Intensive nitrogen loss over the Omani Shelf due to anammox coupled with dissimilatory nitrite reduction to ammonium

A combination of stable isotopes (¹⁵N) and molecular ecological approaches was used to investigate the vertical distribution and mechanisms of biological N₂ production along a transect from the Omani coast to the central–northeastern (NE) Arabian Sea. The Arabian Sea harbors the thickest oxygen minimum zone (OMZ) in the world''s oceans, and is considered to be a major site of oceanic nitrogen (N) loss. Short (<48 h) anoxic incubations with ¹⁵N-labeled substrates and functional gene expression analyses showed that the anammox process was highly active, whereas denitrification was hardly detectable in the OMZ over the Omani shelf at least at the time of our sampling. Anammox was coupled with dissimilatory nitrite reduction to ammonium (DNRA), resulting in the production of double-¹⁵N-labeled N₂ from ¹⁵NO₂⁻, a signal often taken as the lone evidence for denitrification in the past. Although the central–NE Arabian Sea has conventionally been regarded as the primary N-loss region, low potential N-loss rates at sporadic depths were detected at best. N-loss activities in this region likely experience high spatiotemporal variabilities as linked to the availability of organic matter. Our finding of greater N-loss associated with the more productive Omani upwelling region is consistent with results from other major OMZs. The close reliance of anammox on DNRA also highlights the need to take into account the effects of coupling N-transformations on oceanic N-loss and subsequent N-balance estimates.     

Anaerobic ammonia oxidation in a fertilized paddy soil

Evidence for anaerobic ammonium oxidation in a paddy field was obtained in Southern China using an isotope-pairing technique, quantitative PCR assays and 16S rRNA gene clone libraries, along with nutrient profiles of soil cores. A paddy field with a high load of slurry manure as fertilizer was selected for this study and was shown to contain a high amount of ammonium (6.2–178.8 mg kg⁻¹). The anaerobic oxidation of ammonium (anammox) rates in this paddy soil ranged between 0.5 and 2.9 nmolN per gram of soil per hour in different depths of the soil core, and the specific cellular anammox activity observed in batch tests ranged from 2.9 to 21 fmol per cell per day. Anammox contributed 4–37% to soil N2 production, the remainder being due to denitrification. The 16S rRNA gene sequences of surface soil were closely related to the anammox bacteria ‘Kuenenia'', ‘Anammoxoglobus'' and ‘Jettenia''. Most of the anammox 16S rRNA genes retrieved from the deeper soil were affiliated to ‘Brocadia''. The retrieval of mainly bacterial amoA sequences in the upper part of the paddy soil indicated that nitrifying bacteria may be the major source of nitrite for anammox bacteria in the cultivated horizon. In the deeper oxygen-limited parts, only archaeal amoA sequences were found, indicating that archaea may produce nitrite in this part of the soil. It is estimated that a total loss of 76 g N m⁻² per year is linked to anammox in the paddy field.     

Diverse reactions catalyzed by naphthalene dioxygenase fromPseudomonas sp strain NCIB 9816

Naphthalene dioxygenase (NDO) fromPseudomonas sp strain NCIB 9816 is a multicomponent enzyme system which initiates naphthalene catabolism by catalyzing the addition of both atoms of molecular oxygen and two hydrogen atoms to the substrate to yield enantiomerically pure (+)-cis-(1R,2S)-dihydroxy-1,2-dihydronaphthalene. NDO has a relaxed substrate specificity and catalyzes the dioxygenation of many related 2- and 3-ring aromatic and hydroaromatic (benzocyclic) compounds to their respectivecis-diols. Biotransformations with a diol-accumulating mutant, recombinant strains and purified enzyme components have established that in addition tocis-dihydroxylation, NDO also catalyzes a variety of other oxidations which include monohydroxylation, desaturation (dehydrogenation),O-andN-dealkylation and sulfoxidation reactions. In several cases, the absolute stereochemistry of the oxidation products formed by NDO are opposite to those formed by toluene dioxygenase (TDO). The reactions catalyzed by NDO and other microbial dioxygenases can yield specific hydroxylated compounds which can serve as chiral synthons in the preparation of a variety of compounds of interest to pharmaceutical and specialty chemical industries. We present here recent work documenting the diverse array of oxidation reactions catalyzed by NDO. The trends observed in the oxidation of a series of benzocyclic aromatic compounds are compared to those observed with TDO and provide the basis for prediction of regio- and stereospecificity in the oxidation of related substrates. Based on the types of reactions catalyzed and the biochemical characteristics of NDO, a mechanism for oxygen activation by NDO is proposed.     

