Effect of Removing Seminal Plasma Using a Sperm Filter on the Viability of Refrigerated Stallion Semen

Cooling of equine semen obtained from some stallions results in lower seminal quality and viability when the seminal plasma (SP) is present. The objective of this study was to evaluate the effect of the removal of SP using a Sperm Filter on the viability of cooled stallion semen. For this purpose, 31 stallions were used. Their ejaculates were divided into three groups: CN, semen was diluted with an extender; FLT, SP was removed by filtration; and CT, SP was removed by centrifugation and cooled to 15°C for 24 hours. Sperm kinetics and plasma membrane integrity were evaluated immediately after collection (T0) and after 24 hours of refrigeration (T1). No difference (P > .05) was noted at T1 for total sperm motility (TM), progressive sperm motility, or plasma membrane integrity when semen samples from all the stallions were analyzed. However, when samples from stallions termed bad coolers were analyzed (TM = <30% at T1), a difference was observed in TM and progressive sperm motility for CN compared with FLT and CT at T1. Sperm recovery was greater when SP was removed using the filter (FLT) to that when the SP was removed by centrifugation (CN) (89% vs. 81%). Thus, we concluded that filtering with a Sperm Filter is an efficient and practical method for removal of SP from stallion ejaculates, with lower sperm loss than centrifugation. We also found that the presence of SP reduces the quality and viability of cooled semen from stallions whose semen is sensitive to the process of refrigeration. © 2013 Elsevier Inc.

Comparison of Two Staining Methods for Pollen Viability Studies in Sugarcane

The present work aimed to compare two staining methods for pollen viability evaluation in sugarcane. Pollen from four sugarcane genotypes were collected at three different times (6.00, 8.00 and 9.00 a.m.) and tested for viability using two staining methods (iodine and lactophenol blue). Three anthers, of each genotype were crushed in a glass slide with a drop of the respective stain (iodine 0.1 N and lactophenol blue). The percentage of pollen viability was obtained with an optic microscope (250×) and compared with the pollen germination at culture media where one raquis of each genotype was gentle shaken in a petridish. Three replicates (petri dishes) was performed for each genotype which were maintained at the temperature of 25 °C and air humidity around 95 % for 30 min. The factors (staining methods, genotypes and times) and their interactions were evaluated by the analysis of variance, F test (P < 0.01) and the means compared by the t test (P < 0.05). The lactophenol blue staining was more sensible than the iodine staining method to detect the decrease of pollen viability which occurs naturally in sugarcane. The iodine staining method was more stable and easier than lactophenol to perform the inflorescence classification at any evaluated time (6.00, 8.00 and 9.00 a. m.). Both staining methods overestimated the viability obtained by the germination at culture media when performed at 6.00 a.m. © 2012 Society for Sugar Research & Promotion.

Regulation of visfatin by microbial and biomechanical signals in PDL cells

Objectives: This in vitro study was established to examine whether visfatin thought to be a link between periodontitis and obesity is produced by periodontal ligament (PDL) cells and, if so, whether its synthesis is modulated by microbial and/or biomechanical signals. Materials and methods: PDL cells seeded on BioFlex® plates were exposed to the oral pathogen Fusobacterium nucleatum ATCC 25586 and/or subjected to biomechanical strain for up to 3 days. Gene expression of visfatin and toll-like receptors (TLR) 2 and 4 was analyzed by RT-PCR, visfatin protein synthesis by ELISA and immunocytochemistry, and NFκB nuclear translocation by immunofluorescence. Results: F. nucleatum upregulated the visfatin expression in a dose- and time-dependent fashion. Preincubation with neutralizing antibodies against TLR2 and TLR4 caused a significant inhibition of the F. nucleatum-upregulated visfatin expression at 1 day. F. nucleatum stimulated the NFκB nuclear translocation. Biomechanical loading reduced the stimulatory effects of F. nucleatum on visfatin expression at 1 and 3 days and also abrogated the F. nucleatum-induced NFκB nuclear translocation at 60 min. Biomechanical loading inhibited significantly the expression of TLR2 and TLR4 at 3 days. The regulatory effects of F. nucleatum and/or biomechanical loading on visfatin expression were also observed at protein level. Conclusions: PDL cells produce visfatin, and this production is enhanced by F. nucleatum. Biomechanical loading seems to be protective against the effects of F. nucleatum on visfatin expression. Clinical relevance: Visfatin produced by periodontal tissues could play a major role in the pathogenesis of periodontitis and the interactions with obesity and other systemic diseases. © 2013 Springer-Verlag Berlin Heidelberg.

