Absence of multidrug resistance in Salmonella enterica serotypes Typhi and Paratyphi A isolates with intermediate susceptibility to ciprofloxacin.

BACKGROUND: We describe the antimicrobial susceptibility pattern of 64 blood stream isolates of Salmonella enterica serotypes Typhi and Paratyphi A studied from January 2013 to December 2014 at a tertiary care centre in North India. METHODS: Isolates were identified by standard biochemical reactions and confirmed by slide agglutination using specific antisera. Antimicrobial susceptibility testing was performed by Kirby-Bauer disc diffusion method and by E-test. RESULTS: In this study, 92% (46/50) of Salmonella Typhi and all Paratyphi A (n=14) isolates were susceptible to ampicillin, chloramphenicol and cotrimoxazole. Eighty percent of Typhi (40/50) and 64% (9/14) of Paratyphi A were intermediately susceptible to ciprofloxacin. Nineteen percent (12/64) of isolates were resistant to ciprofloxacin. No resistance to ceftriaxone and azithromycin was detected. CONCLUSIONS: Our study adds to the current knowledge of world-wide reports of multidrug resistance in S. Typhi.

Changing microbiological profile of pathogenic bacteria in diabetic foot infections: time for a rethink on which empirical therapy to choose?

AIMS/HYPOTHESIS: We studied the bacterial aetiology and antibiotic sensitivity pattern of diabetic foot ulcers in India. METHODS: Records of 447 hospitalised patients between 1991 and 2008 were retrospectively analysed between two time periods (before and after 1999) to compare bacterial aetiology and antimicrobial sensitivity patterns. The first three consecutive cultures from the same wound during treatment were evaluated. RESULTS: Of 1,632 cultures, 66% were polymicrobial, 23% monomicrobial and 11% sterile. In the monomicrobial group, 14% (n = 228) of cultures were Gram-negative, whereas 9% (n = 147) were Gram-positive. The most common pathogens in the first culture were Pseudomonas aeruginosa (20.1%), Staphylococcus aureus (17.2%) and Escherichia coli (16.3%). Results for the third cultures showed persistence of P. aeruginosa (15.3%) and E. coli (14.2%). Gram-negative isolates dominated over Gram-positive ones (25.3% vs 15.1%, p < 0.05). Antibiotic sensitivity patterns before and after 1999 were: piperacillin-tazobactam 74% vs 66% (p < 0.005), imipenem 77% vs 85% (NS), cefoperazone-sulbactam 47% vs 44% (p < 0.005), amikacin 62% vs 78% (NS), ceftriaxone 41% vs 36% (p < 0.005), amoxicillin-clavulanate 51% vs 43% (p < 0.05) and clindamycin 43% vs 36% (p < 0.005), respectively. CONCLUSIONS/INTERPRETATION: Unlike in the West, in India Gram-negative bacteria were found to have always been dominant in the wounds of patients with diabetic foot infections. Infection with polymicrobial multidrug-resistant Gram-negative bacilli is common. The policy of empirical antimicrobial therapy at tertiary care needs to be changed.

Assessment of white matter damage in subacute sclerosing panencephalitis using quantitative diffusion tensor MR imaging.

BACKGROUND AND PURPOSE: Subacute sclerosing panencephalitis (SSPE), a rare progressive degenerative disease, is caused by persistent infection with a defective measles virus. The correlation between the clinical staging and MR imaging is usually poor. The aim of the study was to investigate the role of diffusion tensor imaging (DTI) in the early detection of white matter damage in SSPE in the presence of normal findings on conventional imaging. METHODS: DTI was performed in 21 patients in stage II SSPE and 10 age/sex-matched healthy controls. Fractional anisotropy (FA) and mean diffusivity (MD) values were calculated in the periventricular white matter, corpus callosum, and posterior limb of the internal capsule in patients with normal and abnormal findings on conventional imaging as well as healthy controls. RESULTS: The patients were grouped into those with normal (n = 11) and abnormal (n = 10) findings on conventional imaging for the purpose of quantitative DTI analysis. Abnormal- and normal-appearing white matter on T2-weighted images showed significantly decreased FA values in all the regions compared with those in healthy controls. MD values were significantly increased in the periventricular white matter region of the frontal and parietooccipital lobe in patients with normal as well as abnormal findings on conventional imaging compared with those in healthy controls. CONCLUSION: DTI detects early white matter abnormalities that may have significant therapeutic implication, even in the presence of normal findings on conventional imaging, in patients with SSPE.

