Production of glycerol by Lactobacillus plantarum NRRL B-4496 and formation of hexamine during fermentation of pea protein enriched flour.

Suspensions of pea protein enriched flour (PP) inoculated with Lactobacillus plantarum NRRL B-4496 and uninoculated PP suspensions were incubated in vials covered with airtight caps. Organic compound compositions of fermented and unfermented PP suspensions (F-PP and U-PP, respectively) were analyzed using solid phase microextraction (SPME) coupled with gas chromatography - mass-spectrometry (GCMS). Acetic acid was detected in all samples; pH dropped from pH 6.5 to pH 4.1 in L. plantarum F-PP and to pH 5.3 in uninoculated F-PP. Abundance of acetic acid and minuscule presence of lactic acid in L. plantarum F-PP suggested that fermentation proceeded preferentially via the pyruvate formate lyase (PFL) pathway. Nonetheless, glycerol appeared to be the most abundant compound in L. plantarum F-PP samples; colorimetric analysis indicated that its average concentration in these samples was 1.05 g/L. A metabolic switch from the PFL pathway to glycerol production might occur due to acidity tolerance limitations of L. plantarum, glycerol production being associated with the release of phosphate, which can act as a buffer. Fermentation of PP by L. plantarum also led to formation of hexamine, which is a known food preservation agent. Presence of naturally formed hexamine and glycerol in food products may render using chemical additives needless.

Impact of hexamine addition to a nitrite-based additive on fermentation quality, Clostridia and Saccharomyces cerevisiae in a white lupin-wheat silage.

BACKGROUND: Nitrite and hexamine are used as silage additives because of their adverse effects on Clostridia and Clostridia spores. The effect of sodium nitrite and sodium nitrite/hexamine mixtures on silage quality was investigated. A white lupin-wheat mixture was treated with sodium nitrite (NaHe0) (900 g t-1 forage), or mixtures of sodium nitrite (900 g t-1 ) and hexamine. The application rate of hexamine was 300 g t-1 (NaHe300) or 600 g t-1 (NaHe600). Additional treatments were the untreated control (Con), and formic acid (FA) applied at a rate of 4 L t-1 (1000 g kg-1 ). RESULTS: Additives improved silage quality noticeably only by reducing silage ammonia content compared with the control. The addition of hexamine to a sodium nitrite solution did not improve silage quality compared with the solution containing sodium nitrite alone. The increasing addition of hexamine resulted in linearly rising pH values (P < 0.001) and decreasing amounts of lactic acid (P < 0.01). Sodium nitrite based additives were more effective than formic acid in preventing butyric acid formation. Additives did not restrict the growth of Saccharomyces cerevisiae compared to the control. CONCLUSION: The addition of hexamine did not improve silage quality compared with a solution of sodium nitrite. © 2018 Society of Chemical Industry.

Bronchoalveolar Lavage Fluid Cytology in Culture-Documented Invasive Pulmonary Aspergillosis in Patients with Hematologic Diseases: Analysis of 67 Episodes.

There is a paucity of studies on the yield of Gomori-methenamine-silver (GMS) staining in bronchoalveolar lavage (BAL) fluid cytology and its comparison with fluorescent dye staining for the diagnosis of invasive pulmonary aspergillosis (IPA) in patients with hematologic malignancies. To that end, we analyzed the yield of direct fungal visualization in BAL fluid cytology with GMS staining, in a series of culture-positive IPA cases in 67 patients with hematologic malignancies, and we compared the results with those of direct examination with calcofluor white staining and BAL fluid galactomannan assays, when available. GMS staining in BAL fluid cytology was positive in 42% of the 67 cases and revealed coinfections in 7 cases. In contrast, only 2/67 (3.6%) BAL fluid samples were positive in direct smears stained with the fluorescent dye calcofluor white. Positive GMS staining results were significantly more frequent in IPA cases with cavitary lesions and IPA cases caused by >1 Aspergillus species, but the proportions of positive cytology results among Aspergillus species were not different.

