A two-column flash chromatography approach to pyoverdin production from Pseudomonas putida GB1.

Our knowledge of the biological and environmental reactivity of siderophores is limited by the difficulty and cost of obtaining reasonable quantities by purification or synthesis. In this note, we describe a modified procedure for the low-cost, mg-scale purification of pyoverdin-type siderophores using a dual-flash chromatography (reverse-phase absorption and size exclusion) approach.

Combining Community-Based Participatory Research (CBPR) with a Random-Sample Survey to Assess Smoking Prevalence in an Under-Served Community.

ACKNOWLEDGMENTS: The authors would like to thank Laura McClure for her help with the manuscript submission, the Liberty City Community Health Advisory Board for its collaboration on this study, as well as the survey interviewers, and the survey participants. INTRODUCTION: Underserved communities might lag behind Healthy People 2010 objectives of smoking reduction because of smoking behavior disparities. This possibility was investigated through a random-sample survey conducted in a disenfranchised community in Miami-Dade County, Florida, using a Community-Based Participatory Research (CBPR) framework. The survey was triggered by our finding that this community had higher than expected incidence of tobacco-associated cancers. METHODS: Survey methods, resulting from a dialog between the Community Advisory Board and academic researchers, included: (a) surveying adult residents of a public housing complex located within the community; (b) probability sampling; (c) face-to-face interviews administered by trained community residents. 250 households were sampled from 750 addresses provided by the county Public Housing Agency. The completed surveys were reviewed by the academic team, yielding 204 questionnaires for the current analysis. RESULTS: Of the 204 respondents, 38% were current smokers. They estimated the percentages of smokers in their household and among their five best friends at 33% and 42%, respectively, and among adults and youth in the community at 72% and 53%, respectively. CONCLUSIONS: A mix of state-of-art methodology with CBPR principles is seldom encountered in the current literature. It allowed the research team to find a high smoking prevalence in an underserved community, twice the statewide and nationwide estimates. Similar or higher levels of smoking were perceived in respondent's entourage. Such disparity in smoking behavior, unlikely to result from self-selection bias because of our rigorous methodology, calls for community-specific tobacco control efforts commensurate to the magnitude of the problem.

Effects of exogenous pyoverdines on Fe availability and their impacts on Mn(II) oxidation by Pseudomonas putida GB-1.

Pseudomonas putida GB-1 is a Mn(II)-oxidizing bacterium that produces pyoverdine-type siderophores (PVDs), which facilitate the uptake of Fe(III) but also influence MnO2 formation. Recently, a non-ribosomal peptide synthetase mutant that does not synthesize PVD was described. Here we identified a gene encoding the PVDGB-1 (PVD produced by strain GB-1) uptake receptor (PputGB1_4082) of strain GB-1 and confirmed its function by in-frame mutagenesis. Growth and other physiological responses of these two mutants and of wild type were compared during cultivation in the presence of three chemically distinct sets of PVDs (siderotypes n°1, n°2, and n°4) derived from various pseudomonads. Under iron-limiting conditions, Fe(III) complexes of various siderotype n°1 PVDs (including PVDGB-1) allowed growth of wild type and the synthetase mutant, but not the receptor mutant, confirming that iron uptake with any tested siderotype n°1 PVD depended on PputGB1_4082. Fe(III) complexes of a siderotype n°2 PVD were not utilized by any strain and strongly induced PVD synthesis. In contrast, Fe(III) complexes of siderotype n°4 PVDs promoted the growth of all three strains and did not induce PVD synthesis by the wild type, implying these complexes were utilized for iron uptake independent of PputGB1_4082. These differing properties of the three PVD types provided a way to differentiate between effects on MnO2 formation that resulted from iron limitation and others that required participation of the PVDGB-1 receptor. Specifically, MnO2 production was inhibited by siderotype n°1 but not n°4 PVDs indicating PVD synthesis or PputGB1_4082 involvement rather than iron-limitation caused the inhibition. In contrast, iron limitation was sufficient to explain the inhibition of Mn(II) oxidation by siderotype n°2 PVDs. Collectively, our results provide insight into how competition for iron via siderophores influences growth, iron nutrition and MnO2 formation in more complex environmental systems.