Sequence of a cDNA for mouse thymidylate synthase reveals striking similarity with the prokaryotic enzyme

We report the nucleotide sequence of a cloned cDNA, pMTS-3, that contains a 1-kb insert corresponding to mouse thymidylate synthase (E.C. 2.1.1.45). The open reading frame of 921 nucleotides from the first AUG to the termination codon specifies a protein with a molecular mass of 34,962 daltons. The predicted amino acid sequence is 90% identical with that of the human enzyme. The mouse sequence also has an extremely high degree of similarity (as much as 55% identity) with prokaryotic thymidylate synthase sequences, indicating that thymidylate synthase is among the most highly conserved proteins studied to date. The similarity is especially pronounced (as much as 80% identity) in the 44-amino-acid region encompassing the binding site for deoxyuridylic acid. The cDNA sequence also suggests that mouse thymidylate synthase mRNA lacks a 3' untranslated region, since the termination codon, UAA, is followed immediately by a poly(A) segment.      

Cytotoxic Efficacy of Photodynamic Therapy in Osteosarcoma Cells In Vitro

In recent years, there has been the difficulty in finding more effective therapies against cancer with less systemic side effects. Therefore Photodynamic Therapy is a novel approach for a more tumor selective treatment.Photodynamic Therapy (PDT) that makes use of a nontoxic photosensitizer (PS), which, upon activation with light of a specific wavelength in the presence of oxygen, generates oxygen radicals that elicit a cytotoxic response¹. Despite its approval almost twenty years ago by the FDA, PDT is nowadays only used to treat a limited number of cancer types (skin, bladder) and nononcological diseases (psoriasis, actinic keratosis)².The major advantage of the use of PDT is the ability to perform a local treatment, which prevents systemic side effects. Moreover, it allows the treatment of tumors at delicate sites (e.g. around nerves or blood vessels). Here, an intraoperative application of PDT is considered in osteosarcoma (OS), a tumor of the bone, to target primary tumor satellites left behind in tumor surrounding tissue after surgical tumor resection. The treatment aims at decreasing the number of recurrences and at reducing the risk for (postoperative) metastasis.In the present study, we present in vitro PDT procedures to establish the optimal PDT settings for effective treatment of widely used OS cell lines that are used to reproduce the human disease in well established intratibial OS mouse models. The uptake of the PS mTHPC was examined with a spectrophotometer and phototoxicity was provoked with laser light excitation of mTHPC at 652 nm to induce cell death assessed with a WST-1 assay and by the counting of surviving cells. The established techniques enable us to define the optimal PDT settings for future studies in animal models. They are an easy and quick tool for the evaluation of the efficacy of PDT in vitro before an application in vivo.     

Evaluation of F8-TNF-α in Models of Early and Progressive Metastatic Osteosarcoma

The targeted delivery of tumor necrosis factor-α (TNF-α) with antibodies specific to splice isoforms of fibronectin [e.g., F8-TNF, specific to the extra-domain A (EDA) domain of fibronectin] has already shown efficacy against experimental sarcomas but has not yet been investigated in orthotopic sarcomas. Here, we investigated F8-TNF in a syngeneic K7 M2–derived orthotopic model of osteosarcoma as a treatment against pulmonary metastases, the most frequent cause of osteosarcoma-related death. Immunofluorescence on human osteosarcoma tissue confirmed the presence of EDA in primary tumors (PTs) as well as metastases. In mice, the efficacy of F8-TNF against PTs and early pulmonary metastases was evaluated. Intratibial PT growth was not affected by F8-TNF, yet early micrometastases were reduced possibly due to an F8-TNF–dependent attraction of pulmonary CD4⁺, CD8⁺, and natural killer cells. Furthermore, immunofluorescence revealed stronger expression of EDA in early pulmonary metastases compared with PT tissue. To study progressing pulmonary metastases, a hind limb amputation model was established, and the efficacy of F8-TNF, alone or combined with doxorubicin, was investigated. Despite the presence of EDA in metastases, no inhibition of progressive metastatic growth was detected. No significant differences in numbers of CD4⁺ or CD8⁺ cells or F4/80⁺ and Ly6G⁺ myeloid-derived cells were observed, although a strong association between metastatic growth and presence of pulmonary Ly6G⁺ myeloid-derived cells was detected. In summary, these findings demonstrate the potential of F8-TNF in activating the immune system and reducing early metastatic growth yet suggest a lack of efficacy of F8-TNF alone or combined with doxorubicin against progressing osteosarcoma metastases.     

