Spectrophotometric Readings Research Articles

Optimisation of total RNA extraction from bovine oocytes and embryos for gene expression studies and effects of cryoprotectants on total RNA extraction

Gene expression is required for understanding bovine oocytes meiotic maturation as well as the potential of embryonic development. In the present study a standardized reagent protocol for total RNA extraction was designed for bovine oocytes and embryos, which is considered specific and less expensive. For such purpose oocytes (n = 795) recovered from about 80 ovaries were divided in three groups: Group 1 modified Trizol (MTP, n = 355); Group 2 Guanidinium thiocyanate protocol (GNTC, n = 140) and Group 3 Commercial Kit protocol (CKP, n = 60). Oocytes belonging to group 1 (n = 100) and 3 (n = 20) were subjected to vitrification using two cryoprotectants 1,2 propandiol (PROH) or Dimethylsulfoxide (DMSO). The 240 remaining oocytes were divided into 3 groups in which 100 were used, in fresh, for in vitro fertilization, and 140 oocytes were vitrified using PROH (n = 70) and DMSO (n = 70) as cryoprotectants, being then fertilized in vitro after thawing. Embryos were used nine days after fertilization. Gene amplification (SDHA, (GAPDH and DNMT1) was performed in oocytes, and gene quantification (DNMT1) in in vitro produced embryos at the stage of blastocyst (n = 10). Efficiency of the extraction was further compared. The purity of all samples to different protocols ranged from 1.10 to 1.25 for GNTC protocol; from 2.05 to 2.63 for the CKP and from 1.50 to 2.11 for the developed MTP, being the last one nearest to the expected purity levels for RNA samples (1.7 to 2.0). On average, for 30 fresh oocytes, from spectrophotometer readings, total RNA concentration was 127.8 ± 9.3 ng μl(-1) for MTP, against 46.4 ± 9.5 ng μl(-1) from CKP and 476 ± 12.9 ng μl(-1) for GNTC protocol. Using the MTP to evaluate RNA in 30 vitrified/thawed oocytes, resulted in a total RNA concentration of 61.3 ± 3.3 ng μl(-1) and 40.0 μ 12.4 ng μ(-1), respectively for DMSO and PROH. Regarding total RNA concentration and purity, in blastocyst stage, more purity was observed in DMSO as compared to PROH (1.8 vs 1.2) (p < 0.05). Better results were also observed on the MTP for gene amplification when compared with the other protocols. For gene quantification, the proposed protocol quantified DNMT1 gene with PCR efficiency (0.933) after normalization against GAPDH and SDHA. Amplification and quantification of genes proved specificity and efficiency of the MTP over the other protocols.

Determination of loratadine in pharmaceuticals by a spectrophotometric method

Abstract The spectrophotometric method for determination of loratadine using tetraiodomercurate has been applied in various pharmaceutical formulations. The results confirmed that recovery value is optimum and the method is valid, thus it can be used in quality control and evaluation of loratadine tablets, oral formulations of mixed composition, oral solutions, etc. The method is easy and simple to apply, does not require complicated equipment and spectrophotometric reading time is reduced, which allows a large number of analyzes in a relatively short time.

Effects of Heat Shock and 2,4-D Treatment on Morphological and Physiological Characteristics of Microspores and Microspore-Derived Doubled Haploid Plants in Brassica napus L.

Stresses such as heat shock, starvation, or osmotic is essential to lead isolated microspores towards embryogenesis. Despite the effectiveness of stresses in embryogenesis, they exert adverse effects on metabolism and growth of the regenerated plants. The effects of heat shock and 2,4-D treatment on total protein content of treated microspores, morphological and physiological characteristics of the doubled haploid (DH) plants were assessed. Buds containing mid- to late- uninucleate microspores were used for microspore culture. Microspores were isolated and cultured in NLN-13 medium and incubated at 30ºC for 14 days or treated with 2,4-D (35 mg.L-1) for 30 min to induce embryogenesis. Microspore-derived embryos were transferred onto B5 medium for plantlet regeneration. Ploidy level of the regenerated plantlets was determined using Partec flow cytometry. Spectrophotometric readings were carried out at 490, 663 and 645 nm to determine Chl a-b and carotenoids contents. TRIzol and cetyl-threeethyl-ammonium bromide (CTAB) were used for protein extraction from microspores and leaves. Length and width of stomata and pollen grains were also photographed using light microscope (Olympus). Applied stressors significantly reduced total protein content of treated microspores however, protein content and concentration of chlorophyll a and b of the DH plants were only increased by heat shock treatment when compared with the donor plant 'Hyola 420'. In contrast, carotenoids were not affected by applied stressors. Longer and wider stomata were observed by 2,4-D treatment but, the length of pollen grains was significantly decreased following heat shock and 2,4-D treatment. Total protein content of cultured microspores, concentration of chlorophyll a and b, length and width of stomata of microspore-derived doubled haploid plants were significantly affected by the type of inductive stresses. However, carotenoids were more stable and not affected by applied stressors.