Pyoverdine synthesis by the Mn(II)-oxidizing bacterium Pseudomonas putida GB-1.

When iron-starved, the Mn(II)-oxidizing bacteria Pseudomonas putida strains GB-1 and MnB1 produce pyoverdines (PVDGB-1 and PVDMnB1), siderophores that both influence iron uptake and inhibit manganese(II) oxidation by these strains. To explore the properties and genetics of a PVD that can affect manganese oxidation, LC-MS/MS, and various siderotyping techniques were used to identify the peptides of PVDGB-1 and PVDMnB1 as being (for both PVDs): chromophore-Asp-Lys-OHAsp-Ser-Gly-aThr-Lys-cOHOrn, resembling a structure previously reported for P. putida CFML 90-51, which does not oxidize Mn. All three strains also produced an azotobactin and a sulfonated PVD, each with the peptide sequence above, but with unknown regulatory or metabolic effects. Bioinformatic analysis of the sequenced genome of P. putida GB-1 suggested that a particular non-ribosomal peptide synthetase (NRPS), coded by the operon PputGB1_4083-4086, could produce the peptide backbone of PVDGB-1. To verify this prediction, plasmid integration disruption of PputGB1_4083 was performed and the resulting mutant failed to produce detectable PVD. In silico analysis of the modules in PputGB1_4083-4086 predicted a peptide sequence of Asp-Lys-Asp-Ser-Ala-Thr-Lsy-Orn, which closely matches the peptide determined by MS/MS. To extend these studies to other organisms, various Mn(II)-oxidizing and non-oxidizing isolates of P. putida, P. fluorescens, P. marincola, P. fluorescens-syringae group, P. mendocina-resinovorans group, and P. stutzerii group were screened for PVD synthesis. The PVD producers (12 out of 16 tested strains) were siderotyped and placed into four sets of differing PVD structures, some corresponding to previously characterized PVDs and some to novel PVDs. These results combined with previous studies suggested that the presence of OHAsp or the flexibility of the pyoverdine polypeptide may enable efficient binding of Mn(III).

Developing an urban community-campus partnership: lessons learned in infrastructure development and communication.

BACKGROUND: A low-income, African American neighborhood in Miami, Florida, experiences health disparities including an excess burden of cancer. Many residents are disenfranchised from the healthcare system, and may not participate in cancer prevention and screening services. OBJECTIVE: We sought to describe the development of a partnership between a university and this community and lessons learned in using a community-based participatory research (CBPR) model. METHODS: To better understand the community's health behaviors and status, a randomized door-to-door survey was conducted in collaboration with a community partner. LESSONS LEARNED: This collaboration helped foster a mutual understanding of the benefits of CBPR. We also describe challenges of adhering to study protocols, quality control, and sharing fiscal responsibility with organizations that do not have an established infrastructure. CONCLUSIONS: Understanding the organizational dynamics of a community is necessary for developing a CBPR model that will be effective in that community. Once established, it can help to inform future collaborations.

The molecular biogeochemistry of manganese(II) oxidation.

Micro-organisms capable of oxidizing the redox-active transition metal manganese play an important role in the biogeochemical cycle of manganese. In the present mini-review, we focus specifically on Mn(II)-oxidizing bacteria. The mechanisms by which bacteria oxidize Mn(II) include a two-electron oxidation reaction catalysed by a novel multicopper oxidase that produces Mn(IV) oxides as the primary product. Bacteria also produce organic ligands, such as siderophores, that bind to and stabilize Mn(III). The realization that this stabilized Mn(III) is present in many environments and can affect the redox cycles of other elements such as sulfur has made it clear that manganese and the bacteria that oxidize it profoundly affect the Earth's biogeochemistry.

The Response of Shewanella oneidensis MR-1 to Cr(III) Toxicity Differs from that to Cr(VI).

