 3 out of the 4 anti-HCV negative viraemic patients who were in the window period. The sensitivity, specificity and accuracy of the test were higher than that of anti HCV antibodies detection test (95.45%, 94.1% and 94.87% versus 81.8%, 88.23% and 84.6%) and they were raised to 100% on combining its positivity with liver enzymes elevation results. Therefore, this simple combined Ag/Ab test can be applied for early detection of HCV infection during window period among HD patients as an alternative to HCV RNA detection.
  • Human T-Lymphotropic virus (HTLV) type I in vivo integration in oral keratinocytes Genetics And Molecular Microbiology

    Domínguez, Martha C; González, Norma Enith; Sánchez, Adalberto; García Vallejo, Felipe

    Abstract in English:

    Although the infection of HTLV-1 to cell components of the mouth have been previously reported, there was not until this report, a detailed study to show the characteristics of such infection. From 14 Tropical Spastic Paraparesis/ HTLV-1-Associated Myelopathy (HAM/TSP) patients and 11 asymptomatic carrier individuals (AC) coming from HTLV-1 endemic areas of southwest Pacific of Colombia, infected oral mucosa cells were primary cultured during five days. These cell cultures were immunophenotyped by dual color fluorescence cell assortment using different lymphocyte CD markers and also were immunohistochemically processed using a polyclonal anti-keratin antibody. Five days old primary cultures were characterized as oral keratinocytes, whose phenotype was CD3- /CD4-/CD8-/CD19-/CD14-/CD45-/A575-keratin+. From DNA extracted of primary cultures LTR, pol, env and tax HTLV-1 proviral DNA regions were differentially amplified by PCR showing proviral integration. Using poly A+ RNA obtained of these primary cultures, we amplify by RT-PCR cDNA of tax and pol in 57.14% (8/14) HAM/TSP patients and 27.28% (3/11) AC. Tax and pol poly A+ RNA were expressed only in those sIgA positive subjects. Our results showed that proviral integration and viral gene expression in oral keratinocytes are associated with a HTLV-1 specific local mucosal immune response only in those HTLV-1 infected individuals with detectable levels of sIgA in their oral fluids. Altogether the results gave strong evidence that oral mucosa infection would be parte of the systemic spreading of HTLV-1 infection.
  • Evaluation of a nested-pcr for Mycobacterium tuberculosis detection in blood and urine samples Genetics And Molecular Microbiology

    Cruz, Heidi Lacerda Alves da; Montenegro, Rosana de Albuquerque; Lima, Juliana Falcão de Araújo; Poroca, Diogo da Rocha; Lima, Juliana Figueirêdo da Costa; Montenegro, Lílian Maria Lapa; Crovella, Sergio; Schindler, Haiana Charifker

    Abstract in English:

    The polymerase chain reaction (PCR) and its variations, such as the nested-PCR, have been described as promising techniques for rapid diagnosis of tuberculosis (TB). With the aim of evaluating the usefulness of a nested-PCR method on samples of blood and urine of patients suspected of tuberculosis we analyzed 192 clinical samples, using as a molecular target the insertion element IS6110 specific of M. tuberculosis genome. Nested-PCR method showed higher sensitivity in patients with extrapulmonary tuberculosis (47.8% and 52% in blood and urine) when compared to patients with the pulmonary form of the disease (sensitivity of 29% and 26.9% in blood and urine), regardless of the type of biological sample used. The nested-PCR is a rapid technique that, even if not showing a good sensitivity, should be considered as a helpful tool especially in the extrapulmonary cases or in cases where confirmatory diagnosis is quite difficult to be achieved by routine methods. The performance of PCR-based techniques should be considered and tested in future works on other types of biological specimens besides sputum, like blood and urine, readily obtainable in most cases. The improving of M. tuberculosis nested-PCR detection in TB affected patients will give the possibility of an earlier detection of bacilli thus interrupting the transmission chain of the disease.
  • Lactobacillus acidophilus-derived biosurfactant effect on gtfB and gtfC expression level in Streptococcus mutans biofilm cells Genetics And Molecular Microbiology