Effect of Storage Time and Temperature of Equine Epididymis on the Viability, Motion Parameters, and Freezability of Epididymal Sperm

The recovery of sperm from the epididymal cauda may be the last chance to obtain genetic material when sudden death or serious injuries occur in valuable stallions. However, the lack of technical knowledge regarding the storage and transportation of the epididymis often prevents the preservation of the sperm. Therefore, the aim of this study was to compare sperm parameters of sperm obtained immediately after orchiectomy with sperm recovered from epididymal cauda at different times after storage at 5°C and at room temperature (RT). For that, 48 stallions of different breeds were used. In group 1 (control group), eight stallions were used, and the harvest of the epididymal sperm was performed immediately after orchiectomy. In group 2, 40 stallions were used, which were divided into five groups according to the storage time of the epididymis after orchiectomy (6, 12, 18, 24, or 30 hours), making a total of eight stallions per group. One epididymis of each stallion was stored at 5°C, and the contralateral epididymis was stored at RT, both for the same period. The sperm parameters of total motility, progressive motility, progressive linear velocity, curvilinear velocity, percentage of rapid sperm, and plasma membrane integrity were evaluated in all the groups after sperm recovery, resuspension in a sperm freezing diluent, and thawing. In conclusion, the storage of the testis-epididymis complex at 5°C provided better preservation of epididymal sperm than the storage at RT, and regardless of the temperature, the progressive motility is the sperm parameter that is most sensitive to storage time. © 2013 Elsevier Inc.

Guided implant surgery: What is the influence of this new technique on bone cell viability?

Purpose: To evaluate the effect of implant osteotomy on immediate bone cell viability, comparing guided surgery for implant placement with the classic drilling procedure. Materials and Methods: For this study, 20 rabbits were used. The animals were divided into a guided surgery group (GG) and a control group (CG) and were then divided into 4 subgroups - subgroups 1, 2, 3, and 4 - corresponding to drills used 10, 20, 30, and 40 times, respectively. All animals received 5 osteotomies in each tibia, by use of the classic drilling procedure in one tibia and guided surgery in the other tibia. The osteotomized areas were removed and processed immunohistochemically for detection of osteocalcin, receptor activator of nuclear factor κB ligand (RANKL), osteoprotegerin (OPG), and caspase 3. Results: Immunohistochemical analysis showed that osteocalcin expression was initially higher in the CG and remained constant after drill reutilization. Although the expressions of RANKL and OPG were not statistically different for the GG and CG, the RANKL/OPG ratio tended to be higher for the GG. Moreover, caspase 3 expression was elevated in the GG, proportionally to the number of osteotomies, indicating an increase in the apoptosis index in the GG. Conclusions: The classic drilling procedure is more favorable to cell viability than guided surgery.© 2013 American Association of Oral and Maxillofacial Surgeons.

Photodynamic inactivation of Streptococcus mutans and Streptococcus sanguinis biofilms in vitro

The purpose of this study was to evaluate specific effects of photodynamic inactivation (PDI) using erythrosine (ER) and Rose Bengal (RB) photosensitizers and a blue light-emitting diode (LED) on the viability of Streptococcus mutans and Streptococcus sanguinis biofilms. Biofilms were grown in acrylic disks immersed in broth to production of biofilms, inoculated with microbial suspension (106 cells/mL) and incubated for 48 h. After the formation of biofilms, the effects of the photosensitizers ER and RB at a concentration of 5 μM for 5 min and blue LED (455 ± 20 nm) for 180 s, photosensitizers alone and conjugated were evaluated. Next, the disks were placed in tubes with sterile physiological solution (0.9 % sodium chloride) and sonicated for to disperse the biofilms. Tenfold serial dilutions were carried and aliquots seeded in brain heart infusion agar which were then incubated for 48 h. Then the numbers colony-forming units per milliliter (CFU/mL; log 10) were counted and analyzed statistically (ANOVA, Tukey test, P ≤ 0.05). Significant decreases in the viability of all microorganisms were observed for biofilms exposed to PDI mediated by both photosensitizers. The reductions with RB and ER were, 0.62 and 0.52 log10 CFU mL -1 for S. mutans biofilms (p = 0.001), and 0.95 and 0.88 log 10 CFU mL-1 for S. sanguinis biofilms (p = 0.001), respectively. The results showed that biofilms formed in vitro by S. mutans and S. sanguinis, were sensitive to PDI using a blue LED associated with photosensitizers ER or RB, indicating its use in the control of caries and periodontal diseases. © 2012 Springer-Verlag London Ltd.