Chromium is a contaminant of concern that is found in drinking water in its soluble, hexavalent form [Cr(VI)] and that is known to be toxic to eukaryotes and prokaryotes. Trivalent chromium [Cr(III)] is thought to be largely harmless due to its low solubility and inability to enter cells. Previous work has suggested that Cr(III) may also be toxic to microorganisms but the mechanism remained elusive. In this work, we probe the toxicity of Cr(III) to Shewanella oneidensis MR-1, a bacterium able to reduce Cr(VI) to Cr(III) and compare it to Cr(VI) toxicity. We found evidence for Cr(III) toxicity both under Cr(VI) reducing conditions, during which Cr(III) was generated by the reduction process, and under non-reducing conditions, when Cr(III) was amended exogenously. Interestingly, cells exposed to Cr(III) (200 µM) experienced rapid viability loss as measured by colony forming units on Luria-Bertani (LB) agar plates. In contrast, they maintained some enzymatic activity and cellular integrity. Cr(VI)-exposed cells exhibited loss of enzymatic activity and cell lysis. The loss of viability of Cr(III)-exposed cells was not due to membrane damage or to enzymatic inhibition but rather appeared to be associated with an abnormal morphology that consisted of chains of membrane-enclosed units of irregular size. Exposure of abnormal cells to growth conditions resulted in membrane damage and cell death, which is consistent with the observed viability loss on LB plates. While Cr(VI) was taken up intracellularly and caused cell lysis, the toxic effect of Cr(III) appeared to be associated with extracellular interactions leading to an ultimately lethal cell morphology.

Menthol cigarette smoking and health, Florida 2007 BRFSS.

OBJECTIVE: To examine associations between menthol cigarette smoking and nicotine dependence, quit attempts, and physical and mental health. METHODS: Data were drawn from the 2007 Florida Behavioral Risk Factor Surveillance System (BRFSS) and a follow-up survey among current smokers (N = 3396). Univariate and multivariate logistic regression analyses were conducted. RESULTS: In multivariate analyses, menthol cigarette smoking was associated with women, African American and Hispanic race/ethnicity, and greater mental distress. CONCLUSIONS: Women, racial/ethnic minorities, and individuals reporting a greater frequency of mental distress are more likely to smoke menthol versus nonmenthol cigarettes. Implications for public health policy and cessation interventions are discussed.

Cancer screening behaviors among smokers and non-smokers.

OBJECTIVE: We explored whether smoking is associated with cancer screening behaviors. METHODS: We used data from the 2007 Florida Behavioral Risk Factor Surveillance System and the Florida Tobacco Callback Survey to examine screening behaviors related to four cancer types (breast, cervical, prostate, and colorectal). Using multiple logistic regression analyses, we examined the association between smoking status and health screening behaviors. RESULTS: For 10 of the 11 cancer screening variables, being a current smoker was significantly associated with being less likely to ever have been screened and also less likely to be compliant with screening guidelines. For breast and cervical cancer, level of nicotine dependence was also significantly related to compliance with screening recommendations; women with higher levels of dependence were less likely to be compliant. CONCLUSIONS: Our results support the notion that individuals' actions related to their health are consistent across different types of behaviors. We found that smokers were less likely to engage in cancer screening behaviors. In addition, among smokers, individuals with greater nicotine dependence had lower compliance with some screening tests. Physicians should ensure that their patients who smoke are receiving appropriate and adequate screening for cancer.

Disparity in melanoma: a trend analysis of melanoma incidence and stage at diagnosis among whites, Hispanics, and blacks in Florida.

OBJECTIVE: To examine and compare the temporal trends in melanoma incidence and stage at diagnosis among whites, Hispanics, and blacks in Florida from 1990 to 2004. DESIGN: Cross-sectional and retrospective analysis. SETTING: Florida Cancer Data System. PATIENTS: Melanoma cases with known stage and race/ethnicity reported from 1990 to 2004. MAIN OUTCOME MEASURES: Age-adjusted melanoma incidence and stage at diagnosis. RESULTS: Of 41 072 cases of melanoma, 39 670 cases were reported for white non-Hispanics (WNHs), 1148 for white Hispanics (WHs), and 254 for blacks. Melanoma incidence rates increased by 3.0% per year among WNH men (P < .001), 3.6% among WNH women (P < .001), 3.4% among WH women (P = .01), and 0.9% among WH men (P = .52), while remaining relatively stable among black men and women. Both WHs and blacks had significantly more advanced melanoma at presentation: 18% of WH and 26% of black patients had either regional or distant-stage melanoma at diagnosis compared with 12% of WNH patients. The proportion of distant-stage melanoma diagnosed among WHs and blacks changed little from 1990 to 2004, compared with a steady decrease in the percentage of melanoma cases diagnosed at distant stage among WNHs (P < .001). Such differences in the time trends of the proportion of distant-stage melanoma remained after excluding in situ cases. CONCLUSIONS: The rising melanoma incidence among WNHs and WHs emphasizes the need for primary prevention. The persistence of disparity in melanoma stage at diagnosis among WHs, blacks, and WNHs warrants closer examination of secondary prevention efforts in minority groups.