    Tahmourespour, Arezoo; Salehi, Rasoul; Kermanshahi, Rooha Kasra

    Abstract in English:

    Streptococcus mutans (S. mutans), harboring biofilm formation, considered as a main aetiological factor of dental caries. Gtf genes play an important role in S. mutans biofilm formation. The purpose of this study was to investigate the effect of Lactobacillus acidophilus-derived biosurfactant on S. mutans biofilm formation and gtfB/C expression level (S. mutans standard strain ATCC35668 and isolated S. mutans strain (22) from dental plaque). The Lactobacillus acidophilus (L. acidophilus) DSM 20079 was selected as a probiotic strain to produce biosurfactant. The FTIR analysis of its biosurfactant showed that it appears to have a protein-like component. Due to the release of such biosurfactants, L. acidophilus was able to interfere in the adhesion and biofilm formation of the S. mutans to glass slide. It also could make streptococcal chains shorter. Using realtime RT-PCR quantitation method made it clear that gtfB and gtfC gene expression were decreased in the presence of L. acidophilus-derived biosurfactant fraction. Several properties of S. mutans cells (the surface properties, biofilm formation, adhesion ability and gene expression) were changed after L. acidophilus-derived biosurfactant treatment. It is also concluded that biosurfacant treatment can provide an optional way to control biofilm development. On the basis of our findings, we can suggest that the prepared biosurfactant may interfere with adhesion processes of S. mutans to teeth surfaces, provided additional evaluation produce satisfactory results.
  • Structural analysis of human respiratory syncytial virus P protein: identification of intrinsically disordered domains Genetics And Molecular Microbiology

    Simabuco, Fernando M; Asara, John M; Guerrero, Manuel C; Libermann, Towia A; Zerbini, Luiz F; Ventura, Armando M

    Abstract in English:

    Human Respiratory Syncytial Virus P protein plus the viral RNA, N and L viral proteins, constitute the viral replication complex. In this report we describe that HRSV P protein has putative intrinsically disordered domains predicted by in silico methods. These two domains, located at the amino and caboxi terminus, were identified by mass spectrometry analysis of peptides obtained from degradation fragments observed in purified P protein expressed in bacteria. The degradation is not occurring at the central oligomerization domain, since we also demonstrate that the purified fragments are able to oligomerize, similarly to the protein expressed in cells infected by HRSV. Disordered domains can play a role in protein interaction, and the present data contribute to the comprehension of HRSV P protein interactions in the viral replication complex.
  • Vegetative compatibility and heterokaryon formation between different isolates of Colletotrichum lindemuthianum by using the nit mutant system Genetics And Molecular Microbiology

    Carvalho, Camila Rodrigues de; Mendes-Costa, Maria Cristina

    Abstract in English:

    Colletotrichum lindemuthianum, the causative agent of bean anthracnose, is one of the most common pathogens leading to expressive damage to plants beyond presenting noticeable variability. The knowledge on vegetative compatibility groups (VCGs) is of particular interest in asexual fungi as they subdivide the population in groups that can exchange genetic information via heterokaryosis and the parasexual cycle. Among the techniques used in studies about vegetative compatibility groups, the obtainment of nit mutants is apparent. This paper is aimed at obtaining heterokaryons between different isolates of C. lindemuthianum, grouping them in VCGs and evaluating their genetic variability by using the nit mutants system. Nit mutants were obtained from 20 single spore isolates. The mutants were phenotypically classified and paired for complementation and formation of heterokaryons so as to group them in VCGs. Seventeen mutants from the different phenotypic-rates were recovered: nit1, nit2, nit3 and nitM. At the same time, 10 mutants were selected for pairing and division of the anastomosis groups. Nine heterokaryons were obtained and the isolates were divided into 9 vegetative compatibility groups. In the combinations for the formation of anastomosis, 31 compatible combinations and 24 incompatible combinations were observed. It was concluded that the methodology used to select nit mutants in C. lindemuthianum made it possible to determine the vegetative compatibility groups and that such a technique was adequate to prove genetic variability.
  • Biological activities of a mixture of biosurfactant from Bacillus subtilis and alkaline lipase from Fusarium oxysporum Industrial Microbiology