AN EVALUATION ON THE GENETIC PROFILING AND MICROBIAL DETECTION IN PRAWNS

The shrimps of India belong to three major families namely: Pedacidac, Palacmonidae and Sergestidae of the decapods group. They are especially rich in niacin, essential for a healthy skin and for the release of energy in the body and vitamin B complex needed for metabolic processes. The study aims at isolating the gut microorganisms such as vibriocholerae and Staphylococcus aureus from P. monodon and A. indicus .Vibrio were isolated by streak plate methods and staphylococcus were isolated by spread plate methods. There microorganisms can cause several diseases to man and including food poisoning .Staphylococcus aureus cause diarrhoea, increase dehydration, etc., while vibriocholerae causes cholera, renal failure, cramps, hypertension , sunken eyes, etc. CTAB extraction method was used for the isolation of genomic DNA. The isolated DNA samples were subjected to agrose gel electrophoresis .The buffer used here was 1xTAE and the DNA bands were observed in UV transilluminator. Spectrophotometric readings of the isolated DNA samples were taken at 260 and 280 nm and these readings were used to estimate the quantity and quality of DNA samples isolated. Prawns are excellent source of protein, a good source of omega 3 fatty acids and a great way to get iron, zinc and vitamin E

Ability of Fungi, Isolated from Nsukka Peppers and Garden-Egg Plant Rhizospheres, to Solubilize Phosphate and Tolerate Cadmium

Fungi were isolated from the rhizosphere of pepper and garden-egg plants, with the aim of solubilizing the insoluble phosphate, identifying the most potent isolate from each rhizosphere and testing their tolerance to cadmium metal. The fungi isolates were obtained from the rhizosphere of pepper and garden-egg plants, using PVK agar and NBRIP-BPB broth culture to determine the quantity of phosphate solubilized. From the spectrophotometric readings, isolates GF1 (Penicillium spp) and PF7 (Aspergillus niger) were identified as most potent isolates. Various concentrations of cadmium (100, 50, 10 and 1 μg/ml) were added to the broth containing different isolates for tolerance determination and a control group for different isolates were also obtained. At four-day interval, a quantity of phosphates solubilized at different concentrations of cadmium and control were recorded. The quantity solubilized in 100, 50, 10 and 1 μg/ml treatment groups increased progressively from Day 4 to Day 12 in both isolates, with a sharp increase observed in isolate GF1. The tolerance of GF1 to cadmium showed that there was no significant difference (P two isolates could be used for phosphate solubilization in cadmium metal environment though GF1 tolerates more than PF7, and could be employed for bioremediation of cadmium heavy metals.

Real-time detection of histone deacetylase activity with a small molecule fluorescent and spectrophotometric probe.

Histone deacetylases (HDACs) are central players in transcription regulation and important targets in cancer treatment. Activity assays are critical tools for the study of the function and regulation of these enzymes, as well as for the screening of potential inhibitors. We report a small-molecule probe for single-step, continuous detection of deacetylase activity based on an acetyl-lysine mimic functionalized with an amine-reactive fluorophore, designed to undergo rapid intramolecular imine formation upon deacetylation. The probe exhibits a bathochromic shift in the absorption spectrum and changes in fluorescence emission intensity that enable unprecedented real-time detection of HDAC activity of purified enzymes or in cell lysates, and offers a means to evaluate HDAC inhibitors via simple spectrophotometric or fluorescence readings without the need of additional reagents.

