Exploring microbial diversity in hot springs of Surajkund, India through 16S rRNA analysis and thermozyme characterization from endogenous isolates.

Hot springs are a valuable source of biologically significant chemicals due to their high microbial diversity. To investigate the possibilities for industrial uses of these bacteria, researchers collected water and sediment samples from variety of hot springs. Our investigation employed both culture-dependent and culture-independent techniques, including 16S-based marker gene analysis of the microbiota from the hot springs of Surajkund, Jharkhand. In addition, we cultivated thermophilic isolates and screened for their ability to produce amylase, xylanase, and cellulase. After the optimized production of amylase the enzyme was partially purified and characterized using UPLC, DLS-ZP, and TGA. The retention time for the amylase was observed to be around 0.5 min. We confirmed the stability of the amylase at higher temperatures through observation of a steady thermo gravimetric profile at 400 °C. One of the thermophilic isolates obtained from the kund, demonstrated the potential to degrade lignocellulosic agricultural waste.

Fournier's gangrene in a female with spinal cord injury: a case report.

INTRODUCTION: We report the case of a woman with spinal cord injury (SCI) who presented to us with Fournier's gangrene. CASE REPORT: A 60-year-old patient with SCI, ASIA A, neurological level D6, on clean intermittent catheterization presented with rapid necrosis of the perineal region, including the labia and anus, which developed after traumatic catheterizations for clean intermittent catheterization. She required repeated debridement and loop colostomy for management. CONCLUSION: We conclude that patients with SCI are rarely at risk for Fournier's gangrene secondary to neurogenic bladder and bowel, as well as to impaired sensations and genital flora. Treating physicians need to be aware of this complication in order to prevent mortality.

Ganglioside patterns and phenotypic characteristics in a normal variant and a transformed back variant of a simian virus 40-induced hamster tumor cell line.

Ganglioside patterns of a cloned Simian virus 40- (SV40) induced hamster tumor cell (Cl(2)TSV(5)-S), its normal variant (Cl(2)TSV(5)-R) which are Cl(2)TSV(5)-S gradually adapted to grow in the presence of 2 microg/ml actinomycin D and exhibit certain normal phenotypic characteristics, and its back variant (Cl(2)TSV(5)-RR), which are Cl(2)TSV(5)-R cells grown in the absence of actinomycin D for more than 60 passages and which exhibit greater phenotypic similarity to Cl(2)TSV(5)-S cells, have been analyzed. All three cell lines contain N(acetylneuraminyl) galactosylglycosyl ceramide (hematoside, GM(3)), N-acetylgalactosaminyl (N-acetylneuraminyl) galactosylglucosyl ceramide (GM(2)), and a higher ganglioside tentatively identified as disialohematoside. However, Cl(2)TSV(5)-R have more GM(2) than Cl(2)TSV(5)-S whereas Cl(2)TSV(5)-RR contain an intermediate amount of GM(2). The amount of GM(2) is correlative with the activity of UDP-N-acetylgalactosamine: hematoside N-acetylgalactosaminyl transferase in the extract of the three cell lines and with their agglutination by wheat germ agglutinin.

Growth of normal and transformed rat embryo fibroblasts: effects of glycolipids from Salmonella minnesota R mutants.

Addition of glycolipids obtained from Salmonella minnesota R mutants to normal, spontaneously transformed, and SV40-transformed rat embryo fibroblasts in culture results in an inhibition of growth of transformed cells but not of normal cells. In the presence of the glycolipid with the smallest carbohydrate chain length, spontaneously transformed cells stop growing when they reach confluency. Inhibition of growth of transformed cells is inversely related to the chain length of the core sugars. Glycolipid mR595 is shown to bind with the cell membrane of transformed cells and elicits an augmentation in the intracellular level of cyclic AMP. Normal cells bind relatively less glycolipid mR595 and show a lower percent of increase in cyclic AMP due to glycolipid mR595 than do transformed cells.

Nitrilase-catalysed conversion of acrylonitrile by free and immobilized cells of Streptomyces sp.