The mycoflora and toxicity of feedstuffs from a production plant in córdoba, Argentina

Feedstuffs used for poultry nutrition in Argentina were analyzed for fungal flora and natural incidence of mycotoxins. Survey of 120 samples of poultry feeds, taken from May 1998 to April 1999, showed the presence of 15 genera of filamentous fungi. The predominant genera wereFusarium spp. andPenicillium ssp., isolated in 67.5 % of the samples, followed byAspergillus spp. (57.5 %). Yeast, were significantly isolated from most of the samples. Species identification was carried down for the toxigenic genera. Fungal total counts of poultry feeds ranged from 2.0 × 10³ to 3.0 × 10⁵ CFU g^-1 The fungal total counts during two months of sampling, were slightly over the limit value of 1 × 105 CFU g^-1, which ensure the hygienic quality of the feed. Potentially toxicogenic species presented moderate mean colony counts. Many of the fungi isolated from poultry feeds are mycotoxin producers. Fumonisins had the highest incidence, and were found in 97 % of the analyzed samples followed by aflatoxin B₁ (46 %), zearalenone (18 %) and deoxynivalenol (6 %). On the co-occurrence of both carcinogenic mycotoxins, all of the FBs contaminated feed samples were co-contaminated with AFB₁. The results show the relevance of the samples screening for viable fungi propagules and the surveillance of their associated mycotoxins in poultry feeds.     

Feasibility study of detecting surface electromyograms in severely obese patients

The aims of this study were to examine if surface EMG signals can be detected from the quadriceps femoris muscle of severely obese patients and to investigate if differences exist in quadriceps force and myoelectric manifestations of fatigue between obese patients and lean controls.Fourteen severely obese patients (body mass index, BMI, mean ± SD: 44.9 ± 6.3 kg/m²) and fourteen healthy controls (BMI: 23.7 ± 2.5 kg/m²) were studied. The vastus medialis and lateralis of the dominant thigh were concurrently investigated during voluntary isometric contractions (10-s long at submaximal and maximal intensities and intermittent submaximal contractions until exhaustion) and sustained (120-s long) electrically elicited contractions.We found that the detection of surface EMG signals from the quadriceps is feasible also in severely obese subjects presenting increased thickness of the subcutaneous fat tissue. In addition, we confirmed and extended previous findings showing that the volume conductor properties determine the amplitude and spectral features of the detected surface EMG signals: the lower the subcutaneous tissue thickness, the higher the amplitude and mean frequency estimates. Further, we found no differences in the mechanical and myoelectric manifestations of fatigue during intermittent voluntary and sustained electrically elicited contractions between obese patients and lean controls.     

Sprint,agility, strength and endurance capacity in wheelchair basketball players

The aims of the present study were, firstly, to determine the reliability and reproducibility of an agility T-test and Yo-Yo 10 m recovery test; and secondly, to analyse the physical characteristics measured by sprint, agility, strength and endurance field tests in wheelchair basketball (WB) players. 16 WB players (33.06 ± 7.36 years, 71.89 ± 21.71 kg and sitting body height 86.07 ± 6.82 cm) belonging to the national WB league participated in this study. Wheelchair sprint (5 and 20 m without ball, and 5 and 20 m with ball) agility (T-test and pick-up test) strength (handgrip and maximal pass) and endurance (Yo-Yo 10 m recovery test) were performed. T-test and Yo-Yo 10 m recovery test showed good reproducibility values (intraclass correlation coefficient, ICC = 0.74-0.94). The WB players’ results in 5 and 20 m sprints without a ball were 1.87 ± 0.21 s and 5.70 ± 0.43 s and with a ball 2.10 ± 0.30 s and 6.59 ± 0.61 s, being better than those reported in the literature. Regarding the pick-up test results (16.05 ± 0.52 s) and maximal pass (8.39 ± 1.77 m), players showed worse values than those obtained in elite players. The main contribution of the present study is the characterization of the physical performance profile of WB players using a field test battery. Furthermore, we demonstrated that the agility T-test and the aerobic Yo-Yo 10 m recovery test are reliable; consequently they may be appropriate instruments for measuring physical fitness in WB.