Ghost Factors of Laboratory Carbonate Chemistry Are Haunting Our Experiments.

AbstractFor many historical and contemporary experimental studies in marine biology, seawater carbonate chemistry remains a ghost factor, an uncontrolled, unmeasured, and often dynamic variable affecting experimental organisms or the treatments to which investigators subject them. We highlight how environmental variability, such as seasonal upwelling and biological respiration, drive variation in seawater carbonate chemistry that can influence laboratory experiments in unintended ways and introduce a signal consistent with ocean acidification. As the impacts of carbonate chemistry on biochemical pathways that underlie growth, development, reproduction, and behavior become better understood, the hidden effects of this previously overlooked variable need to be acknowledged. Here we bring this emerging challenge to the attention of the wider community of experimental biologists who rely on access to organisms and water from marine and estuarine laboratories and who may benefit from explicit considerations of a growing literature on the pervasive effects of aquatic carbonate chemistry changes.

Eelgrass pathogen Labyrinthula zosterae synthesizes essential fatty acids.

Negative consequences of parasites and disease on hosts are usually better understood than their multifaceted ecosystem effects. The pathogen Labyrinthula zosterae (Lz) causes eelgrass wasting disease but has relatives that produce large quantities of nutritionally valuable long-chain polyunsaturated fatty acids (LCPUFA) such as docosahexaenoic acid (DHA). Here we quantify the fatty acids (FA) of Lz cultured on artificial media, eelgrass-based media, and eelgrass segments to investigate whether Lz may similarly produce LCPUFA. We also assess whether field-collected lesions show similar FA patterns to laboratory-inoculated eelgrass. We find that Lz produces DHA as its dominant FA along with other essential FA on both artificial and eelgrass-based media. DHA content was greater in both laboratory-inoculated and field-collected diseased eelgrass relative to their respective controls. If Lz's production scales in situ, it may present an unrecognized source of LCPUFA in eelgrass ecosystems.

Environment, dosage, and pathogen isolate moderate virulence in eelgrass wasting disease.

Eelgrass wasting disease, caused by the marine pathogen Labyrinthula zosterae, has the potential to devastate important eelgrass habitats worldwide. Although this host-pathogen interaction may increase under certain environmental conditions, little is known about how disease severity is impacted by multiple components of a changing environment. In this study, we investigated the effects of variation in 3 different L. zosterae isolates, pathogen dosage, temperature, and light on severity of infections. Severity of lesions on eelgrass varied among the 3 different isolates inoculated in laboratory trials. Our methods to control dosage of inoculum showed that disease severity increased with pathogen dosage from 104 to 106 cells ml-1. In a dosage-controlled light and temperature 2-way factorial experiment consisting of 2 light regimes (diel light cycle and complete darkness) and 2 temperatures (11 and 18°C), L. zosterae cell growth rate in vitro was higher at the warmer temperature. In a companion experiment that tested the effects of light and temperature in in vivo inoculations, disease severity was higher in dark treatments and temperature was marginally significant. We suggest that the much greater impact of light in the in vivo inoculation experiment indicates an important role for plant physiology and the need for photosynthesis in slowing severity of infections. Our work with controlled inoculation of distinct L. zosterae isolates shows that pathogen isolate, increasing dosage of inoculum, increasing temperature, and diminishing light increase disease severity, suggesting L. zosterae will cause increased damage to eelgrass beds with changing environmental conditions.

Effective multiple sideband generation using an electro-optic modulator for a multiple isotope magneto-optical trap.

Herein, we report an effective method for the generation of radio-frequency (RF) sidebands in an electro-optic modulator for the simultaneous magneto-optical trapping of two isotopes. This is achieved by switching the RF signals alternately, which suppresses the generation of unwanted frequency signals and improves the laser power per sideband. The generated sidebands are successfully applied to a dual-rubidium-isotope magneto-optical trap (MOT), which results in an increased number of trapped atoms. This simple, flexible, and robust technique can be implemented in experiments that require a large number of atoms in multiple-isotope MOTs and for various applications.

Impact of hospital volume on hospital mortality, length of stay and total costs after pancreaticoduodenectomy.