Suboptimal chlorine treatment of drinking water leads to selection of multidrug-resistant Pseudomonas aeruginosa.

The present study was undertaken to investigate the spectrum of bacteria present in the River Gomti water before and after chlorination for drinking purposes. We observed that the strains of Pseudomonas aeruginosa that survived chlorination on three out of seven occasions were resistant to almost all the antibiotics tested. The chlorine-resistant bacteria had mucoid colonies and grew better at 24 degrees C. All attempts to isolate the plasmid responsible for chlorine resistance were unsuccessful. Laboratory experiments using different strains of the P. aeruginosa in distilled water showed that only the resistant strain survived chlorine treatment at a dose of < or =500 microg/L. Similar results were obtained when water collected from seven different sites on the River Gomti was treated with graded doses of chlorine. At the higher dose of chlorine, all the bacteria died in 30 min, whereas with lower doses all the bacteria survived. The present study underscores the importance of measuring water chlorine concentrations to assure they are sufficiently high to remove pathogenic bacteria from drinking water. To our knowledge, this is the first report in the literature of the selection of multidrug-resistant bacteria by suboptimal chlorine treatment of water.

Pros and cons of antioxidant use during radiation therapy.

Radiation therapy is one of the major treatment modalities in the management of human cancer. While impressive progress like more accurate dosimetry and more precise methods of radiation targeting to tumor tissue has been made, the value of radiation therapy in tumor control may have reached a plateau. At present, two opposing hypotheses regarding the use of antioxidants during radiation therapy have been proposed. One hypothesis states that supplementation with high doses of multiple micronutrients including high dose dietary antioxidants (vitamins C and E, and carotenoids) may improve the efficacy of radiation therapy by increasing tumor response and decreasing some of its toxicity on normal cells. The other hypothesis suggests that antioxidants (dietary or endogenously made) should not be used during radiation therapy, because they would protect cancer cells against radiation damage. Each of these hypotheses is based on different conceptual frameworks that are derived from results obtained from specific experimental designs, and thus, each may be correct within its parameters. The question arises whether any of these concepts and experimental designs can be used during radiation therapy to improve the management of human cancer by this modality. This review has analyzed published data that are used in support of each hypothesis, and has revealed that the current controversies can be resolved, if the results obtained from one experimental design are not extrapolated to the other. This review has also discussed the scientific rationale for a micronutrient protocol that includes high doses of dietary antioxidants (vitamin C, vitamin E succinate and natural beta-carotene) which can be used adjunctively with radiation therapy.

Scientific rationale for using high-dose multiple micronutrients as an adjunct to standard and experimental cancer therapies.

We have hypothesized that high-dose multiple micronutrients, including antioxidants, as an adjunct to standard (radiation therapy and chemotherapy) or experimental therapy (hyperthermia and immunotherapy), may improve the efficacy of cancer therapy by increasing tumor response and decreasing toxicity. Several in vitro studies and some in vivo investigations support this hypothesis. A second hypothesis is that antioxidants may interfere with the efficacy of radiation therapy and chemotherapy. This hypothesis is based on the concept that antioxidants will destroy free radicals that are generated during therapy, thereby protecting cancer cells against death. None of the published data on the effect of antioxidants in combination with radiation or chemotherapeutic agents on tumor cells supports the second hypothesis. Scientific rationale in support of a micronutrient protocol to be used as an adjunct to standard or experimental cancer therapy is presented.

Multiple antioxidants in the prevention and treatment of Parkinson's disease.