Hypoxia-inducible Factor-1α (HIF-1α) Protein Diminishes Sodium Glucose Transport 1 (SGLT1) and SGLT2 Protein Expression in Renal Epithelial Tubular Cells (LLC-PK1) under Hypoxia

In this work, we demonstrated the regulation of glucose transporters by hypoxia inducible factor-1α (HIF-1α) activation in renal epithelial cells. LLC-PK1 monolayers were incubated for 1, 3, 6, or 12 h with 0% or 5% O2 or 300 μm cobalt (CoCl2). We evaluated the effects of hypoxia on the mRNA and protein expression of HIF-1α and of the glucose transporters SGLT1, SGLT2, and GLUT1. The data showed an increase in HIF-1α mRNA and protein expression under the three evaluated conditions (p < 0.05 versus t = 0). An increase in GLUT1 mRNA (12 h) and protein expression (at 3, 6, and 12 h) was observed (p < 0.05 versus t = 0). SGLT1 and SGLT2 mRNA and protein expression decreased under the three evaluated conditions (p < 0.05 versus t = 0). In conclusion, our results suggest a clear decrease in the expression of the glucose transporters SGLT1 and SGLT2 under hypoxic conditions which implies a possible correlation with increased expression of HIF-1α.

A QUICK, EFFICIENT, AND COST-EFFECTIVE METHOD FOR ISOLATING HIGH-QUALITY TOTAL RNA FROM TOMATO FRUITS, SUITABLE FOR MOLECULAR BIOLOGY STUDIES

Tomato (Solanum lycopersicum L.) is the primary model for the study of fleshy fruits, and research on this species has elucidated many aspects of fruit physiology, development, and metabolism. However, for advancing such studies at molecular biology levels, the RNA isolation from fruit tissues is often essential. The RNA isolation from tomato fruits is complicated because of the presence of high levels of polysaccharides, polyphenolics, pigments, and secondary metabolites and also the varying water content during development. Here, we present an optimized protocol for the isolation of total RNA from the fruit tissues at different developmental stages. In comparison to the previous methods described for the RNA isolation from tomato fruit, this method has the advantages that it does not involve the use of guanidine salts, lyophilizers, and commercial reagents, reduces the time and cost of extraction, overcomes the high water content problem, and promotes RNA quality by inhibiting RNA degradation and minimizing the gDNA, polyphenolic and polysaccharide contaminations. Using this method, high yields of high-purity and intact RNA samples were obtained as confirmed by the spectrophotometric readings and the electrophoresis on denaturing agarose gels. The isolated RNA was employed as a robust template for cDNA synthesis, reverse transcriptase-polymerase chain reaction (RT-PCR), and temporal gene expression analysis. The functionality of the isolated RNA was further demonstrated through cloning full-length cDNAs encoding β-galactosidase proteins by RT-PCR and sequencing.

Spectrophotometric Analysis of Crown Discoloration Induced by Various Antibiotic Pastes Used in Revascularization

IntroductionAntibiotic pastes are used for disinfection in regenerative endodontic procedures. This study evaluated the crown discoloration induced by various antibiotic pastes including the mixture of metronidazole and ciprofloxacin with minocycline, doxycycline, amoxicillin, or cefaclor. MethodsSeventy extracted bovine incisors were sectioned to obtain a standardized root length of 10 mm above the facial cementoenamel junction. After pulp tissue removal, irrigation with sodium hypochlorite and the placement of temporary filling material and cotton pellet were performed from the apical aspect. The specimens were then randomly divided into 7 groups (n = 10 for each group), and each group received the following antibiotic paste fillings: no filling (control group), calcium hydroxide, double antibiotic paste (DAP), triple antibiotic paste (TAP) with minocycline, TAP with doxycycline, TAP with amoxicillin, and TAP with cefaclor. Spectrophotometric readings were obtained on the buccal surfaces of the crown on day 1 to week 3 after filling, and the ΔE value was calculated. Data were analyzed with 2-way analysis of variance and the Tukey post hoc tests (P = .05), and the human perceptibility threshold was set to 3.7. ResultsTAP with minocycline, doxycycline, and cefaclor induced more coronal discoloration compared with the control group (P < .05). The control, calcium hydroxide, and DAP groups showed no color changes exceeding the perceptibility threshold at all time points. ConclusionsThe results indicated that all antibiotic pastes, except DAP, induced crown discoloration.