BACKGROUND: High morbidity and mortality rates after pancreaticoduodenectomy (PD) have led to concentration of this surgery in high-volume centres, with improved outcomes. The extent to which better outcomes might be apparent in a healthcare system where the mortality rate is already low is unclear. METHODS: The Japanese Diagnosis Procedure Combination database was used to identify patients undergoing PD between 2007 and 2010. Patient data included age, sex, co-morbidities at admission, type of hospital, type of PD, and the year in which the patient was treated. Hospital volume was defined as the number of PDs performed annually at each hospital, and categorized into quintiles: very low-, low-, medium-, high- and very high-volume groups. The Charlson co-morbidity index was calculated using the International Classification of Diseases, tenth revision, codes of co-morbidities. RESULTS: A total of 10 652 patients who underwent PD in 848 hospitals were identified. The overall in-hospital mortality rate after PD was 3·3 per cent (350 of 10 652), and for the groups ranged from 5·0 per cent for the very low-volume group to 1·4 per cent for the very high-volume group (P < 0·001). Multivariable analysis revealed a significant linear relationship between higher hospital volume and shorter postoperative length of stay compared with the very low-volume group, and between increasing hospital volume and lower total costs. CONCLUSION: A significant relationship exists between increasing hospital volume, lower in-hospital mortality, shorter length of stay and lower costs for patients undergoing PD in Japan. Centralization of PD in this healthcare system is therefore justified.

The clinical efficacy of dietary fat restriction in treatment of dogs with intestinal lymphangiectasia.

BACKGROUND: Intestinal lymphangiectasia (IL), a type of protein-losing enteropathy (PLE), is a dilatation of lymphatic vessels within the gastrointestinal tract. Dietary fat restriction previously has been proposed as an effective treatment for dogs with PLE, but limited objective clinical data are available on the efficacy of this treatment. HYPOTHESIS/OBJECTIVES: To investigate the clinical efficacy of dietary fat restriction in dogs with IL that were unresponsive to prednisolone treatment or showed relapse of clinical signs and hypoalbuminemia when the prednisolone dosage was decreased. ANIMALS: Twenty-four dogs with IL. METHODS: Retrospective study. Body weight, clinical activity score, and hematologic and biochemical variables were compared before and 1 and 2 months after treatment. Furthermore, the data were compared between the group fed only an ultra low-fat (ULF) diet and the group fed ULF and a low-fat (LF) diet. RESULTS: Nineteen of 24 (79%) dogs responded satisfactorily to dietary fat restriction, and the prednisolone dosage could be decreased. Clinical activity score was significantly decreased after dietary treatment compared with before treatment. In addition, albumin (ALB), total protein (TP), and blood urea nitrogen (BUN) concentration were significantly increased after dietary fat restriction. At 2 months posttreatment, the ALB concentrations in the ULF group were significantly higher than that of the ULF + LF group. CONCLUSIONS AND CLINICAL IMPORTANCE: Dietary fat restriction appears to be an effective treatment in dogs with IL that are unresponsive to prednisolone treatment or that have recurrent clinical signs and hypoalbuminemia when the dosage of prednisolone is decreased.

Indocyanine green fluorescence imaging in the surgical management of liver cancers: current facts and future implications.

Imaging detection of liver cancers and identification of the bile ducts during surgery, based on the fluorescence properties of indocyanine green, has recently been developed in liver surgery. The principle of this imaging technique relies on the intravenous administration of indocyanine green before surgery and the illumination of the surface of the liver by an infrared camera that simultaneously induces and collects the fluorescence. Detection by fluorescence is based on the contrast between the (fluorescent) tumoral or peri-tumoral tissues and the healthy (non-fluorescent) liver. Results suggest that indocyanine green fluorescence imaging is capable of identification of new liver cancers and enables the characterization of known hepatic lesions in real time during liver resection. The purpose of this paper is to present the fundamental principles of fluorescence imaging detection, to describe successively the practical and technical aspects of its use and the appearance of hepatic lesions in fluorescence, and to expose the diagnostic and therapeutic perspectives of this innovative imaging technique in liver surgery.

Hypergravity-induced changes in gene expression in Arabidopsis hypocotyls.

Under hypergravity conditions, the cell wall of stem organs becomes mechanically rigid and elongation growth is suppressed, which can be recognized as the mechanism for plants to resist gravitational force. The changes in gene expression by hypergravity treatment were analyzed in Arabidopsis hypocotyls by the differential display method, for identifying genes involved in hypergravity-induced growth suppression. Sixty-two cDNA clones were expressed differentially between the control and 300 g conditions: the expression levels of 39 clones increased, whereas those of 23 clones decreased under hypergravity conditions. Sequence analysis and database searching revealed that 12 clones, 9 up-regulated and 3 down-regulated, have homology to known proteins. The expression of these genes was further analyzed using RT-PCR. Finally, six genes were confirmed to be up-regulated by hypergravity. One of such genes encoded 3-hydroxy-3-methylglutaryl-Coenzyme A reductase (HMGR), which catalyzes a reaction producing mevalonic acid, a key precursor of terpenoids such as membrane sterols and several types of hormones. The expression of HMGR gene increased within several hours after hypergravity treatment. Also, compactin, an inhibitor of HMGR, prevented hypergravity-induced growth suppression, suggesting that HMGR is involved in suppression of Arabidopsis hypocotyl growth by hypergravity. In addition, hypergravity increased the expression levels of genes encoding CCR1 and ERD15, which were shown to take part in the signaling pathway of environmental stimuli such as temperature and water, and those of the alpha-tubulin gene. These genes may be involved in a series of cellular events leading to growth suppression of stem organs under hypergravity conditions.