The biotransformation of acrylonitrile was investigated using thermophilic nitrilase produced from a new isolate Streptomyces sp. MTCC 7546 in both the free and immobilized state. Under optimal conditions,the enzyme converts nitriles to acids without the formation of amides.The whole cells of the isolate were immobilized in agar-agar and the beads so formed were evaluated for 25 cycles at 50 degrees C. The enzyme showed a little loss of activity during reuse. Seventy-one per cent of 0.5 M acrylonitrile was converted to acid at 6 h of incubation at a very low density of immobilized cells, while 100% conversion was observed at 3 h by free cells.

Autologous olfactory [corrected] mucosal transplant in chronic spinal cord injury: an Indian Pilot Study.

STUDY DESIGN: Prospective Pilot Study. OBJECTIVES: To determine the safety and feasibility of autologous olfactory mucosal transplantation into the spinal cord in chronic spinal cord injured using the technique developed by Carlos Lima et al. SETTING: Spinal Injury Center, New Delhi. METHODS: Five chronic, motor complete, traumatic spinal cord injury (SCI) patients with neurological level C5-T12 underwent the procedure. Participants were assessed at baseline and at 6 monthly intervals. Safety and tolerability were evaluated through monitoring for any adverse events and tests including magnetic resonance imaging (MRI) evaluation. Efficacy assessment was done through neurological, functional and psychological evaluation, electrophysiological studies and urodynamics. RESULTS: Surgery was tolerated well by all American Spinal Injury Association (ASIA) Impairment Scale (AIS) A participants. The only AIS B participant lost sensory scores significantly after surgery but is gradually regaining it. MRI evaluation revealed a syrinx in one participant and increase in length of myelomalacia in four participants. There were no other adverse findings on MRI evaluation. There was no significant improvement in any of the neurological, electrophysiological or urodynamic efficacy variables. Statistically significant improvement was seen in functional scores as evaluated by Spinal Cord Independence Measure, Beck Depression Inventory scores and life impact scores on International Spinal Cord Injury Scale. CONCLUSIONS: The procedure is relatively safe and feasible in AIS A participants with thoracic level injuries at 18 month follow-up. No efficacy could be demonstrated which could be attributed to the procedure.

A simplicity-based fuzzy clustering approach for detection and extraction of murmurs from the phonocardiogram.

Almost all cardiac abnormalities manifest themselves as murmurs in a phonocardiogram (PCG). The location of a murmur in the PCG over a cardiac cycle depends on the underlying cardiac abnormality. Locating murmurs with respect to the cardiac cycle is useful for diagnosing cardiac dysfunction. Locating murmurs is difficult due to spectral similarity and time overlap with other heart sounds. Moreover, the wide variation in murmur amplitudes and murmur spectral characteristics across different patients and abnormalities has hindered the design of a generic time segmentation algorithm to isolate murmurs within the PCG. In this paper, we present a method to locate cardiac murmurs within the PCG that is based on their visual simplicity, which does not depend upon their absolute amplitude and frequency characteristics, and hence, results in their better localization. Then, we identify fuzzy sets to cluster simplicity values, due to murmurs, to determine the time duration over which the murmur occurs. The overall accuracy of the proposed algorithm in detecting systolic murmurs is 80%. The sensitivity of the algorithm in locating systolic murmurs is 73% and its specificity is 100%. The isolated murmur can then be further processed to extract clinically relevant features to diagnose the nature of the cardiac abnormality. Experimental results with a variety of murmurs are provided.

GATA-4 and Nkx-2.5 coactivate Nkx-2 DNA binding targets: role for regulating early cardiac gene expression.