Rapid isolation of DNA from Dioscorea species suitable for PCR, restriction digestion and pathogen screening

The methods employed for DNA extraction from many plants is difficult because of the metabolites that interfere with DNA isolation procedures. We have developed a reliable and efficient method for isolating genomic DNA free from polysaccharide, polyphenols and protein contaminants from Dioscorea spp. The method involves inactivation of contaminant proteins by using CTAB/Proteinase K and precipitation of polysaccharides in the presence of high concentration of salt. The purity of genomic DNA was confirmed by A260/280 and A260/230 ratios calculated from the spectrophotometric readings and further by restriction analysis of the isolated DNA using restriction enzymes Eco RI. The total genomic DNA extracted by the new protocol was used for polymerase chain reaction amplification, RAPD analysis, restriction digestion and pathogen screening. The new protocol can be successfully used for both small- and large-scale preparation of genomic DNA from different tissues of Dioscorea spp. The quarantine of seed tubers and use of pathogen-free tubers for planting is a prerequisite for integrated disease management strategy. The protocol can be used for the isolation of genomic DNA from other crop plants too.

Comparative effectiveness of nonpurpuragenic 595-nm pulsed dye laser and microsecond 1064-nm neodymium:yttrium-aluminum-garnet laser for treatment of diffuse facial erythema: A double-blind randomized controlled trial

Facial erythema is a common symptom that responds to vascular laser treatment, but there are few comparative studies. We sought to compare the effectiveness of microsecond 1064-nm neodymium:yttrium-aluminum-garnet (Nd:YAG) laser with nonpurpuragenic 595-nm pulsed dye laser (PDL) for diffuse facial erythema. This was a split-face, double-blind randomized controlled trial. Bilateral cheeks received 4 treatments each at one month intervals with PDL or Nd:YAG. Spectrophotometer measurements, digital photographs, pain scores, and patient preferences were recorded. Sixteen patients enrolled and 2 dropped out. Fourteen patients, all skin types I to III, 57% women, mean age 42 years, completed the study and were analyzed. Spectrophotometer readings changed after both PDL (8.9%) and Nd:YAG (2.5%), but varied by treatment type, with PDL reducing facial redness 6.4% more from baseline than Nd:YAG (P = .0199; 95% confidence interval -11.6 to -1.2). Pain varied (P = .0028), with Nd:YAG associated with less pain, at 3.07, than PDL at 3.87. Subjects rated redness as improved by 52% as a result of PDL, and 34% as a result of Nd:YAG (P = .031; 95% confidence interval -34.6 to -1.94). No serious adverse events were observed. Lasers settings are not standardized across devices. Facial erythema is safely and effectively treated with PDL and Nd:YAG. Nonpupuragenic PDL may be more effective for lighter-skinned patients, but microsecond Nd:YAG may be less painful.

A high-throughput seed germination assay for root parasitic plants

BackgroundSome root-parasitic plants belonging to the Orobanche, Phelipanche or Striga genus represent one of the most destructive and intractable weed problems to agricultural production in both developed and developing countries. Compared with most of the other weeds, parasitic weeds are difficult to control by conventional methods because of their life style. The main difficulties that currently limit the development of successful control methods are the ability of the parasite to produce a tremendous number of tiny seeds that may remain viable in the soil for more than 15 years. Seed germination requires induction by stimulants present in root exudates of host plants. Researches performed on these minute seeds are until now tedious and time-consuming because germination rate is usually evaluated in Petri-dish by counting germinated seeds under a binocular microscope.ResultsWe developed an easy and fast method for germination rate determination based on a standardized 96-well plate test coupled with spectrophotometric reading of tetrazolium salt (MTT) reduction. We adapted the Mosmann’s protocol for cell cultures to germinating seeds and determined the conditions of seed stimulation and germination, MTT staining and formazan salt solubilization required to obtain a linear relationship between absorbance and germination rate. Dose–response analyses were presented as applications of interest for assessing half maximal effective or inhibitory concentrations of germination stimulants (strigolactones) or inhibitors (ABA), respectively, using four parameter logistic curves.ConclusionThe developed MTT system is simple and accurate. It yields reproducible results for germination bioassays of parasitic plant seeds. This method is adapted to high-throughput screenings of allelochemicals (stimulants, inhibitors) or biological extracts on parasitic plant seed germination, and strengthens the investigations of distinctive features of parasitic plant germination.