Spray Nozzles, Pressures, Additives and Stirring Time on Viability and Pathogenicity of Entomopathogenic Nematodes (Nematoda: Rhabditida) for Greenhouses

The objective of this study was to evaluate different strategies for the application of entomopathogenic nematodes (EPN). Three different models of spray nozzles with air induction (AI 11003, TTI 11003 and AD-IA 11004), three spray pressures (207, 413 and 720 kPa), four different additives for tank mixtures (cane molasses, mineral oil, vegetable oil and glycerin) and the influence of tank mixture stirring time were all evaluated for their effect on EPN (Steinernema feltiae) viability and pathogenicity. The different nozzles, at pressures of up to 620 kPa, were found to be compatible with S. feltiae. Vegetable oil, mineral oil and molasses were found to be compatible adjuvants for S. feltiae, and stirring in a motorized backpack sprayer for 30 minutes did not impact the viability or pathogenicity of this nematode. Appropriate techniques for the application of nematodes with backpack sprayers are discussed. © 2013 Moreira et al.

Performance of laying hens and economic viability of different climatization systems

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Photodynamic inactivation of biofilm: Taking a lightly colored approach to stubborn infection

Microbial biofilms are responsible for a variety of microbial infections in different parts of the body, such as urinary tract infections, catheter infections, middle-ear infections, gingivitis, caries, periodontitis, orthopedic implants, and so on. The microbial biofilm cells have properties and gene expression patterns distinct from planktonic cells, including phenotypic variations in enzymic activity, cell wall composition and surface structure, which increase the resistance to antibiotics and other antimicrobial treatments. There is consequently an urgent need for new approaches to attack biofilm-associated microorganisms, and antimicrobial photodynamic therapy (aPDT) may be a promising candidate. aPDT involves the combination of a nontoxic dye and low-intensity visible light which, in the presence of oxygen, produces cytotoxic reactive oxygen species. It has been demonstrated that many biofilms are susceptible to aPDT, particularly in dental disease. This review will focus on aspects of aPDT that are designed to increase efficiency against biofilms modalities to enhance penetration of photosensitizer into biofilm, and a combination of aPDT with biofilm-disrupting agents. © 2013 Informa UK Ltd.

Evaluation of the microbial diversity of denitrifying bacteria in batch reactor

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Development of equations to estimate microbial contamination in ruminal incubation residues of forage produced under tropical conditions using 15N as a label1

The objective of this study was to use 15N to label microbial cells to allow development of equations for estimating the microbial contamination in ruminal in situ incubation residues of forage produced under tropical conditions. A total of 24 tropical forages were ruminal incubated in 3 steers at 3 separate times. To determine microbial contamination of the incubated residues, ruminal bacteria were labeled with 15N by continuous intraruminal infusion 60 h before the first incubation and continued until the last day of incubation. Ruminal digesta was collected for the isolation of bacteria before the first infusion of 15N on adaptation period and after the infusion of 15N on collection period. To determine the microbial contamination of CP fractions, restricted models were compared with the full model using the model identity test. A value of the corrected fraction A was estimated from the corresponding noncorrected fraction by this equation: Corrected A fraction (ACPC) = 1.99286 + 0.98256 × A fraction without correction (ACPWC). The corrected fraction B was estimated from the corresponding noncorrected fraction and from CP, NDF, neutral detergent insoluble protein (NDIP), and indigestible NDF (iNDF) using the equation corrected B fraction (BCPC) = -17.2181 - 0.0344 × fraction B without correction (BCPWC) + 0.65433 × CP + 1.03787 × NDF + 2.66010 × NDIP - 0.85979 × iNDF. The corrected degradation rate of B fraction (kd)was estimated using the equation corrected degradation rate of B fraction (kdCPC) = 0.04667 + 0.35139 × degradation rate of B fraction without correction (kdCPWC) + 0.0020 × CP - 0.00055839 × NDF - 0.00336 × NDIP + 0.00075089 × iNDF. This equation was obtained to estimate the contamination using CP of the feeds: %C = 79.21 × (1 - e-0.0555t) × e-0.0874CP. It was concluded that A and B fractions and kd of CP could be highly biased by microbial CP contamination, and therefore these corrected values could be obtained mathematically, replacing the use of microbial markers. The percentage of contamination and the corrected apparent degradability of CP could be obtained from values of CP and time of incubation for each feed, which could reduce cost and labor involved when using 15N. © 2013 American Society of Animal Science. All rights reserved.