Utilization of ATP-binding cassette exporter for hyperproduction of an exoprotein: construction of lipase-hyperproducing recombinant strains of Serratia marcescens.

The Serratia marcescens extracellular lipase (LipA) is an enzyme applicable to enantioselective hydrolysis of racemic substrates. The enzyme is secreted through an ATP-binding cassette (ABC) exporter, the Lip system, encoded by the lipBCD genes. The S. marcescens recombinant carrying pLIPE121, which encodes the lipA gene in pUC19, exhibited a higher LipA production level than the wild-type strain. However, the level was lower than expected, and secretion was suggested to be a bottleneck. lipBCD plasmids were introduced into S. marcescens recombinants harboring lipA plasmids and the effectiveness of the lipBCD plasmids in elevating LipA productivity was investigated. S. marcescens strains harboring both lipA and lipBCD plasmids showed sevenfold greater extracellular LipA activity than the strain harboring the lipA plasmid alone. A high level of extracellular LipA production (1,300 kU/ml) and high plasmid stability (enough to carry out large-scale cultivation) were observed under non-selective conditions. Addition of L-proline and Tween 80 was effective in increasing cell growth of the recombinant, which led to high LipA production. In batch cultivation using a 30-l jar fermentor, LipA production was achieved at a high level of 5,200 kU/ml. This is the first report describing utilization of ABC exporter for the overproduction of an industrially important extracellular protein.

[Induction of mucosal immunity to mycobacterial heat shock protein (hsp) 65 by colonic inoculation of plasmid DNA encoding hsp65].

Mycobacterial heat shock protein (hsp) 65 has more than 50% sequence homology with human hsp60 and immune responses against mycobacterial hsp65 may cross-react with human hsp60 and could cause autoimmune diseases including inflammatory bowel diseases (IBD). Since the colonic mucosa is a main inflammatory site in IBD, mucosal immunity to hsp65 may be more important for the mucosal inflammation than systemic immunity to hsp65. We inoculated plasmid DNA (pDNA) encoding mycobacterial hsp65 (pACB-hsp 65) into the colon of Wistar rats and evaluated the mucosal humoral immune response and the effect of these immune responses on the colonic mucosa. Four weeks after pDNA inoculation, significantly elevated titers of hsp65-specific IgA antibody were seen in fecal extracts of rats immunized intra-colonic mucosa with pACB-hsp65 (40 +/- 9 U/ml), whereas the fecal IgA antibody titers of rats inoculated intradermal with pACB-hsp65 did not arise (8 +/- 5 U/ml). Colonic inoculation of pACB-hsp65 induced systemic and mucosal immune responses to hsp65. However, macroscopic and histological examinations of the colonic mucosa inoculated with pACB-hsp65 showed no evidence of mucosal damage. These results suggested that the mucosal immunity to hsp65 on the colonic mucosa may not play a crucial role in the induction of colonic mucosal inflammation as was seen in IBD.

QT dispersion is increased in diabetic patients with foot ulcer.

QT dispersion, a measure of inhomogenous ventricular repolarization, was measured in diabetic patients with foot ulcer. We recruited 75 patients with non insulin-dependent diabetes mellitus: patients with neuropathic ulcer (n=15, NU group), with ischemic ulcer (n=20, IU group), with previous myocardial infarction (n=20, MI group) and without any diabetic microangiopathies (n=20, DC group). We also studied normal control subjects (n=15, NC group). The interlead variability of rate corrected QT interval (QTc dispersion) was calculated. QTc interval in the MI group was significantly higher than that in the NC or DC but showed no difference in the NU and IU groups. QTc dispersion in the IU (54+/-15 msec) as well as MI (60+/-21 msec) group were significantly higher than the NC (36+/-18 msec) or DC group (39+/-14 msec). This may be due to complicated coronary artery disease in the IU group. Furthermore, QTc dispersion was also increased (49+/-14 msec) in the NU group in which cardiac autonomic nervous dysfunction was suggested. Patients with both types of diabetic ulcer demonstrated increased QT dispersion due to atherosclerosis or neurological disorder.