Integrated biological and advanced oxidation based treatment of hexamine bearing wastewater: Effect of cow-dung as a co-substrate.

This work examines the treatment of hexamethylenetetramine (HMT) bearing effluent from N, N-dinitroso pentamethylene tetra-mine producing industrial plants in India. Chemical treatment using Fenton's reagent and aerobic treatment using batch reactors with co-substrate were investigated. Aerobic batch reactors integrated with advanced oxidation process of Fenton's reagent provides effective treatment of HMT effluents. Influence of Fenton's reagent dose reaction/contact and effect of varying co-substrate with effluent initial concentration was observed. Higher dose 100 mL of Fenton's reagent with higher reaction time 20 h resulted better degradation (34.88%) of wastewater. HMT hydrolyzes in acidic environment to ammonia and formaldehyde. Formaldehyde under normal conditions is toxic for biological treatment processes. When hydrolysis and acidification in the reactors are accompanied by low pH, aerobic batch reactors with use of co-substrates glucose, sucrose, and cow-dung extract separately in different proportion to wastewater ranging from 0.67 to 4.00, degraded wastewater effectively. Higher proportion of co-substrate to wastewater resulted better degradation. The relationships between nitrate, pH, turbidity and COD are discussed.

Utility of reflexive gomori methenamine silver and Acid-fast bacillus staining on bronchoalveolar lavage specimens.

OBJECTIVE: Reflexive testing of bronchoalveolar lavage (BAL) specimens with Gomori methenamine silver (GMS) and acid-fast bacillus (AFB) stains is not routinely performed by most institutions. Instead, these stains are usually ordered to evaluate for the presence of fungal elements and/or acid-fast organisms if initial histopathologic assessment suggests the presence of these pathogens. Our institution, however, performs these stains on all BAL specimens. Thus, we sought to determine whether this practice was cost effective, considering the turnaround time and diagnostic efficacy of these tests. METHODS: We retrospectively reviewed 488 BAL specimens performed at two military healthcare institutions over a 2-year period and performed a cost analysis with review of the impact on turnaround time. RESULTS: Of the 488 cases, we identified only 3 (~0.6%) with infections by acid-fast or fungal organisms, at an estimated total cost of $12,151.20 and an average delay of 3.0 to 3.5 hours for slide preparation. CONCLUSION: Our results suggest that in a largely young and healthy population such as ours, it may be more feasible to perform these stains on BAL specimens on a case-by-case basis rather than automatically on every specimen, to control costs and enhance productivity.

Repolymerization of actin from actin:thymosin beta4 complex induced by diaphanous related formins and gelsolin.

The beta-thymosins are peptides of about 5 kDa molecular mass. Thymosin beta4 (Tbeta4) is the most ubiquitous member of this family and composed of 43 residues. Initially the beta-thymosins were supposed to be specifically produced and released by the thymic gland and to possess hormonal activities modulating the immune response. However, it was later noticed that beta-thymosins are present in the cytoplasm of almost all eukaryotic cells. Especially high concentrations of Tbeta4 were detected in hematopoetic cells, like polymorpho-nuclear leucocytes and in platelets. In these cells the main intracellular function of the beta-thymosins is to bind to monomeric actin and to inhibit its polymerization to filamentous actin. Thus Tbeta4 allows resting eukaryotic cells to maintain a high concentration of monomeric actin, although the intracellular ionic conditions would favor its almost complete polymerization to F-actin. Thereby monomeric actin is sequestered from the dynamic assembly and disassembly processes of the actin cytoskeleton that constantly occur intracellularly.

Regulation of the yeast formin Bni1p by the actin-regulating kinase Prk1p.