280 OSTEOGENIC ACTIVITY OF IN HOUSE-PRODUCED PORCINE BMP2 ON ADIPOSE-DERIVED STEM CELLS

In our laboratory, we extensively study the possibility of using adipose-derived stem cells (ASC) for maxillofacial bone regeneration. This includes also the tissue repair of large critical-size osteotomies requiring the use of tridimensional scaffolds. Bone regeneration in scaffolds can be greatly enhanced by the use of specific growth factors such as BMP2. In the present study, we compared the activity of commercially available human BMP2 (hBMP2) with in house-produced porcine BMP2 (pBMP2). The latter was synthesised using the BMP2 coding sequence from mRNA obtained from porcine ASC cell cultures. The coding sequence of the mature protein was cloned into a pET-21 plasmid and produced in E. coli as inclusion bodies. The activity of pBMP2 and hBMP2 was tested on ASC isolated from male pigs at passage 4 and at approximately 80% confluence in 48-well plates. Cells were treated in triplicate with hBMP2 or pBMP2 at 0.5, 5, 50, 500, or 1000 ng mL–1, adipogenic medium (AM), osteogenic medium (OM), or normal DMEM medium supplemented with acetic acid (used to resuspend BMP2 as the control) for 5 or 17 days. Cells were harvested for Alizarin Red S (AR) quantification and expression of osteogenic genes. For the AR analysis, cells were fixed with formalin and treated with AR. The AR was then extracted by acetic acid and neutralized with ammonium hydroxide before spectrophotometer reading at an absorbance of 420 nm. Data were analysed using GLM of SAS (SAS Institute Inc., Cary, NC, USA) with treatment, time, concentration, and all interactions as main effects. Using an inverted robotic stage microscope, images of the entire well for each replicate were taken every 2 to 3 days. Images revealed formation of osteogenic nodules in OM and characteristic large cells filled with lipid droplets in AM. No evident nodule formation was observed in the other treated cells at any time point. The AR was higher than control in both hBMP2 and pBMP2 at 0.5, 50, and 1000 ng mL–1 but not at 5 and 500 ng mL–1. There was no overall difference between hBMP2 and pBMP2 but the former had the highest AR value at 5 days in cells treated with 0.5 ng mL–1 and pBMP2 at 17 days with 1000 ng mL–1. Interestingly, both had higher values compared to OM, particularly at 5 days. We also observed an increase of AR due to time in cells treated with acetic acid (control). Overall, the data appear to indicate an increase in calcium accumulation in cells treated with both hBMP2 and pBMP2, with an early increase in the former and a late and larger increase in the latter. This might indicate a larger but slower activity of pBMP2 compared with hBMP2. The lack of formation of osteogenic nodules by both BMP2 might indicate an insufficiency of BMP2 to induce osteogenesis in porcine ASC. This last observation, together with the lack of increased AR accumulation compared with control at the 5 and 50 ng mL–1 doses, suggests the need for a more accurate analysis of BMP2 activity by measuring expression of BMP2-related genes. Finally, the data provide preliminary support for the equivalency of activity of pBMP2 and hBMP2 for in vivo bone regeneration.

Developing methods to compare tablet formulations of atorvastatin

Atorvastatin (ATV) is an antilipemic drug of great interest to the pharmaceutical industry. ATV does not appear in the monographs of Brazilian pharmacopoeia, and analytical methodologies for its determination have been validated. The chromatographic conditions used included: RP-18 column-octadecylsilane (250 x 4.6 mm, 5 mm), detection at 238 nm, mobile phase containing 0.1% phosphoric acid and acetonitrile (35:65% v/v), flow at 1.5 mL min-1, oven temperature at 30ºC, and injection volume of 10 mL. ATV is classified as a class II product, according to the biopharmaceutical classification system. As such, a dissolution test was proposed to evaluate pharmaceutical formulations on the market today, under the following conditions: water as a dissolution medium, 1000 mL as a volume, paddle apparatus at a rotation speed of 50 rpm, 80% (Q) in 15 minutes with UV spectrophotometer readings at 238 nm. In the pattern condition proposed as the ideal dissolution test, which appropriately differentiates amongst formulations, the generic product was not considered pharmaceutically equivalent; however, in other less differential dissolution methods, which also fall within appropriate legal parameters, this product could come to be regarded as generic.