Parkinson's disease (PD) is one of the major progressive neurological disorders for which no preventative or long-term effective treatment strategies are available. Epidemiologic studies have failed to identify specific environmental, dietary or lifestyle risk factors for PD except for toxic exposure to manganese, meperidine (Demerol, the "designer drug" version of which often contains a toxic byproduct of the synthesis, 1-methyl-4-phenyl 1,2,3,6 tetrahydropyridine [MPTP]), and some herbicides and pesticides. The search for genetic risk factors such as mutation, overexpression or underexpression of nuclear genes in DA neurons in idiopathic PD has not been successful as yet. Polymorphism in certain genes appears to be a risk factor, but there is no direct evidence for the causal relationship between polymorphism and increased risk of PD. In familial PD, mutation in the alpha-synuclein gene is associated with the disease, but a direct role of this gene in degeneration of DA neurons remains to be established. Although mutations in the Parkin gene has been associated with autosomal recessive juvenile Parkinson's disease, the role of this gene mutation in causing degeneration of DA neurons has not been defined. We have reported that in hereditary PD, a mutation in the alpha-synuclein gene may increase the sensitivity of DA neurons to neurotoxins. We hypothesize that, in idiopathic PD, epigenetic (mitochondria, membranes, protein modifications) rather than genetic events are primary targets which, when impaired, initiate degeneration in DA neurons, eventually leading to cell death. Although the nature of neurotoxins that cause degeneration in DA neurons in PD is not well understood, oxidative stress is one of the intermediary risk factors that could initiate and/or promote degeneration of DA neurons. Therefore, supplementation with antioxidants may prevent or reduce the rate of progression of this disease. Supplementation with multiple antioxidants at appropriate doses is essential because various types of free radicals are produced, antioxidants vary in their ability to quench different free radicals and cellular environments vary with respect to their lipid and aqueous phases. L-dihydroxyphenylalanine (L-dopa) is one of the agents used in the treatment of PD. Since L-dopa is known to produce free radicals during its normal metabolism, the combination of L-dopa with high levels of multiple antioxidants may improve the efficacy of L-dopa therapy.

High doses of multiple antioxidant vitamins: essential ingredients in improving the efficacy of standard cancer therapy.

Numerous articles and several reviews have been published on the role of antioxidants, and diet and lifestyle modifications in cancer prevention. However, the potential role of these factors in the management of human cancer have been largely ignored. Extensive in vitro studies and limited in vivo studies have revealed that individual antioxidants such as vitamin A (retinoids), vitamin E (primarily alpha-tocopheryl succinate), vitamin C (primarily sodium ascorbate) and carotenoids (primarily polar carotenoids) induce cell differentiation and growth inhibition to various degrees in rodent and human cancer cells by complex mechanisms. The proposed mechanisms for these effects include inhibition of protein kinase C activity, prostaglandin E1-stimulated adenylate cyclase activity, expression of c-myc, H-ras, and a transcription factor (E2F), and induction of transforming growth factor-beta and p21 genes. Furthermore, antioxidant vitamins individually or in combination enhance the growth-inhibitory effects of x-irradiation, chemotherapeutic agents, hyperthermia, and biological response modifiers on tumor cells, primarily in vitro. These vitamins, individually, also reduce the toxicity of several standard tumor therapeutic agents on normal cells. Low fat and high fiber diets can further enhance the efficacy of standard cancer therapeutic agents; the proposed mechanisms for these effects include the production of increased levels of butyric acid and binding of potential mutagens in the gastrointestinal tract by high fiber and reduced levels of growth promoting agents such as prostaglandins, certain fatty acids and estrogen by low fat. We propose, therefore, a working hypothesis that multiple antioxidant vitamin supplements together with diet and lifestyle modifications may improve the efficacy of standard and experimental cancer therapies.

Cancer prevention studies: past, present, and future directions.

In spite of extensive research on vitamins and diet, a consistent beneficial role of vitamin supplements, together with diet modification in human cancer prevention, has not been demonstrated. Published results of human intervention trials with vitamin supplements have been contradictory. This review critically, but briefly, evaluates (a) current concepts of human carcinogenesis, (b) effects of vitamins on biochemical parameters that are pertinent to cancer prevention, and (c) whether past or current protocols for intervention trials among high-risk populations adopt specific scientific rationales that are based on laboratory and human epidemiology studies. In addition, we propose a novel experimental design for intervention trials among high-risk human populations that is based on sound scientific principles derived from laboratory and human epidemiologic data on vitamins, diet, lifestyle, and cancer prevention. Such trials would answer a fundamental public health issue of today: Does supplementation with multiple vitamins, together with diet and lifestyle modifications, reduce the risk of cancer?

Vitamins regulate gene expression and induce differentiation and growth inhibition in cancer cells. Their relevance in cancer prevention.