Correlates of smoking quit attempts: Florida Tobacco Callback Survey, 2007.

OBJECTIVE: The public health burden of tobacco-associated diseases in the USA remains high, in part because many people's attempts to quit are unsuccessful. This study examined factors associated with having lifetime or recent attempts to quit smoking among current smokers, based on a telephone survey of Florida adults. METHODS: Data from the 2007 telephone-based Florida Behavioral Risk Factor Surveillance System (BRFSS) and its follow-up survey, the Tobacco Callback Survey, were used to assess determinants of having ever attempted to quit smoking and attempted to quit smoking in the past 12 months. All analyses were conducted using SAS. RESULTS: Among 3,560 current smokers, 41.5% reported having tried to quit smoking in the past 12 months while 83.4% reported having ever tried to quit. Having a history of a tobacco-related medical condition was significantly associated with both recent (Adjusted Odds Ratio (AOR) 1.41 [Confidence Interval 1.19-1.65]) and lifetime quit attempts (AOR 1.43 [1.15-1.79]). Greater nicotine dependence and being advised by a physician to quit smoking were also positively associated with lifetime quit attempts. Receipt of healthcare provider advice to quit smoking in the past 12 months and a strong belief that quitting following a long history of regular smoking would not result in health benefits and belief that there are health benefits to quitting smoking were associated with lifetime quit attempts. CONCLUSION: Targeted smoking cessation interventions are needed for smokers with selected medical conditions and with high nicotine dependence. The importance of physician advice in encouraging individuals to quit is further highlighted.

Unequal burden of head and neck cancer in the United States.

BACKGROUND: Black Americans are adversely affected by many types of malignancies. METHODS: We reviewed data from the National Cancer Institute's Surveillance, Epidemiology, and End Results (SEER) program to evaluate racial disparities in head and neck cancer incidence, mortality, and survival. RESULTS: Head and neck cancer incidence is greater in the black population and peaks at a younger age. The incidence disparity is decreasing over time and is less for cancers of the oral cavity/pharynx (OCP) than for cancers of the larynx. The disparity in survival after diagnosis is substantial for both sites and is increasing over time because of improvement in survival for the white population, but not for the black population. Some, but not all, of the survival disparity is due to more advanced stage at the time of diagnosis within the black population. The age-adjusted mortality rate for black men is approximately twice the rate for white men. CONCLUSION: Black Americans clearly bear a greater burden from head and neck cancer. The underlying causes are largely unknown, but are most likely due to a complex interplay of differences in access to health care, quality of medical care, biologic/genetic factors, incidence of comorbid conditions, exposure to carcinogens, diet, and cultural beliefs. Prospective studies are needed to define the relative importance of these factors and to inform intervention strategies.

Understanding and meeting the needs of those using growth hormone injection devices.

BACKGROUND: Recombinant human growth hormone (r-hGH) is used to treat: growth hormone deficiency in children and adults; children born small for gestational age; Turner's syndrome; and chronic renal failure. r-hGH is administered by daily subcutaneous injection and may be given using a number of different administration devices. The aim of this survey was, firstly, to identify which attributes of an r-hGH administration device are considered most important to physicians, teenage patients, parents of young children requiring GH and nurses who have experience of r-hGH administration, and, secondly, to determine how they rate existing devices in each of these key attributes. METHODS: The opinions of 67 individuals with experience in r-hGH administration were captured in discussion sessions. Parents, physicians and nurses were asked to rate 19 device attributes by completing a questionnaire, and to rank four different r-hGH administration devices (including a conceptual electronic device) in order of preference. RESULTS: Reliability, ease of use, lack of pain during injection, safety in use, storage, and number of steps in preparation before use, during use and after were considered to be the five most desirable attributes of an r-hGH administration device. An electronic device was preferred to an automatic, multi-dose injection device, a needle-free injection device or a manual, ready-to-use, disposable injection device. CONCLUSION: In the opinion of physicians, nurses and parents using r-hGH injection devices, an ideal device must combine reliability with simplicity, while delivering treatment with minimal pain. An electronic device, which combines many of the most useful features of existing devices with novel functions, was the preferred option for r-hGH administration.

