Free Essay

Anti-Legionella Activity of Staphylococcal Hemolytic Peptides

In: Science

Submitted By thesunightsong
Words 3768
Pages 16
Peptides 32 (2011) 845–851

Contents lists available at ScienceDirect

Peptides journal homepage:

Anti-Legionella activity of staphylococcal hemolytic peptides
A. Marchand, J. Verdon, C. Lacombe, S. Crapart, Y. Héchard, J.M. Berjeaud ∗
Université de Poitiers, Laboratoire de Chimie et Microbiologie de l’Eau – UMR 6008 CNRS, IBMIG – UFR Sciences Fondamentales et Appliquées, 1 rue du Georges Bonnet, 86022 Poitiers Cedex, France

a r t i c l e

i n f o

a b s t r a c t
A collection of various Staphylococci was screened for their anti-Legionella activity. Nine of the tested strains were found to secrete anti-Legionella compounds. The culture supernatants of the strains, described in the literature to produce hemolytic peptides, were successfully submitted to a two step purification process. All the purified compounds, except one, corresponded to previously described hemolytic peptides and were not known for their anti-Legionella activity. By comparison of the minimal inhibitory concentrations, minimal permeabilization concentrations, decrease in the number of cultivable bacteria, hemolytic activity and selectivity, the purified peptides could be separated in two groups. First group, with warnericin RK as a leader, corresponds to the more hemolytic and bactericidal peptides. The peptides of the second group, represented by the PSM from Staphylococcus epidermidis, appeared bacteriostatic and poorly hemolytic. © 2011 Elsevier Inc. All rights reserved.

Article history: Received 15 December 2010 Received in revised form 19 January 2011 Accepted 19 January 2011 Available online 1 February 2011 Keywords: Legionella pneumophila Staphylococci Hemolytic peptides

1. Introduction Legionella pneumophila is a waterborne pathogenic bacterium responsible for severe pneumonia called Legionnaire’s disease [6,17]. In the environment, L. pneumophila is ubiquitously found in fresh water and could survive within biofilms and free-living amoeba [2]. L. pneumophila could be found at high level in manmade water systems such as air conditioning and cooling towers, spas. These systems are mainly responsible for outbreaks as they might produce contaminated water droplets, which are inhaled by people. L. pneumophila reaches the lungs and multiplies within macrophages [15]. In 2006, there were more than 6000 reported cases in Europe leading to 400 deaths ca. Concerning L. pneumophila growth in water systems, various methods of control have been used: chlorine, monochloramine, heat. However, these treatments are not fully efficient and, after a lag period following the treatment, L. pneumophila might be able to recolonize the system [19]. Therefore, it is of primary importance to find new treatments to restrain L. pneumophila growth. Recently, it was shown that hemolytic peptides secreted by Staphylococcus warneri RK inhibited the growth of Legionella spp. [8]. Beside an original 22 amino acids long peptide named warnericin RK, two delta-hemolysins were characterized [20]. Hemolysins are among the most important virulence factors of Staphylococci. Staphylococcus aureus, which is a potentially pathogenic coagulase-positive specie of this genus, produces four

∗ Corresponding author. Tel.: +33 5 49 45 40 06; fax: +33 5 49 45 35 03. E-mail address: (J.M. Berjeaud). 0196-9781/$ – see front matter © 2011 Elsevier Inc. All rights reserved. doi:10.1016/j.peptides.2011.01.025

hemolysins – , , and [26]. -Hemolysin, also named -toxin or -lysin, is a well known peptide, which mechanism of action has been described [4,22]. -Hemolysins were found in most species of coagulase-negative Staphylococci and some sequences (Table 1) were characterized for S. intermedius [9], S. epidermidis [12], S. simulans and S. warneri [18]. Numerous hemolytic peptides (Table 1) with various other activities were found to be secreted by different Staphylococci. Thus S. epidermidis was described to produce a complex of peptides, named phenol-soluble modulin (PSM) [13], combining proinflammatory activity and a role in the detachment of biofilm [23,27]. The PSM complex is composed of three peptides of 22, 44 and 25 amino acids named PSM , PSM and PSM , respectively. PSM corresponds to the S. epidermidis -hemolysin [13]. Genes encoding PSM related peptides are present in all sequenced S. aureus strains. The production of the peptides, PSM , PSM and -toxin, was shown to be higher in the community-associated methicillin-resistant S. aureus (CA-MRSA) strains, than in the hospital associated strains (HA-MRSA), and contributes significantly to their virulence [24]. S. haemolyticus secretes three hemolytic peptides, which inhibit the growth of Neisseria gonorrhoeae. They were named gonococcal growth inhibitor (GGI) I, II and III and consist of 44 amino acids linear peptides with high sequence homologies (65–75%) [7,25]. The SLUSH A, B and C peptides, each composed of 43 amino acids with closely related sequences (>76%), are secreted by S. lugdunensis. Their sequences are clearly distinct from known hemolysins but are similar to the S. haemolyticus GGIs [5]. From the three hemolytic peptides secreted from S. cohnii, H1 and H3 are composed of 43 amino acids and resemble to SLUSH peptides, whereas the shorter H2, containing only 18 amino acids,


