A polymerase chain reaction for detection of Brucella canis in vaginal swabs of naturally infected bitches

A PCR assay for the detection of Brucella canis in canine vaginal swab samples was evaluated, comparing its performance with that of bacterial isolation, serological tests, and a blood PCR assay. One hundred and forty-four female dogs were clinically examined to detect reproductive problems and they were tested by the rapid slide agglutination test, with and without 2-mercaptoethanol (2ME-RSAT and RSAT, respectively). In addition, microbiological culture and PCR were performed on blood and vaginal swab samples. The results of the vaginal swab PCR were compared to those of the other tests using the Kappa coefficient and McNemar test. of the 144 females that were examined, 66 (45.8%) were RSAT positive, 23 (15.9%) were 2ME-RSAT positive, 49 (34.02%) were blood culture positive, 6 (4.1%) were vaginal swab culture positive, 54 (37.5%) were blood PCR positive, 52 (36.2%) were vaginal swab PCR positive, and 50.69% (73/144) were positive by the combined PCR. The PCR was able to detect as few as 3.8 fg of B. canis DNA experimentally diluted in 54 ng of canine DNA, extracted from vaginal swab samples of non-infected bitches. In addition, the PCR assay amplified B. canis genetic sequences from vaginal swab samples containing 1.0 x 10(0) cfu/mL. In conclusion, vaginal swab PCR was a good candidate as a confirmatory test for brucellosis diagnosis in bitches suspected to be infected, especially those negative on blood culture or blood PCR; these animals may be important reservoirs of infection and could complicate attempts to eradicate the disease in confined populations. (C) 2007 Elsevier B.V. All rights reserved.

Adaptation and evaluation of polymerase chain reaction for Brucella ovis detection in semen, urine and organs of rams experimentally infected

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

Genotype by environment interaction for 450-day weight of Nelore cattle analyzed by reaction norm models

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Effect of Progesterone and Ionomycin on Domestic Cat Sperm Motility Patterns and Acrosome Reaction

This study was aimed at assessing the changes in sperm motion patterns and the percentage of acrosome reaction (AR) in domestic cat semen after treatment with either ionomycin or progesterone (P(4)). Ten ejaculates were collected from five tomcats using an artificial vagina, and were diluted, centrifuged and resuspended in a capacitation medium. Samples were evaluated and divided into seven equal aliquots and, after 2 h at 25 degrees C, were incubated for 30 min at 38 degrees C in 5% CO(2) and then analyzed. Computer-assisted sperm analysis and a combination of three fluorescent probes were used to assess sperm plasma, acrosomal membrane integrity and mitochondrial transmembrane potential. Thirty minutes after the start of incubation, P(4) was added (10 mu g/ml) to the P1 group. Groups P2 and P3 were supplemented with P(4) (10 and 20 mu g/ml, respectively) only after 2 h of incubation, and groups I1 and I2 were supplemented with ionomycin (4 and 8 mu M, respectively) 2 h after incubation. Group E was supplemented with ethanol (0.6%) at 2 h after incubation and group C received no supplementation. Ionomycin and P(4) treatments led to a hyperactivation-like sperm motion and an increase (p < 0.05) in the percentage of AR. Although a higher (p < 0.05) percentage of AR was obtained in group I2 when compared with all P(4) groups, a decrease (p < 0.05) in total and progressive motility was observed in I2 group. As I1 group was similar to I2 to induce AR without diminishing sperm motility, we can conclude that ionomycin at 4 mu M seems to be more suitable to trigger AR in domestic cat sperm.

Taenia saginata: Differential diagnosis of human taeniasis by polymerase chain reaction-restriction fragment length polymorphism assay

Speciation of Taenia in human stool is important because of their different clinical and epidemiological features. DNA analysis has recently become possible which overcomes the problems of differentiating human taeniid cestodes morphologically. In the present study, we evaluated PCR coupled to restriction fragment length polymorphism to differentiate Taenia solium from Taenia saginata eggs present in fecal samples from naturally infected patients. A different Dral-RFLP pattern: a two-band pattern (421 and 100 bp) for T saginata and a three-band pattern (234, 188, and 99 bp) for T solium was observed allowing the two species to be separated.. The lower detection limit of the PCR-RFLP using a non-infected fecal sample prepared with a given number of T saginata eggs was 34 eggs in 2 g stool sediment. The 521 bp mtDNA fragment was detected in 8 out of 12 Taenia sp. carriers (66.6%). of these, three showed a T solium pattern and five a T saginata pattern. (c) 2005 Elsevier B.V. All rights reserved.