The formin family of proteins promotes the assembly of linear actin filaments in the cells of diverse eukaryotic organisms. The predominant formins in mammalian cells are self-inhibited by an intramolecular interaction between two terminal domains and are activated by the binding of the Rho GTPases and other factors. In this study, we show that Bni1p, a formin required for the assembly of actin cables in budding yeast, is also regulated by an autoinhibitory mechanism and phosphorylation by the actin regulatory kinase Prk1p, and possibly Ark1p as well, plays a key role in unlocking the inhibition. Bni1p is phosphorylated by Prk1p at three [L/V/I]xxxxTG motifs in vitro, and the phosphorylation is sufficient to activate Bni1p by disrupting its intramolecular interaction. This finding extends the roles of Prk1p in the regulation of actin dynamics to be associated with both anterograde and retrograde transport pathways, i.e. exocytosis and endocytosis, in yeast.

RNAi-mediated knockdown of dystrophin expression in adult mice does not lead to overt muscular dystrophy pathology.

Duchenne muscular dystrophy (DMD) is a fatal muscle wasting disorder caused by mutations in the dystrophin gene. DMD has a complex and as yet incompletely defined molecular pathophysiology. The peak of the pathology attributed to dystrophin deficiency happens between 3 and 8 weeks of age in mdx mice, the animal model of DMD. Accordingly, we hypothesized that the pathology observed with dystrophin deficiency may be developmentally regulated. Initially, we demonstrated that profound small interfering RNA-mediated dystrophin knockdown could be achieved in mouse primary muscle cultures. The use of adeno-associated virus vectors to express short-hairpin RNAs targeting dystrophin in skeletal muscle in vivo yielded a potent and specific dystrophin knockdown, but only after approximately 5 months, indicating the very long half-life of dystrophin. Interestingly, and in contrast to what is observed in congenital dystrophin deficiency, long-term ( approximately 1 year) dystrophin knockdown in adult mice did not result, per se, in overt dystrophic pathology or upregulation of utrophin. This supports our hypothesis and suggests new pathophysiology of the disease. Furthermore, taking into account the rather long half-life of dystrophin, and the notion that the development of pathology is age-dependent, it indicates that a single gene therapy approach before the onset of pathology might convey a long-term cure for DMD.

Utilization of hexamethylenetetramine (urotropine) by bacteria and yeasts.

A slow growing bacterial population able to utilize hexamethylelenetetramine (urotropine) as sole source of carbon, nitrogen and energy was isolated from soil. From this crude enrichment culture two bacteria were isolated and identified as Brevundimonas diminuta and a Phyllobacterium sp. by sequencing of 16S ribosomal DNA. These bacteria also grew on urotropine but at a lower rate than the enrichment culture. Addition of glucose to the latter resulted in growth of some yeasts that overgrew the bacteria. Assimilation of urotropine as sole nitrogen source is very common among yeasts, 46 out of 60 species tested showed this characteristic.

Utility of reflex Gomori methenamine silver staining for Pneumocystis jirovecii on bronchoalveolar lavage cytologic specimens: a review.

Pneumocystis jiroveci (Pj; formerly Pneumocystis carinii) is an opportunistic pathogen causing life-threatening pneumonia (Pneumocystis pneumonia) in immunosuppressed individuals. Its diagnosis is dependent on identification in bronchoalveolar lavage (BAL) specimens. Gomori's methenamine silver nitrate (GMS) stain has been advocated to highlight the organisms in BAL specimens. This study was performed to determine the utility of reflex GMS staining on all BAL specimens for identifying Pj.All BAL specimens from years 2000 to 2004 were processed as cytospins and stained with Papanicolaou (Pap) and GMS stains. A total of 2,984 BAL specimens were identified. A total of 116 (3.9% of total BAL) BAL specimens were diagnostic of Pj. The diagnostic specimens were grouped as follows: 103 (88.8% of total positive cases) Pj identified with both Pap and GMS staining; 11 (9.5% of total positive cases) Pj identified only with Pap staining; and 2 (1.7% of total positive cases) Pj identified only with GMS staining. In conclusion, the prevalence of Pj in BAL specimens is 3.9%, which can be attributed to improved management of immunocompromised patients. Performing reflex GMS staining on all BAL specimens does not improve the diagnostic identification of Pj since the majority (98.3%) of diagnoses can be rendered on Pap stained slides. A cost analysis for GMS staining on 2,879 GMS-negative BAL specimens was estimated at $143,950. Thus, from diagnostic and cost benefit perspectives, GMS staining can be recommended only on cases where Pap stain is negative, and the clinical presentation is consistent with Pneumocystis pneumonia.