The cardiogenic homeodomain factor Nkx-2.5 and serum response factor (SRF) provide strong transcriptional coactivation of the cardiac alpha-actin (alphaCA) promoter in fibroblasts (C. Y. Chen and R. J. Schwartz, Mol. Cell. Biol. 16:6372-6384, 1996). We demonstrate here that Nkx-2.5 also cooperates with GATA-4, a dual C-4 zinc finger transcription factor expressed in early cardiac progenitor cells, to activate the alphaCA promoter and a minimal promoter, containing only multimerized Nkx-2.5 DNA binding sites (NKEs), in heterologous CV-1 fibroblasts. Transcriptional activity requires the N-terminal activation domain of Nkx-2.5 and Nkx-2.5 binding activity through its homeodomain but does not require GATA-4's activation domain. The minimal interactive regions were mapped to the homeodomain of Nkx-2.5 and the second zinc finger of GATA-4. Removal of Nkx-2.5's C-terminal inhibitory domain stimulated robust transcriptional activity, comparable to the effects of GATA-4 on wild-type Nkx-2.5, which in part facilitated Nkx-2.5 DNA binding activity. We postulate the following simple model: GATA-4 induces a conformational change in Nkx-2.5 that displaces the C-terminal inhibitory domain, thus eliciting transcriptional activation of promoters containing Nkx-2.5 DNA binding targets. Therefore, alphaCa promoter activity appears to be regulated through the combinatorial interactions of at least three cardiac tissue-enriched transcription factors, Nkx-2.5, GATA-4, and SRF.

Cardiac tissue enriched factors serum response factor and GATA-4 are mutual coregulators.

Combinatorial interaction among cardiac tissue-restricted enriched transcription factors may facilitate the expression of cardiac tissue-restricted genes. Here we show that the MADS box factor serum response factor (SRF) cooperates with the zinc finger protein GATA-4 to synergistically activate numerous myogenic and nonmyogenic serum response element (SRE)-dependent promoters in CV1 fibroblasts. In the absence of GATA binding sites, synergistic activation depends on binding of SRF to the proximal CArG box sequence in the cardiac and skeletal alpha-actin promoter. GATA-4's C-terminal activation domain is obligatory for synergistic coactivation with SRF, and its N-terminal domain and first zinc finger are inhibitory. SRF and GATA-4 physically associate both in vivo and in vitro through their MADS box and the second zinc finger domains as determined by protein A pullout assays and by in vivo one-hybrid transfection assays using Gal4 fusion proteins. Other cardiovascular tissue-restricted GATA factors, such as GATA-5 and GATA-6, were equivalent to GATA-4 in coactivating SRE-dependent targets. Thus, interaction between the MADS box and C4 zinc finger proteins, a novel regulatory paradigm, mediates activation of SRF-dependent gene expression.

Effect of core lipopolysaccharides from Salmonella minnesota R mutants on the survival times of mice bearing Ehrlich tumor.

Injection of a homologous series of bacterial core lipopolysaccharides obtained from Salmonella minnesota R mutants to Ehrlich solid tumor-beating mice results in an increase of survival times of treated animals. Lower chain length favors greater antitumor activity. Oligosaccharides and polysaccharides derived from lipopolysaccharides were found to be ineffective in increasing survial time of tumore-bearing mice. Smaller-sugar-chain-length core lipopolysaccharides were found to be better adjuvant than were those with longer sugar chains. Implication of adjuvant action of lipopolysaccharides in the elicitation of antitumor activity is suggested.

Maltose tetrapalmitate, a nontoxic immunopotentiator with antitumor activity.

The attempt to synthesize a lipid A-like component (the active portion of lipopolysaccharides, but lacking its endotoxic activity) resulted in the production of fatty acyl sugars of which maltose tetrapalmitate was seen to yield the most promising results. It shows no endotoxic activity and elicits an antitumor response in tumor-transplanted animals as shown by (a) an enhancement of the host's capacity to reject a large number of tumor cells, (b) retardation of growth in tumor size, and (c) induction of hemorrhagic necrosis in certain tumors. Experiments with mammary ascites carcinoma show maltose tetrapalmitate to be as effective as is bacterial glycolipid mR595 in its antitumor activity. The degree of sensitivity to maltose tetrapalmitate varies with the tumor-host system: mammary ascites carcinoma less than NH = Cl2TSV5S = B16 less than L26. The mode of action of maltose tetrapalmitate appears to be via its modulation of the immune system. It is itself noncytotoxic to tumor cells in vitro. It is seen to stimulate the spleen cells of certain animals mitogenically, although it causes tumor rejection in all the types of animals tested. Also, it activates peritoneal exudate macrophages in tumor-bearing animals; whether specifically or nonspecifically has not yet been established.