Activity of MK-3118, a new oral glucan synthase inhibitor, tested against Candida spp. by two international methods (CLSI and EUCAST)

To evaluate the activity of the orally bioavailable enfumafungin derivative MK-3118 and comparator antifungal agents tested against a collection of 113 clinical isolates of Candida spp. using CLSI and EUCAST broth microdilution (BMD) methods. Candida spp. isolates (n=113) were tested by CLSI and EUCAST methods. The collection contained 29 Candida albicans, 29 Candida glabrata, 21 Candida tropicalis, 15 Candida parapsilosis and 19 Candida krusei, including azole- and echinocandin-resistant isolates. CLSI and EUCAST MIC endpoints of 50% and 100% inhibition were determined using visual reading at 24 and 48 h of incubation and spectrophotometric reading at 24 h of incubation, respectively. MK-3118 CLSI MIC results ranged from 0.06 to 16 mg/L depending on species, duration of incubation and endpoint criteria (EC) used. Comparison of CLSI and EUCAST following 24 h of incubation and either 50% or 100% inhibition revealed an essential agreement (EA; ± 2 doubling dilutions) of 99.1% using the 50% inhibition EC and 93.2% using the 100% inhibition EC. MK-3118 (24 h of incubation and 50% EC) was active against all the species tested and displayed similar potency to caspofungin (using CLSI BMD) against C. albicans (MIC90, 1 and 2 mg/L, respectively), C. tropicalis (1 and 1 mg/L, respectively), C. parapsilosis (0.5 and 0.5 mg/L, respectively) and C. krusei (2 and 1 mg/L, respectively), but was 8-fold more potent than caspofungin against C. glabrata strains (MIC90, 2 and 16 mg/L, respectively). MK-3118 was active against fluconazole-resistant strains as well as caspofungin-resistant strains with documented fks mutations. MK-3118 was documented to have potent in vitro activity against Candida spp. when tested by both CLSI and EUCAST BMD methods, with the highest overall EA (99.1%) obtained when MK-3118 MIC results were read after 24 h of incubation using a partial inhibition EC (50%).

Lochia Patterns Among Normal Women: A Systematic Review

We conducted a systematic review of the literature to determine the amount and duration of blood loss 24 hours to 12 weeks after delivery. We searched MEDLINE, CINAHL, and PubMed for studies between the years 1950 and 2011 that prospectively evaluated the amount and duration of blood loss from 24 hours to 12 weeks after delivery. Excluded were those that were only case studies, retrospective studies, studies not published in English, studies outside of the time frame, and studies that included only subjects from special populations. From the 333 identified studies, 18 met inclusion criteria. There was variability in how the amount of blood loss was determined, ranging from subject self-assessment to objective measures, such as pad weight and spectrophotometric readings of hematin concentration. The reported duration of normal blood loss after delivery varied among the studies. Whereas the average duration of blood loss in these studies ranged from 24 to 36 days, in only 1 study was bleeding followed to cessation. An understanding of bleeding patterns after delivery is important for clinicians to recognize deviations from normal, identify women at risk for delayed postpartum hemorrhage, and limit unnecessary interventions, yet studies reveal significant variability in amount and duration of normal lochial blood loss and methods of assessment that are inconsistent. This review draws attention to the need for the establishment of valid, reliable, and feasible methods to quantify normal and abnormal postpartum blood loss.

The biofilm formation ability of Listeria monocytogenes isolated from meat, poultry, fish and processing plant environments is related to serotype and pathogenic profile of the strains

In the present study, the relationships between serotype, pathogenic profile and &lt;em&gt;in vitro&lt;/em&gt; biofilm formation of 106 &lt;em&gt;Listeria monocytogenes&lt;/em&gt; strains, having no epidemiological correlation and isolated from different environmental and food sources, were analyzed. The quantitative assessment of the &lt;em&gt;in vitro&lt;/em&gt; biofilm formation was carried out by using a microtiter plate assay with spectrophotometric reading (OD620). The isolates were also submitted to serogrouping using the target genes &lt;em&gt;lmo0737&lt;/em&gt;, &lt;em&gt;lmo1118, ORF2819, ORF2110, prs&lt;/em&gt;, and to the evaluation of the presence of the following virulence genes: &lt;em&gt;prfA, hlyA, rrn, inlA, inlB, iap, plcA, plcB, actA&lt;/em&gt; and &lt;em&gt;mpl&lt;/em&gt;, by multiplex PCRs. The 62% of the strains showed weak or moderate &lt;em&gt;in vitro&lt;/em&gt; ability in biofilm formation, in particular serotypes 1/2b and 4b, frequently associated with sporadic or epidemic listeriosis cases. The 25% of these isolates showed polymorphism for the &lt;em&gt;actA&lt;/em&gt; gene, producing a fragment of 268-bp instead of the expected 385-bp. The deletion of nucleotides in this gene seems to be related to enhanced virulence properties among these strains. Strains belonging to serotypes associated with human infections and characterized by pathogenic potential are capable to persist within the processing plants forming biofilm.