Biodegradation of formaldehyde and its derivatives in industrial wastewater with methylotrophic yeast Hansenula polymorpha and with the yeast-bioaugmented activated sludge.

Methylotrophic yeast Hansenula polymorpha were shown to cooperate with activated sludge from biological wastewater treatment stations, enhancing substantially its potential to biodegrade formaldehyde in industrial wastewater. After integration with yeast cells the modified sludge retained its original structure and activity whereas its resistance to elevated formaldehyde concentrations was significantly improved. The applicability of the yeast in the utilization of formaldehyde derivatives, as exemplified by urotropine and trioxane, was also investigated. The treatment of urotropine-containing wastewater with methylotrophic yeast was found to be effective at acidic conditions (pH below 5.5). Trioxane was not degraded due to the stability of an ether bond which made themolecule recalcitrant to oxidation via methylotrophic pathway reactions. It is concluded that the yeast species may be applied to treat wastewater containing formaldehyde and some of its derivatives as either monocultures or as an integrated, specialized element of the activated sludge biocenosis.

Use of 2H2O for estimating rates of gluconeogenesis. Application to the fasted state.

A method is introduced for estimating the contribution of gluconeogenesis to glucose production. 2H2O is administered orally to achieve 0.5% deuterium enrichment in body water. Enrichments are determined in the hydrogens bound to carbons 2 and 6 of blood glucose and in urinary water. Enrichment at carbon 6 of glucose is assayed in hexamethylenetetramine, formed from formaldehyde produced by periodate oxidation of the glucose. Enrichment at carbon 2 is assayed in lactate formed by enzymatic transfer of the hydrogen from glucose via sorbitol to pyruvate. The fraction gluconeogenesis contributes to glucose production equals the ratio of the enrichment at carbon 6 to that at carbon 2 or in urinary water. Applying the method, the contribution of gluconeogenesis in healthy subjects was 23-42% after fasting 14 h, increasing to 59-84% after fasting 42 h. Enrichment at carbon 2 to that in urinary water was 1.12 +/- 0.13. Therefore, the assumption that hydrogen equilibrated during hexose-6-P isomerization was fulfilled. The 3H/14C ratio in glucose formed from [3-3H,3-14C]lactate given to healthy subjects was 0.1 to 0.2 of that in the lactate. Therefore equilibration during gluconeogenesis of the hydrogen bound to carbon 6 with that in body water was 80-90% complete, so that gluconeogenesis is underestimated by 10-20%. Glycerol's contribution to gluconeogenesis is not included in these estimates. The method is applicable to studies in humans of gluconeogenesis at safe doses of 2H2O.

[A study on fixatives for the simultaneous histological estimation of the gastric mucous gel layer and mucous cells in rat gastric surface mucosa].