Orthotopic implantation of primary N-[4-(5-Nitro-2-furyl)-2-thiazolyl]formamide-induced bladder cancer in bladder submucosa: an animal model for bladder cancer study.

Primary bladder tumors induced in Fischer 344 inbred rats by N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide were transplanted in syngeneic rats by the intravesical, s.c., i.v., and orthotopic routes. Attempts were made to establish bladder cancer cell lines in vitro. No success was achieved in transplantation by either the s.c., i.v., or intravesical routes when primary tumor cells were transplanted as cell suspensions. Cell suspensions of primary tumors also failed to grow in culture. However, orthotopic implantation into the bladder submucosa gave 45% success. Tumor fragments obtained from either the primary tumor or its lung metastases resulted in 10.6 and 36% tumor takes, respectively, when implanted s.c. However, after one orthotopic passage in the bladder submucosa, the tumor cells injected as cell suspension grew s.c. in 14% and orthotopically in 79% of the animals. Tumor fragments obtained from orthotopic tumors and implanted s.c. resulted in 15% tumor takes. After the second orthotopic passage, tumor cells could be grown in cultures and orthotopically in 100% of animals. The technique of orthotopic implantation as well as the usefulness of this tumor model for bladder cancer studies are described.

Combined cyclophosphamide chemotherapy and maltose tetrapalmitate immunotherapy in the treatment of transplanted bladder and prostate carcinoma of the rat.

Chemoimmunotherapy in two animal models for urological cancers was studied. The models were Dunning R3327A prostatic carcinoma transplanted s.c. in Fischer X Copenhagen F1 hybrids and a well-differentiated bladder carcinoma transplanted orthotopically in the bladder submucosa of female Fischer rats. Cyclophosphamide, cis-platinum, and Adriamycin were initially used as anticancer chemotherapeutic agents, and the most effective ones were used in combination with maltose tetrapalmitate (MTP), which was used as an immunopotentiator. In the case of prostatic carcinoma, cyclophosphamide was the most effective among the anticancer agents in controlling tumor growth after inoculation of either 10(4) or 10(5) tumor cells. Combination of cyclophosphamide with i.p. MTP delayed tumor take and controlled tumor size more effectively than did either of the treatments given alone. Similar results were obtained in the case of bladder tumor. A combination of cis-platinum with MTP significantly controlled bladder tumor size, and a combination of cyclophosphamide with MTP cured 75% of the rats. The remaining 25% of this group had a small tumor that did not increase in size during the subsequent 2 weeks of observation without treatment. The incidence of metastasis of bladder tumor to lymph nodes and lung was reduced by MTP and cis-platinum and eliminated by cyclophosphamide alone and in combination with MTP. Nonspecific immunity as measured by phytohemagglutinin stimulation of spleen lymphocytes and antitumor immunity as measured by cytotoxicity and macrophage migration inhibition assays were highest in rats subjected to cyclophosphamide and MTP combined therapies.

Single-step conversion of cephalosporin-C to 7-aminocephalosporanic acid by free and immobilized cells of Pseudomonas diminuta.

7-Aminocephalosporanic acid (7-ACA), the starting material for the production of a number of clinically used semisynthetic cephalosporins, is produced by deacylation of cephalosporin-C. The production of 7-ACA was studied in various modes, at the optimal conditions using free and immobilized whole cells of Pseudomonas diminuta.

Combination of 60Co gamma-radiation, misonidazole, and maltose tetrapalmitate in the treatment of Dunning prostatic tumor in the rat.

Maltose tetrapalmitate (MTP), a synthetic nontoxic immunoadjuvant, the radiosensitizer misonidazole (MISO), and 60Co gamma-radiation, alone or in combination, were used in the management of Dunning prostatic tumor in the rat. Nine groups of 10 rats each were used to assess the efficacy of various therapeutic modalities. Tumor growth rates and animal survival times were determined for each group. Radiation was more effective when combined with MTP, but the adjuvant must be present when radiation is given for synergism to occur. MISO was as effective as MTP when used with radiation, but combining them cancels out their individual effects. In a clinical situation it would be advantageous to use separately the synergisms existing between MISO and radiation on the one hand and MTP and radiation on the other hand.