Taenia saginata: polymerase chain reaction for taeniasis diagnosis in human fecal samples

The taeniasis-cysticercosis complex is a zoonosis of great medical and economic importance where humans play an important role as the carrier of adult stage of Taenia solium and Taenia saginata. This paper describes PCR standardization that can be applied in human fecal samples for taeniasis diagnosis. DNA extraction was achieved with DNAzol reagent, after egg disruption with glass beads. DNA prepared from fecal specimens was first purified and PCR amplified generating fragments of 170 and 600 bp. The assay described herein provides an important tool for T saginata identification in human fecal samples. (C) 2003 Elsevier B.V. (USA). All rights reserved.

Detection of Leptospira spp. from pure cultures and from experimentally contaminated bovine semen by polymerase chain reaction

Considerando a importância do sêmen na transmissão da leptospira bovina, foi realizado o presente estudo que teve como objetivo aplicar a reação em cadeia pela polimerase (PCR) para a detecção de leptospiras em sêmen bovino experimentalmente contaminado. A reação de PCR foi capaz de amplificar um fragmento de DNA específico de 330 pares de bases a partir de cultivos puros de 26 sorovares de Leptospira spp. A contaminação experimental de sêmen com Leptospira interrogans serovar hardjo revelou que a técnica de PCR conseguiu detectar 10 bactérias/ml, concentração sensivelmente mais baixa que as 1.000 bactérias/ml detectadas através do cultivo microbiológico. Os resultados observados revelam o grande potencial da reação de PCR para a detecção de Leptospira spp. em sêmen bovino, notadamente em centrais de inseminação artificial.

A novel in situ Polymerase chain reaction hybridisation assay for the direct detection of bovine herpesvirus type 5 in formalin-fixed, paraffin-embedded tissues

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

Evaluation of tissue reaction to Aroeira (Myracrodruon urundeuva) extracts: a histologic and edemogenic study

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Reaction of the lateral periodontium of dogs' teeth to contaminated and noncontaminated perforations filled with mineral trioxide aggregate

It has been shown that the mineral trioxide aggregate (MTA) used to seal lateral/furcal perforations stimulates the deposition of newly formed cementum. Nevertheless, when the site of the perforation is contaminated, the healing process might occur under less favorable conditions. This study evaluated the repair healing process of noncontaminated and contaminated lateral perforations filled with MTA and the effect of previously filling the contaminated perforations with a bactericidal agent. Thirty lateral root perforations were prepared in endodontically treated dog's teeth, thus forming 3 groups with 10 specimens each. In group 1 the perforations were immediately sealed with MTA. In group 2 the perforations were left open for 7 days and thereafter sealed with MTA. In group 3 the perforations were left open for 7 days, filled temporarily with a calcium hydroxide-based paste for 14 days, and then sealed with MTA. The animals were killed after 90 days, and the pieces were prepared for histomorphologic and histomicrobiologic evaluations. The statistical analysis showed that group 1 had significantly better repair than groups 2 (P <.05) and 3 (P <.05), which validates the superior results obtained when MTA was immediately used to seal root perforations. Groups 2 and 3 had statistically similar repair to each other (P >.05). There were a larger number of cases of complete or partial biologic seal in group 1 compared with the contaminated groups. It might be concluded that the lateral root perforations sealed with MTA after contamination presented worse repair than the noncontaminated, immediately sealed perforations. The temporary filling with a bactericidal agent (calcium hydroxide-based paste) did not improve the repair of perforations exposed to contamination, and the contaminated groups presented similar results to each other.