The biofilm formation ability of Listeria monocytogenes isolated from meat, poultry, fish and processing plant environments is related to serotype and pathogenic profile of the strains

In the present study, the relationships between serotype, pathogenic profile and in vitro biofilm formation of 106 Listeria monocytogenes strains, having no epidemiological correlation and isolated from different environmental and food sources, were analyzed. The quantitative assessment of the in vitro biofilm formation was carried out by using a microtiter plate assay with spectrophotometric reading (OD620). The isolates were also submitted to serogrouping using the target genes lmo0737, lmo1118, ORF2819, ORF2110, prs, and to the evaluation of the presence of the following virulence genes: prfA, hlyA, rrn, inlA, inlB, iap, plcA, plcB, actA and mpl, by multiplex PCRs. The 62% of the strains showed weak or moderate in vitro ability in biofilm formation, in particular serotypes 1/2b and 4b, frequently associated with sporadic or epidemic listeriosis cases. The 25% of these isolates showed polymorphism for the actA gene, producing a fragment of 268-bp instead of the expected 385-bp. The deletion of nucleotides in this gene seems to be related to enhanced virulence properties among these strains. Strains belonging to serotypes associated with human infections and characterized by pathogenic potential are capable to persist within the processing plants forming biofilm.

Relationship between antibiotic susceptibility and biofilm production in a clinical strain of Burkholderia cepacia complex

Infections caused by Burkholderia cepacia complex in patients with cystic fibrosis (CF) are often related to increased mortality. One of the mechanisms that makes the bacteria more resistant to the action of antibiotics is the ability to produce biofilm. It has been compared the antibiotic resistance of two clinical isolates of a biofilm-producing strain called BTS-2, responsible for chronic pulmonary infection in a CF patient. Bacteria were incubated in both Yeast Extract Mannitol Medium (YEM) and Mueller Hinton Broth (MHB) at 30°C for 24 hours. Biofilm was quantified by spectrophotometer readings (OD570) of the extracted crystal violet. By microdilution broth, we evaluated the minimum concentration of antibiotic inhibiting the growth of the planktonic form (MIC); by the technology “Calgary Biofilm Device” and by Resazurin, we evaluated the susceptibility of sessile forms (BIC). Data demonstrate that the microorganism in the course of time changed its sensitivity to some of the antibiotics tested. The comparison between sessile and planktonic forms of BTS2 pre-incubated in MHB showed that sessile forms increased their resistance to antibiotics in a few cases only. BICs of BTS2 pre-incubated in YEM, where the strain produced an amount of biofilm significantly lower than in MHB, showed a higher susceptibility to 4 of the 10 antibiotics tested. Our data showed that the susceptibility of sessile forms of a strain is not always lower than the susceptibility of its planktonic forms and that culture medium can affect significantly the biofilm production.

COAL ACTIVE BABASSU ADSORPTION IN COLOUR FOR THE TREATMENT OF INDUSTRIAL WASTE

The presence of chemical residues in aquatic environment can cause noticeable adverse physiological effects in humans and animals. In view of this, one needs methods to try to control its disposal, among these methods highlight the adsorption. The adsorption on solids is a phenomenon that focus on the surface of existing substances in liquids or gases. Due to the adsorption capacity of the derivatives of babassu (Orbignya phalerata, Mart.) and its abundance geographical took place this study was conducted with the purpose of transforming the coal into a potent babassu adsorbent useful for the treatment of industrial effluents, including pharmaceutical. The sieve analysis was performed and showed charcoal powder with a mean diameter of 500.475 mM, obtaining therefore a coarse powder. Potential adsorption system was elevated. According to the spectrophotometric reading, the initial concentration of methylene blue (3,85 mg/ml) was reduced to 0.106 g/ml in the system containing activated charcoal, thus resulting in a percentage adsorption equivalent to 97%. This study noted a keen perspective on the adsorption capacity of activated carbon from babassu front of the methodology employed. This material shows great potential for use in the manufacture of filters for the decontamination of industrial waste.