Although several hypotheses for human carcinogenesis have been proposed, the specific genetic changes that cause normal cells to become cancer cells have not been identified. In spite of uncertainties regarding the mechanisms of carcinogenesis, several vitamins such as beta-carotene and vitamins A, C, and E, which can reduce the risk of cancer, have been identified, using animal and in vitro models of carcinogenesis. These studies have led to a hypothesis that the supplemental intake of these vitamins may reduce the risk of cancer. This hypothesis in humans can be tested only by intervention trials that are in progress. Prospective and retrospective case-controlled experimental designs are not suitable for testing the above hypothesis. The fact that some vitamins induce cell differentiation and/or growth inhibition in tumor cells in culture suggests that the use of these vitamins in cancer prevention has a cellular basis. In addition to having a direct effect on tumor cells, vitamins such as alpha-tocopheryl succinate and beta-carotene enhance the effect of other agents that induce differentiation in tumor cells. Some vitamins like beta-carotene, retinoic acid, alpha-tocopheryl succinate, and vitamin D also regulate the expressions of certain oncogenes and cellular genes. These are exciting new functions of vitamins that nobody could have predicted only a few years ago.

Vitamin E and cancer prevention: recent advances and future potentials.

Many animal and in vitro experiments have shown that the supplementation of diet with vitamin E within a certain dose range reduced the risk of chemical- and radiation-induced cancers. In vitro studies revealed that alpha-tocopheryl succinate (TS) induced differentiation and growth-inhibition in certain animal and human tumor cells in culture, whereas alpha-tocopherol (alpha-T), alpha-tocopheryl acetate (alpha-TA) and alpha-tocopheryl nicotinate (alpha-TN) were ineffective, alpha-TS also reduced basal and ligand-stimulated adenylate cyclase activity, and expression of c-myc and H-ras oncogenes in certain tumor cells in culture. The relative efficacy of various forms of vitamin E in cancer prevention in animal or human models has not been evaluated. Human epidemiologic studies utilizing retrospective and prospective case-control experimental designs are not suitable for evaluating the role of vitamin E in cancer prevention due to several inherent problems associated with these methodologies. Intervention trials utilizing vitamin E with appropriate biological and statistical rationales are most suitable for testing the role of vitamin E in cancer prevention in humans. Some human trials utilizing vitamin E alone or in combination with other nutrients are in progress.

Establishment of primary cultures of rat and human parotid epithelial cells for transfection experiments.

The molecular mechanisms that regulate the synthesis of salivary proteins are unknown. The paucity of homogeneous cell populations of parotid acinar cells has become a limiting factor for such a study. Therefore, the establishment of immortalized clones of acinar cells is essential. This study has established primary cultures of rat and human parotid epithelial cells that are suitable for transfection with plasmid vectors, pSV2, pSV3, and pSV5 to generate immortalized cells in vitro. Among various techniques used, the rat and human parotid tissue or cellular clumps when restrained in chicken plasma clot allowed the outgrowth of epithelial cells that maintained epithelial cell morphology for over 4 wk. However, the initial growth requirements for rat and human parotid cells were different. The presence of 10% heat inactivated fetal bovine serum in supplemented MCDB-LB medium was essential for the outgrowth of rat parotid epithelial cells, but this was not needed for the outgrowth of human parotid epithelial cells. The growth of both human and rat parotid epithelial cells can be maintained in serum-free supplemented MCDB-LB. These primary cultures contained amylase-producing cells as demonstrated by immunofluorescent technique, and they were transfected with pSV2, pSV3, and pSV5 using primarily the calcium phosphate-DNA co-precipitation technique. After initial extensive cell death, many cells with epithelial cell morphology survived.

Ayurvedic (science of life) agents induce differentiation in murine neuroblastoma cells in culture.

Many Indian Ayurvedic (science of life) agents have been introduced into the U.S.A. as food supplements. Two of them, Maharishi Amrit Kalash-Ambrosia (MAK-A) and Maharishi Amrit Kalash-Nectar (MAK-N) are under investigation. This study shows that an ethanol extract of MAK-A induced morphological (neurite formation) and biochemical (increase of activity of tyrosine hydroxylase by about 15-fold) differentiation in murine neuroblastoma (NBP2) cells in culture, whereas an aqueous extract of MAK-A increased only the activity of tyrosine hydroxylase but to a much lesser extent. The treatment time of 3 days was needed for the expression of maximum differentiation. Ethanol extracts of MAK-A and aqueous extracts of MAK-A increased the intracellular level of adenosine 3',5'-cyclic monophosphate (cAMP) by about 4-fold in 3 days but they did not do so in 15 min. Ethanol extracts of MAK-A also induced neurite formation in neuroblastoma cells grown in serum free medium but the concentration requirement was about a fifth of that needed in serum. The treatment time of 24 hr was sufficient to induce optimal differentiation in neuroblastoma cells grown in serum free medium. The differentiating agents in ethanol-MAK-A were resistant to heat and light and could not be removed by treatment with activated charcoal. Neither ethanol-MAK-N nor aqueous-MAK-N induced differentiation in neuroblastoma cells, suggesting that the differentiating agents were present only in MAK-A.

