New methods for selective isolation of bacterial DNA from human clinical specimens

Separation of bacterial DNA from human DNA in clinical samples may have an important impact on downstream applications, involving microbial diagnostic systems. We evaluated two commercially available reagents (MolYsis^®, Molzym GmbH & Co. KG, Bremen and Pureprove^®, SIRS-Lab GmbH, Jena, both Germany) for their potential to isolate and purify bacterial DNA from human DNA. We chose oral samples, which usually contain very high amounts of both human and bacterial cells. Three different DNA preparations each were made from eight caries and eight periodontal specimens using the two reagents above and a conventional DNA extraction strategy as reference. Based on target-specific real-time-quantitative PCR assays we compared the reduction of human DNA versus loss of bacterial DNA. Human DNA was monitored by targeting the β-2-microglobulin gene, while bacteria were monitored by targeting 16S rDNA (total bacteria and Porphyromonas gingivalis) or the glycosyltransferase gene (Streptococcus mutans).We found that in most cases at least 90% of human DNA could successfully be removed, with complete removal in eight of 16 cases using MolYsis, and two (of 16) cases using Pureprove. Conversely, detection of bacterial DNA was possible in all cases with a recovery rate generally ranging from 35% to 50%. In conclusion, both strategies have the potential to reduce background interference from the host DNA which may be of remarkable value for nucleic-acid based microbial diagnostic systems.     

Aversive Learning under Different Training Conditions: Effects of NMDA Receptor Blockade in Area CA1 of the Hippocampus

Adult male Wistar rats were given either a single training trial or one training trial per day during 3 days followed by a retention test trial in an inhibitory avoidance (IA) task. In animals given a single training trial, pretraining, but not pretest bilateral infusion of the NMDA glutamate receptor antagonist d,l-2-amino-5-phosphonopentanoic acid (AP5) (5.0 μg) into the CA1 hippocampal area blocked IA retention. In animals given three training trials, infusions of AP5 given prior to each of the three training trials severely impaired, but did not block retention. The results indicate that NMDA receptors in the hippocampus are involved in the formation, but not in expression, of aversive memory. In addition, rats given repeated training were able to show a mild improvement of performance across training trials, possibly through mechanisms that do not depend on NMDA receptor activation in the dorsal hippocampus.     

Micronucleus test on gas station attendants

In the present study, the micronucleus test was applied in exfoliated cells of buccal mucosa to assess the mutagenicity risk associated with occupational exposure for gas station attendants. For each individual, 2000 exfoliated buccal cells were analyzed for micronucleus frequency. A highly significant difference was found between exposed and control groups. Likewise, a significant difference was found between these groups regarding the frequency of binucleated and broken egg cells. To determine whether smoking, alcohol habit, age, gender, or working time could exert any additional effect, we determined the frequency of micronuclei and binucleated and broken egg cells amongst exposed and control individuals. The results allowed us to conclude that the individuals studied belong to a risk group and should periodically undergo biological monitoring and appropriate care.     

7th SOSORT consensus paper: conservative treatment of idiopathic & Scheuermann's kyphosis

Abstract

Thoracic hyperkyphosis is a frequent problem and can impact greatly on patient's quality of life during adolescence. This condition can be idiopathic or secondary to Scheuermann disease, a disease disturbing vertebral growth. To date, there is no sound scientific data available on the management of this condition. Some studies discuss the effects of bracing, however no guidelines, protocols or indication's of treatment for this condition were found. The aim of this paper was to develop and verify the consensus on managing thoracic hyperkyphosis patients treated with braces and/or physiotherapy.

Methods

The Delphi process was utilised in four steps gradually modified according to the results of a set of recommendations: we involved the SOSORT Board twice, then all SOSORT members twice, with a Pre-Meeting Questionnaire (PMQ), and during a Consensus Session at the SOSORT Lyon Meeting with a Meeting Questionnaire (MQ).

Results

There was an unanimous agreement on the general efficacy of bracing and physiotherapy for this condition. Most experts suggested the use of 4-5 point bracing systems, however there was some controversy with regards to physiotherapeutic aims and modalities.

Conclusion

The SOSORT panel of experts suggest the use of rigid braces and physiotherapy to correct thoracic hyperkyphosis during adolescence. The evaluation of specific braces and physiotherapy techniques has been recommended.     

Sex differences in carotid baroreflex control of arterial blood pressure in humans: relative contribution of cardiac output and total vascular conductance

It is presently unknown whether there are sex differences in the magnitude of blood pressure (BP) responses to baroreceptor perturbation or if the relative contribution of cardiac output (CO) and total vascular conductance (TVC) to baroreflex-mediated changes in BP differs in young women and men. Since sympathetic vasoconstrictor tone is attenuated in women, we hypothesized that carotid baroreflex-mediated BP responses would be attenuated in women by virtue of a blunted vascular response (i.e., an attenuated TVC response). BP, heart rate (HR), and stroke volume were continuously recorded during the application of 5-s pulses of neck pressure (NP; carotid hypotension) and neck suction (NS; carotid hypertension) ranging from +40 to -80 Torr in women (n = 20, 21 ± 0.5 yr) and men (n = 20, 21 ± 0.4 yr). CO and TVC were calculated on a beat-to-beat basis. Women demonstrated greater depressor responses to NS (e.g., -60 Torr, -17 ± 1%baseline in women vs. -11 ± 1%baseline in men, P < 0.05), which were driven by augmented decreases in HR that, in turn, contributed to larger reductions in CO (-60 Torr, -15 ± 2%baseline in women vs. -6 ± 2%baseline in men, P < 0.05). In contrast, pressor responses to NP were similar in women and men (e.g., +40 Torr, +14 ± 2%baseline in women vs. +10 ± 1%baseline in men, P > 0.05), with TVC being the primary mediating factor in both groups. Our findings indicate that sex differences in the baroreflex control of BP are evident during carotid hypertension but not carotid hypotension. Furthermore, in contrast to our hypothesis, young women exhibited greater BP responses to carotid hypertension by virtue of a greater cardiac responsiveness.     