Rat tissue reaction to MTA FILLAPEX (R)

The aim of this study was to evaluate the rat subcutaneous tissue reaction to implanted polyethylene tubes filled with mineral trioxide aggregate (MTA) FILLAPEX (R) compared to the reaction to tubes filled with Sealapex (R) or Angelus MTA (R). These materials were placed in polyethylene tubes and implanted into the dorsal connective tissue of Wistar rats for 7, 15, 30, 60, and 90 days. The specimens were stained with hematoxylin and eosin or Von Kossa or left unstained for examination under polarized light. Qualitative and quantitative evaluations of the reaction were performed. All materials caused moderate reactions after 7 days, which decreased with time. The reactions were moderate and similar to that evoked by the control and Sealapex (R) on the 15th day. MTA FILLAPEX (R) and Angelus MTA caused mild reactions beginning after 15 days. Mineralization and granulation birefringent to polarized light were observed with all materials. It was concluded that MTA FILLAPEX (R) was biocompatible and stimulated mineralization.

Tissue reaction to Endométhasone sealer in root canal fillings short of or beyond the apical foramen

Objective: This study evaluated the response of periapical tissues to the endodontic sealer Endomethasone in root canal fillings short of or beyond the apical foramen. Material and Methods: Twenty root canals of premolars and incisors of 2 mongrel dogs were used. After coronal access and pulp extirpation, the canals were instrumented up to a size 55 K-file and the apical cemental barrier was penetrated with a size 15 K-file to obtain a main apical foramen, which was widened to a size 25 K-file. The canals were irrigated with saline at each change of file. The root canals were obturated either short of or beyond the apical foramen by the lateral condensation of gutta-percha and Endomethasone, originating 2 experimental groups: G1: Endomethasone/short of the apical foramen; G2: Endomethasone/beyond the apical foramen. The animals were killed by anesthetic overdose 90 days after endodontic treatment. The individual roots were obtained and serial histological sections were prepared for histomorphological analysis (H&E and Brown & Brenn techniques) under light microscopy. The following parameters were examined: closure of the apical foramen of the main root canal and apical opening of accessory canals, apical cementum resorption, intensity of the inflammatory infiltrate, presence of giant cells and thickness and organization of the apical periodontal ligament. Each parameter was scored 1 to 4, 1 being the best result and 4 the worst. Data were analyzed statistically by the Wilcoxon nonparametric tests (p=0.05). Results: Comparing the 2 groups, the best result (p<0.05) was obtained with root canal filling with Endomethasone short of the apical foramen but a chronic inflammatory infiltrate was present in all specimens. Conclusions: Limiting the filling material to the root canal space apically is important to determine the best treatment outcome when Endomethasone is used as sealer.

Tissue Reaction to a Triantibiotic Paste Used for Endodontic Tissue Self-regeneration of Nonvital Immature Permanent Teeth

Introduction: The endodontic regenerative procedure (ERP), which is an alternative to calcium hydroxide induced apexification, involves the use of a triple antibiotic paste (TAP) as a dressing material. The aim of this study was to evaluate the response of rat subcutaneous tissue to implanted polyethylene tubes that were filled with TAP or calcium hydroxide. Methods: Thirty rats received 2 individual implants of polyethylene tubes filled with TAP or calcium hydroxide paste (CHP) and another empty tube as a control. Thirty additional rats received 2 individual implants consisting of polyethylene tubes filled with dressing material carriers (macrogol and propylene glycol) and a sham procedure. After 7, 15, 30, 60, and 90 days, 12 animals were euthanized, and the tubes and surrounding tissue were removed and processed for histology by using glycol methacrylate and stained with hematoxylin and eosin. The histological score ranged from 0 to 3 depending on the content of inflammatory cells; the fibrous capsule was considered thin or thick, and necrosis and calcification were recorded as present or absent. The results were analyzed using the Kruskal-Wallis test. Results: Both dressing materials induced moderate reactions at 7 and 15 days. These reactions were similar to the control (P>.05) and reduced in intensity (to mild) from day 30 onward (P>.05). The carriers did not interfere with the reaction of the dressing materials. Conclusions: TAP and CHP were biocompatible over the different experimental periods examined. (J Endod 2012;38:91-94)