    Quadros, Cedenir Pereira de; Duarte, Marta Cristina Teixeira; Pastore, Gláucia Maria

    Abstract in English:

    In this study, we investigate the antimicrobial effects of a mixture of a biosurfactant from Bacillus subtilis and an alkaline lipase from Fusarium oxysporum (AL/BS mix) on several types of microorganisms, as well as their abilities to remove Listeria innocua ATCC 33093 biofilm from stainless steel coupons. The AL/BS mix had a surface tension of around 30 mN.m-1, indicating that the presence of alkaline lipase did not interfere in the surface activity properties of the tensoactive component. The antimicrobial activity of the AL/BS mix was determined by minimum inhibitory concentration (MIC) micro-assays. Among all the tested organisms, the presence of the mixture only affected the growth of B. subtilis CCT 2576, B. cereus ATCC 10876 and L. innocua. The most sensitive microorganism was B. cereus (MIC 0.013 mg.mL-1). In addition, the effect of the sanitizer against L. innocua attached to stainless steel coupons was determined by plate count after vortexing. The results showed that the presence of the AL/BS mix improved the removal of adhered cells relative to treatment done without the sanitizer, reducing the count of viable cells by 1.72 log CFU.cm-2. However, there was no significant difference between the sanitizers tested and an SDS detergent standard (p<0.05).
  • Growth and content of Spirulina platensis biomass chlorophyll cultivated at different values of light intensity and temperature using different nitrogen sources Industrial Microbiology

    Danesi, Eliane Dalva Godoy; Rangel-Yagui, Carlota Oliveira; Sato, Sunao; Carvalho, João Carlos Monteiro de

    Abstract in English:

    The effects of light intensity and temperature in S. platensis cultivation with potassium nitrate or urea as nitrogen source were investigated, as well as the biomass chlorophyll contents of this cyanobacteria, through the Response Surface Methodology. Experiments were performed at temperatures from 25 to 34.5ºC and light intensities from 15 to 69 µmol photons m-2 s-1, in mineral medium. In cultivations with both sources of nitrogen, KNO3 and urea, statistic evaluation through multiple regression, no interactions of such independent variables were detected in the results of the dependent variables maximum cell concentration, chlorophyll biomass contents, cell and chlorophyll productivities, as well as in the nitrogen-cell conversion factor. In cultivation performed with both sources of nitrogen, it was possible to obtain satisfactory adjustments to relate the dependent variables to the independent variables. The best results were achieved at temperature of 30ºC, at light intensity of 60 µmol photons m-2s-1, for cell growth, with cell productivity of approximately 95 mg L-1 d-1 in cultivations with urea. For the chlorophyll biomass content, the most adequate light intensity was 24 µmol photons m-2 s-1.
  • Tannase production by Penicillium atramentosum KM under SSF and its applications in wine clarification and tea cream solubilization Industrial Microbiology

    Selwal, Manjit K; Yadav, Anita; Selwal, Krishan K; Aggarwal, N.K; Gupta, Ranjan; Gautam, S. K

    Abstract in English:

    Tannin acyl hydrolase commonly known as tannase is an industrially important enzyme having a wide range of applications, so there is always a scope for novel tannase with better characteristics. A newly isolated tannase-yielding fungal strain identified as Penicillium atramentosum KM was used for tannase production under solid-state fermentation (SSF) using different agro residues like amla (Phyllanthus emblica), ber (Zyzyphus mauritiana), jamun (Syzygium cumini), Jamoa (Eugenia cuspidate) and keekar (Acacia nilotica) leaves. Among these substrates, maximal extracellular tannase production i.e. 170.75 U/gds and 165.56 U/gds was obtained with jamun and keekar leaves respectively at 28ºC after 96 h. A substrate to distilled water ratio of 1:2 (w/v) was found to be the best for tannase production. Supplementation of sodium nitrate (NaNO3) as nitrogen source had enhanced tannase production both in jamun and keekar leaves. Applications of the enzyme were studied in wine clarification and tea cream solubilization. It resulted in 38.05% reduction of tannic acid content in case of jamun wine, 43.59% reduction in case of grape wine and 74% reduction in the tea extract after 3 h at 35ºC.
  • Kinetic analysis of batch ethanol acetylation in isothermal non-stationary multiphase systems by lyophilized mycelium of Aspergillus oryzae Industrial Microbiology