Systemic, bladder wall, and bladder tumor concentration of misonidazole following intravesical administration in the rat.

The hypoxic-cell radiosensitizer misonidazole was administered intravesically to normal and to bladder tumor-bearing female Fischer rats. Drug concentration was measured in the bladder wall, the tumor and in the serum using high pressure liquid chromatography at different times following administration. The data shows that misonidazole is readily adsorbed by the bladder wall and the tumor with tissue levels reaching up to 10 times those measured in the serum.

Increased survival of CD1 mice bearing dimethylhydrazine induced primary colon and anal cancers by difluoromethylornithine with concomitant increase in angiosarcoma incidence.

Sixty CDl mice received dimethylhydrazine 20 mg/kg s.c. once weekly for 26 weeks to induce colorectal cancer. At this time the animals harbored frank colorectal cancer and early epidermoid cancer. The animals were divided into six groups that were subjected to the following treatments: none, MTP immunotherapy (MTP) alone, radiotherapy (R) alone, difluoromethylornithine (DFMO) chemotherapy alone and combinations of R+DFMO and R+DFMO+MTP. Criteria of evaluation of treatment efficacy were: number of colorectal tumor lesion and their staging at death, the incidence and size of anal cancer at death and survival time. Radiotherapy alone was marginally effective and MTP treatment was moderately effective in preventing anal cancer and reducing the number of colorectal tumors as well as their size. DFMO was exceptional in preventing anal cancer in a majority of animals and increasing animal survival; the latter effect was due to its preventive action against pyelonephritis, the major cause for animal death. However, in DFMO treated animals, the incidence of angiosarcoma increased from 10-16% (in the absence of DFMO) to 35-50% (in the presence of DFMO). The most effective treatment of the colorectal tumor was the triple combination of R + DFMO + MTP.

Prophylactic antiangiogenic tumor treatment.

Complementary antitumor treatments are required to increase the cure rate achieved by surgery and/or radiotherapy by avoiding future recurrences and metastases. The growth of most solid tumors, particularly carcinomas, depends upon the simultaneous development of internal tumor vasculature to allow the proliferation of tumor cells. Inhibition of tumor vascularization is an indirect means of limiting tumor expansion. Daily administration of cortisone and maltose tetrapalmitate (MTP) abolished growth of implanted syngeneic C3HBA mammary tumor. Gross and macroscopic examination of these tumors revealed that tumor growth was prevented. Histological examination demonstrated lack of vascularization within the neoplastic tissue. We believe that this combination in an appropriate form could provide a prophylactic treatment regimen after conventional antitumor treatments in humans.

Effect of maltose tetrapalmitate on tumor vascularization and tumor growth.

C3HBA mammary tumor (H-2k) was implanted s.c. by trocar in MTP responder C3H/HeN (H-2k) and non-responder C3H/HeJ (H-2k) female mice. One half of the animals received MTP i.p. The size of the tumor was measured everyday. Tumor growth was slightly slower in the C3H/HeJ than in the C3H/HeN. MTP treatment was effective only for the tumor implanted in the C3H/HeN. Microscopic and microscopic visualization of the tumor 1, 2, 3, and 15 days after tumor implantation and MTP treatment revealed poor vascular development in the MTP-treated C3H/HeN compared to untreated controls at days 2 and 3.

Evaluation of cortisone-heparin and cortisone-maltose tetrapalmitate therapies against rodent tumors. II. Pathological studies.

Pathological studies were undertaken in three tumor-host models which were subjected to cortisone based treatments. The first model was Fisher 344 rats with established orthotopically implanted syngeneic bladder tumor. Cortisone-herapin and cortisone-maltose tetrapalmitate (MTP) treatments induced focal areas of tumor necrosis and necrobiosis, whereas cortisone alone caused necrobiosis. The second model was C3HBA mammary tumor fragments implanted subcutaneously in syngeneic MTP responder C3H/HeN and MTP non-responder C3H/HeJ female mice. Only cortisone-MTP treatment led to an absence of capillary extension from surrounding blood vessels into the scant tumor stroma. The third model, ethyl carbamate induced primary lung cancer in AJ mice, was tested only with cortisone-herapin combination. The treatment caused central zones of necrosis.