Microbial culture collections as pillars for promoting fungal diversity, conservation and exploitation

Fungi are a diverse group of organisms with an overall global number of 1.5 M up to 3.3 M species on Earth. Besides their ecological roles as decomposers, fungi are important in several aspects of applied research. Here, we review how culture collections may promote the knowledge on diversity, conservation and biotechnological exploitation of fungi. The impact of fungi diversity on biotechnological studies is discussed. We point out the major roles of microbial repositories, including fungal preservation, prospecting, identification, authentication and supply. A survey on the World Data Center for Microorganisms (WDCM) powered by the World Federation for Culture Collections and on the Genetic Heritage Management Council (CGEN) database revealed that 46 Brazilian culture collections registered in these databases are dedicate to preserving fungi. Most of these culture collections are located in the Southeast of Brazil. This scenario also demonstrates that Brazil has many collections focused on fungal strains, but the lack of up-to-date information in WDCM as well as of a solid national platform for culture collections registration do not allow accurate assessment of fungal preservation. © 2013 Elsevier Inc. All rights reserved.

Leishmaniasis causes oxidative stress and alteration of oxidative metabolism and viability of neutrophils in dogs

The aim of the present study was to test the hypothesis that oxidative stress and alteration of oxidative metabolism and apoptosis of neutrophils in dogs vary with the stage of leishmaniasis and to determine the contribution of uremia to such alterations. Dogs with leishmaniasis were classified into two stages: moderate (Leish II, n = 20) or very severe (i.e. with concurrent uremia; Leish IV, n = 20) according to the LeishVet Consensus. The two leishmaniasis groups were compared with uremic dogs without leishmaniasis (Uremic, n = 10) and to healthy dogs (Control, n = 30). To determine oxidative stress, total antioxidant/oxidant capacity, lipid peroxidation, total glutathione and the plasma antioxidants albumin, uric acid and bilirubin were quantified. Superoxide production was determined using the hydroethidine probe and viability and apoptosis were measured using annexin V-PE by capillary flow cytometry. Oxidative stress was present in both uremia and leishmaniasis with reduced total antioxidant capacity and was associated with increased induced production of superoxide and apoptosis. The greatest amount of oxidants was observed in animals with moderate disease only. Neutrophils from uremic dogs with and without leishmaniasis had decreased viability and an increased apoptosis rate in addition to increased lipid peroxidation. In conclusion, oxidative stress occurs in both stages of leishmaniasis with differences in intensity and levels of plasma markers; however, uremia does contribute to the decreased spontaneous viability of neutrophils in dogs in the final stage of the disease. © 2013 Elsevier Ltd. All rights reserved.

Implant-abutment gap versus microbial colonization: Clinical significance based on a literature review

Microorganisms from the oral cavity may settle at the implant-abutment interface (IAI). As a result, tissue inflammation could occur around these structures. The databases MEDLINE/PubMed and PubMed Central were used to identify articles published from 1981 through 2012 related to the microbial colonization in the implant-abutment gap and its consequence in terms of crest bone loss and osseointegration. The following considerations could be put forward, with respect to the clinical importance of IAI: (a) the space present at the IAI seems to allow bacterial leakage to occur, in spite of the size of this space; (b) bacterial leakage seems to occur at the IAI, irrespective of the type of connection. More studies are necessary to clarify the relationship between leakage at IAI and abutment connection designs; (c) losses at the peri-implant bone crests cannot be related to the IAI size, since few studies have shown no relationship. Also, the microbial leakage at the IAI cannot be related to the bone crest loss, since there are no articles reporting this relationship; remains controversial the influence of the IAI position on the bone crest losses. © 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 101B: 1321-1328, 2013. Copyright © 2013 Wiley Periodicals, Inc.

Alteration in cellular viability, pro-inflammatory cytokines and nitric oxide production in nephrotoxicity generation by Amphotericin B: Involvement of PKA pathway signaling

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)