    Palazzi, Emilio; Molinari, Francesco; Fabiano, Bruno; Pessoa Jr, Adalberto; Converti, Attilio

    Abstract in English:

    A relatively complex network of reactions has been investigated, using as a network model the isothermal batch esterification of acetic acid with ethanol in n-heptane catalyzed by lyophilized mycelium of Aspergillus oryzae. The kinetic analysis was firstly carried out on the whole system, without any simplification, by means of the well-known integral method. Owing to the poor results obtained by this way, we developed an alternative approach, combining initial rates and integral analysis and reducing the number of empirical parameters to be determined by the use of equilibrium data. All the values of the parameters calculated according to this "composite" approach to kinetic analysis well correlate with experimental data.
  • Citric acid production from orange peel wastes by solid-state fermentation Industrial Microbiology

    Torrado, Ana María; Cortés, Sandra; Salgado, José Manuel; Max, Belén; Rodríguez, Noelia; Bibbins, Belinda P; Converti, Attilio; Domínguez, José Manuel

    Abstract in English:

    Valencia orange (Citrus sinensis) peel was employed in this work as raw material for the production of citric acid (CA) by solid-state fermentation (SSF) of Aspergillus niger CECT-2090 (ATCC 9142, NRRL 599) in Erlenmeyer flasks. To investigate the effects of the main operating variables, the inoculum concentration was varied in the range 0.5·10³ to 0.7·10(8) spores/g dry orange peel, the bed loading from 1.0 to 4.8 g of dry orange peel (corresponding to 35-80 % of the total volume), and the moisture content between 50 and 100 % of the maximum water retention capacity (MWRC) of the material. Moreover, additional experiments were done adding methanol or water in different proportions and ways. The optimal conditions for CA production revealed to be an inoculum of 0.5·10(6) spores/g dry orange peel, a bed loading of 1.0 g of dry orange peel, and a humidification pattern of 70 % MWRC at the beginning of the incubation with posterior addition of 0.12 mL H2O/g dry orange peel (corresponding to 3.3 % of the MWRC) every 12 h starting from 62 h. The addition of methanol was detrimental for the CA production. Under these conditions, the SSF ensured an effective specific production of CA (193 mg CA/g dry orange peel), corresponding to yields of product on total initial and consumed sugars (glucose, fructose and sucrose) of 376 and 383 mg CA/g, respectively. These results, which demonstrate the viability of the CA production by SSF from orange peel without addition of other nutrients, could be of interest to possible, future industrial applications.
  • The exploration of the antibacterial mechanism of fe3+ against bacteria Microbial Physiology

    Sun, Hong-qi; Lu, Xue-mei; Gao, Pei-ji

    Abstract in English:

    This study demonstrated that the bacteria could adsorb Fe3+ and reduce Fe3+ to Fe2+. Iron had significant bacteriostatic effects, which were directly proportional to the iron concentration and under the influence of pH and chelator. It presumed that the inhibition of Fe3+ acts through the formation of hydroxyl free radicals.
Sociedade Brasileira de Microbiologia USP - ICB III - Dep. de Microbiologia, Sociedade Brasileira de Microbiologia, Av. Prof. Lineu Prestes, 2415, Cidade Universitária, 05508-900 São Paulo, SP - Brasil, Ramal USP 7979, Tel. / Fax: (55 11) 3813-9647 ou 3037-7095 - São Paulo - SP - Brazil
E-mail: bjm@sbmicrobiologia.org.br
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