Comparison of stage at diagnosis of melanoma among Hispanic, black, and white patients in Miami-Dade County, Florida.

OBJECTIVE: To compare stage at diagnosis of melanoma between non-Hispanic white, non-Hispanic black, and Hispanic patients. DESIGN: Retrospective analysis. SETTING: Melanoma cases reported to the Florida Cancer Data System, with known stage and race/ethnicity information, for residents of Miami-Dade County, Florida, from 1997 to 2002. PATIENTS: Those diagnosed as having melanoma according to the Florida Cancer Data System. MAIN OUTCOME MEASURE: Stage of melanoma at diagnosis. RESULTS: Of the 1690 melanoma cases reported with both stage and race/ethnicity information, 1176 (70%) were among non-Hispanic white patients, 485 (29%) were among Hispanic patients of any race, and 29 (2%) were among non-Hispanic black patients. Late-stage (regional and distant) diagnosis was more common among Hispanic (26%) and non-Hispanic black patients (52%) compared with non-Hispanic white patients (16%) (P < .001). CONCLUSION: Advanced stage of melanoma diagnosis among Hispanic and black patients suggests suboptimal secondary prevention efforts in minority populations.

Sun protection policies in Miami-Dade County public schools: opportunities for skin cancer prevention.

Childhood exposure to ultraviolet radiation from the sun and a history of sunburns are risk factors for skin cancer. Because children spend time outdoors when they are at school, school sun protection policies are an important health issue, particularly in areas of the country with year-round warm and sunny climates, such as Florida. To better understand the sun protection policies and practices in South Florida schools, a sample (n = 51) of elementary and middle schools in Miami-Dade County public schools were surveyed as part of a CDC-funded cancer control program at the University of Miami. Of the principals and teachers surveyed, most (78%) knew about the county school system's guidelines for avoiding excessive heat exposure, which include two sun protection measures. Two-thirds reported that they shared these guidelines with teachers; 21% shared them with parents. Few schools monitor implementation of the guidelines, although 70% schedule outdoor activities to avoid peak sun hours. No schools required sunscreen, hats, or protective clothing. Physical education teachers and students spend an average of 4.5 and 0.6 hours per day outdoors, respectively. Improved school sun protection policies and monitoring of such policies is needed to reduce sun exposure and skin cancer risk for both students and staff.

Chemical Characterization of Polysaccharide from the Slime Layer of the Cyanobacterium Microcystis flos-aquae C3-40.

Macromolecular material from the slime layer of the cyanobacterium Microcystis flos-aquae C3-40 was defined as material that adhered to cells during centrifugation in growth medium but was dislodged by washing with deionized water and retained within dialysis tubing with a molecular-weight cutoff of 3,500. At each step of this isolation procedure, the slime was observed microscopically. Cells in the centrifugal pellet were surrounded by large amounts of slime that excluded negative stain, whereas cells that had been washed with water lacked visible slime. Two independently isolated lots of slime contained no detectable protein (<1%, wt/wt) and consisted predominantly of anthrone-reacting polysaccharide. Sugars in a hydrolysate of slime polysaccharide were derivatized with trimethylsilylimidazole and examined by gas chromatography-mass spectrometry. The composition of the slime polysaccharide was 1.5% (wt/wt) galactose, 2.0% glucose, 3.0% xylose, 5.0% mannose, 5.5% rhamnose, and 83% galacturonic acid. This composition resembles that of the plant polysaccharide pectin, which was treated in parallel as a control. Consistent with earlier indications that M. flos-aquae slime preferentially binds certain cations, the ratio of Fe to Na in the dialyzed slime was 10 times that in the growth medium. The composition of the slime is discussed with respect to possible mechanisms of cation binding in comparison with other cyanobacterial exopolysaccharides and pectin.