Modification of the hyperthermic response on neuroblastoma cells by cAMP and sodium butyrate.

The role of adenosine 3',5'-cyclic monophosphate (cAMP) and sodium butyrate in modifying the effect of heat on murine neuroblastoma cells (NBP2) in culture was evaluated on the criterion of survival. An elevation of cellular cAMP level by prostaglandin (PG) A2, a stimulator of adenylate cyclase, and 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone (R020-1724), an inhibitor of cyclic nucleotide phosphodiesterase, only during heat treatment (43 degrees C and 40 degrees C) was sufficient to enhance the effect of heat. The extent of enhancement (additive versus synergistic) depended upon the cAMP stimulating agent and the experimental condition. When these agents were added after heat treatment for the entire observation period, they produced similar results. PGA2 and R020-1724 are known to increase the intracellular level of cAMP in these cells by three and fivefold, respectively; therefore, the effect of these agents in enhancing the heat-response may be mediated by cAMP-dependent mechanisms. The presence of sodium butyrate during heat treatment alone was ineffective; however, when it was added before or after heat treatment for the entire observation period, the survival of heated cell was markedly reduced.

Study on the specificity of alpha-tocopheryl (vitamin E) acid succinate effects on melanoma, glioma and neuroblastoma cells in culture.

d- and dl-alpha-tocopheryl succinate inhibited growth and caused morphological changes in mouse melanoma (B-16), mouse neuroblastoma (NBP2), and rat glioma (C-6) cells in culture. To study whether the effects of alpha-tocopheryl (vitamin E) succinate on tumor cells are mediated by antioxidant mechanisms, the effects of lipid-soluble antioxidants, butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) were compared with those of vitamin E succinate. Results showed that these antioxidants produced alterations on the growth and morphology of neuroblastoma, melanoma, and glioma cells which are similar to those produced by vitamin E succinate; however, the extent of the effect depended upon the type of antioxidant and the form of tumor cells. These data suggest that the effects of vitamin E succinate on tumor cells may be mediated, in part, by antioxidant mechanisms.

Effects of tocopherol (vitamin E) acid succinate on morphological alterations and growth inhibition in melanoma cells in culture.

The effects of various forms of tocopherol (vitamin E) on the growth and differentiation of mouse melanoma (B-16) and mouse fibroblast (L-cells) cells in culture were studied. D-alpha-tocopherol acid succinate induced morphological alterations and growth inhibition in melanoma cells. When vitamin E acid succinate was removed 4 days after treatment, the above changes remained irreversible for a period of 24 hr, after which resistant cells and partially affected cells renewed cell division and eventually reached confluency. The relative efficacy of D and DL forms of vitamin E acid succinate remains to be evaluated. However, other forms of vitamin E such as DL-alpha-tocopherol free alcohol, Aquasol DL-alpha-tocopherol acetate, DL-alpha-tocopherol nicotinate, or sodium succinate with an equivalent volume of ethanol, at similar concentrations, were ineffective. Vitamin E acid succinate at similar concentrations did not induce morphological changes in fibroblasts. Melanoma cells were about 2-fold more sensitive to vitamin E acid succinate than were fibroblasts for the criterion of growth inhibition. Vitamin E acid succinate-induced morphological changes and growth inhibition in melanoma cells were expressed in hormone-supplemented serum-free medium, but the concentration requirement was about 5 times less than that needed in serum-supplemented medium. Although cyclic adenosine 3': 5'-monophosphate-stimulating agents are known to cause growth inhibition and morphological changes in melanoma cells in culture, vitamin E acid succinate-induced morphological alterations in melanoma cells are no mediated by a rise in cellular cyclic adenosine 3':5'-monophosphate. Ethanol was sufficient to increase the melanin content in melanoma cells. These data show that vitamin E acid succinate may be a potentially useful tumor therapeutic agent.