A new threat: assessing the main interactions between marine fish and plastic debris from a scientometric perspective

Reviews in Fish Biology and Fisheries - Impacts generated by plastic pollution constitute one of the most significant challenges that humanity must face in the next century, receiving increasing...     

Leptin Signaling in Kiss1 Neurons Arises after Pubertal Development

The adipocyte-derived hormone leptin is required for normal pubertal maturation in mice and humans and, therefore, leptin has been recognized as a crucial metabolic cue linking energy stores and the onset of puberty. Several lines of evidence have suggested that leptin acts via kisspeptin expressing neurons of the arcuate nucleus to exert its effects. Using conditional knockout mice, we have previously demonstrated that deletion of leptin receptors (LepR) from kisspeptin cells cause no puberty or fertility deficits. However, developmental adaptations and system redundancies may have obscured the physiologic relevance of direct leptin signaling in kisspeptin neurons. To overcome these putative effects, we re-expressed endogenous LepR selectively in kisspeptin cells of mice otherwise null for LepR, using the Cre-loxP system. Kiss1-Cre LepR null mice showed no pubertal development and no improvement of the metabolic phenotype, remaining obese, diabetic and infertile. These mice displayed decreased numbers of neurons expressing Kiss1 gene, similar to prepubertal control mice, and an unexpected lack of re-expression of functional LepR. To further assess the temporal coexpression of Kiss1 and Lepr genes, we generated mice with the human renilla green fluorescent protein (hrGFP) driven by Kiss1 regulatory elements and crossed them with mice that express Cre recombinase from the Lepr locus and the R26-tdTomato reporter gene. No coexpression of Kiss1 and LepR was observed in prepubertal mice. Our findings unequivocally demonstrate that kisspeptin neurons are not the direct target of leptin in the onset of puberty. Leptin signaling in kisspeptin neurons arises only after completion of sexual maturation.     

Akt activation by insulin treatment attenuates cachexia in Walker-256 tumor-bearing rats

Cancer-bearing often exhibits hypoinsulinemia, insulin (INS) resistance and glutamine depletion associated with cachexia. However, INS and glutamine effects on cachexia metabolic abnormalities, particularly on tumor-affected proteins related to INS resistance, are poorly known. The main purpose of this study was to investigate the effects of INS and glutamine dipeptide (GDP) treatments on phospho-protein kinase B (p-Akt), and phospho-hormone sensitive lipase (p-HSL) in Walker-256 tumor-bearing rats. INS (NPH, 40 UI/kg, subcutaneous), GDP (1.5 g/kg, oral), INS+GDP or vehicle (control rats) were administered for 13 days, once a day, starting at the day of inoculation of tumor cells. The experiments were performed 4 hours after the last treatment to evaluate acute effects of INS and GDP, besides the chronic effects. INS and/or INS+GDP treatments, which markedly increased the insulinemia, increased the p-Akt: total Akt ratio and prevented the increased p-HSL^Ser552: total HSL ratio in the retroperitoneal fat of tumor-bearing rats, without changing the INS resistance and increased expression of factor tumor necrosis-α (TNF-α) in this tissue. INS and INS+GDP also increased the p-Akt: total Akt ratio, whereas GDP and INS+GDP increased the GLUT4 glucose transporter gene expression, in the gastrocnemius muscle of the tumor-bearing rats. Accordingly, treatments with INS and INS+GDP markedly reduced glycemia, increased retroperitoneal fat and attenuated the body mass loss of tumor-bearing rats. In conclusion, hyperinsulinemia induced by high-dose INS treatments increased Akt phosphorylation and prevented increased p-HSL^Ser552: total HSL ratio, overlapping INS resistance. These effects are consistent with increased fat mass gain and weight loss (cachexia) attenuation of tumor-bearing rats, evidencing that Akt activation is a potential strategy to prevent loss of fat mass in cancer cachexia.     

Molecular and biologic characteristics of Toxoplasma gondii isolates from wildlife in the United States

Toxoplasma gondii isolates can be grouped into 3 genetic lineages. Type I isolates are considered more virulent in outbred mice and have been isolated predominantly from clinical cases of human toxoplasmosis, whereas types II and III isolates are considered less virulent for mice and are found in humans and food animals. Little is known of genotypes of T. gondii isolates from wild animals. In the present report, genotypes of isolates of T. gondii from wildlife in the United States are described. Sera from wildlife were tested for antibodies to T. gondii with the modified agglutination test, and tissues from animals with titers of 1:25 (seropositive) were bioassayed in mice. Toxoplasma gondii was isolated from the hearts of 21 of 34 seropositive white-tailed deer (Odocoileus virginianus) from Mississippi and from 7 of 29 raccoons (Procyon lotor); 5 of 6 bobcats (Lynx rufus); and the gray fox (Urocyon cinereoargenteus), red fox (Vulpes vulpes), and coyote (Canis latrans) from Georgia. Toxoplasma gondii was also isolated from 7 of 10 seropositive black bears (Ursus americanus) from Pennsylvania by bioassay in cats. All 3 genotypes of T. gondii based on the SAG2 locus were circulating among wildlife.