We examined the special characteristics of various kinds of fixatives and tried to find the most suitable method for simultaneous histological estimation of the mucous gel layer and mucous cells in rat gastric paraffin sections stained with Alcian blue (pH2.5)-periodic acid Schiff (AB-PAS). The tested fixatives were (group in parentheses): absolute ethanol at -80 degrees C (A), absolute methanol at -80 degrees C (B), Carnoy's solution (C), formalin-ethanol (D) and formalin-Tyrode (E). The thickness of the mucous gel layer (ML) and the numbers of AB- and PAS-positive cells (AB cells, PAS cells) of both the fundic gland area (F. area) and pyloric gland area (P. area) were measured microscopically (x 200). ML in the F. area was found in the order of (A) > (B) > (C) > (D) = (E). AB cells were in the order of (A) > (E) > (B) > (C) > (D); and PAS cells were in the order of (B) > (A) > (E) > (C) > (D). Effects of various fixatives in the P. area showed the same trend as the F. area. Apart from above mentioned procedures, we observed the effects of ethanol at various temperatures: -80 degrees C, -25 degrees C, -4 degrees C and 20 degrees C. ML, AB and PAS cells showed the highest level at -80 degrees C. Moreover, to confirm that these fixatives retain the mucus, we evaluated the hexose and hexosamine contents contained in the fixative solution after fixation. As a result of various fixations, the hexose and hexosamine values in the fixative solution were the smallest for ethanol at -80 degrees C. In conclusion, ethanol at -80 degrees C was the most suitable fixative for histological estimation of mucous gel and mucous cells in rat gastric surface mucosa.

Efficacy of two proprietary preparations of frusemide in patients with congestive heart failure.

The efficacies of two proprietary preparations of frusemide (Lasix and Urex) were compared in five patients with congestive heart failure. No significant differences in efficacy were found between these preparations in respect of the symptoms experienced by patients and the results of biochemical tests.

Beware of formaldehyde in disguise.

The toxic potential of formaldehyde has recently been widely discussed, including the consequences of its release from certain building materials. The action of methenamine, used in the treatment of urinary infections, is based on the release of formaldehyde in the body. Various aspects of formaldehyde toxicity are discussed as a basis for reevaluation of methenamine and reconsideration of its safety.

Formaldehyde generators: relationship between stability, lipophilicity and carcinogenic potency.

The rate of hydrolysis to formaldehyde of methylenedimorpholine (MDM), hexamethylenetetramine (HMT) and dinitrosopentamethylenetetramine (DNPT) have been compared with the enzyme-mediated formation of formaldehyde from hexamethylphosphoramide (HMPA). The bio-distribution of [14C]HMPA following nasal instillation in rats has also been studied and compared with that of [14C]methyl methanesulphonate (MMS). These data are used to relate the several carcinogenic/genotoxic properties of the chemicals named above. It is concluded from these data and related considerations that three classes of formaldehyde-generators should be recognized (a) labile agents such as MDM (and formaldehyde) which are likely to be locally active as carcinogens, (b) lipophilic and relatively stable agents such as HMPA which may be both locally active (if bio-accumulated) and systemically active as carcinogens, and (c) agents such as HMT and DNPT, of intermediate stability, which are unlikely to be systemically active and to be of low or zero activity locally.

The methenamine misunderstanding in the therapy of phosgene poisoning.

A thorough analysis of the pertinent literature leads to the conclusion that there is no justification for the use of methenamine (hexamethylene tetramin, Urotropin) for the therapy of phosgene poisoning.

Drug therapy reviews: methenamine mandelate and methenamine hippurate.

The mechanism of action, spectrum of antimicrobial activity, pharmacokinetics, adverse effects, therapeutic use, and dosage of methenamine hippurate and methenamine mandelate are reviewed. The antimicrobial activity of methenamine depends on its conversion in the urine to formaldehyde. Formaldehyde's spectrum of antibacterial activity encompasses all urinary tract pathogens. Urinary concentrations of formaldehyde vary with pH and urine volume; however, there is no documentation that acdification of the urine enhances methenamine's therapeutic activity. Adverse reactions to methenamine, including gastrointestinal intolerance and skin reactions, are mild and reversible and occur infrequently. Methenamine mandelate and hippurate are effective in the prevention of recurrent urinary tract infections except in patients with Foley catheters or who require intermittent catheterization.