A. Marchand et al. / Peptides 32 (2011) 845–851


Mehlin et al. [13] Tegmark et al. [18] Wiseman [26] McKevitt et al. [12] Ji et al. [9] Tegmark et al. [18] Mehlin et al. [13] Watson et al. [25]

Donvito et al. [5]

Mak et al. [11]

appears unique [11]. These peptides, whose sequences are presented in Table 1, could be separated in two groups according to their size, from 18 to 26 amino acids for the small peptides, and from 43 to 44 for the long peptides. Nevertheless, like warnericin RK and S. warneri -hemolysins, all these peptides are hydrophobic, induce lysis of erythrocytes but are generally described to be non-active against bacteria, except for GGI peptides active against gonococci. Because warnericin RK presents a high hemolytic activity, which is a major drawback for its therapeutic potency, we decided to test if other Staphylococci could produce anti-Legionella peptides with the final objective to detect a peptide with no or a low hemolytic activity. In this paper we report the selection of Staphylococci producing peptides inhibiting the growth of Legionella. The antibacterial peptides were purified and identified and their activities towards Legionella as well as erythrocytes were measured. Based on the comparison of the activities of the purified peptides, as well as warnericin RK and -hemolysin of S. warneri, two classes of antiLegionella peptides were proposed. 2. Materials and methods 2.1. Bacterial strains, culture media and growth conditions The Staphylococcus strains were grown in Brain Heart Infusion (BHI, Difco) liquid medium at 37 ◦ C for 18–20 h under shaking (250 rpm). L. pneumophila Lens was grown at 37 ◦ C either on buffered charcoal yeast extract (BCYE) agar plates for 96 h or in buffered yeast extract (BYE) liquid medium for 24–30 h under shaking (150 rpm). 2.2. Purification of anti-Legionella peptides In order to characterize anti-Legionella peptides, a two-step purification procedure was conducted as described below. Cells were removed, from an overnight staphylococcal culture, by centrifugation (9000 × g, 15 min, 4 ◦ C) and the supernatant was heated at 70 ◦ C for 15 min. The resulting sample was named crude extract (CE). Chromatographic steps were conducted on a Dionex 3600 HPLC instrument composed of a Dionex P680 HPLC binary pump, an iso-

cratic Dionex UltiMate 3000 pump used as a loading pump and a Dionex UltiMate 3000 UV detector. Firstly, the CE was loaded onto a hydrophobic interaction column (POROS 20 HP2, PerSeptive Biosystems, 4.6 mm × 80 mm). Elution was monitored at 220 nm and 280 nm and carried out, at a flow rate of 2 mL/min, using a water/acetonitrile/trifluoroacetic acid 0.05% (v/v) gradient. After washing for 5 min with 10% acetonitrile, elution was achieved in 25 min with a 20 min linear gradient from 10 to 100% acetonitrile, followed by a 5 min wash with 100% acetonitrile. The collected active fraction was lyophilized. Secondly, the lyophilized fraction was dissolved in 900 L of 50% aqueous acetonitrile and injected onto a Kromasil C8 reverse-phase ˚ HPLC analytical column (5 M, 100 A, 4.6 mm × 250 mm). Separation was carried out using a water/acetonitrile/trifluoroacetic acid 0.05% (v/v) solvent system. After an initial 5 min wash with 50% or 70% (according to the staphylococcal strain) acetonitrile, elution was achieved in 40 min at a flow rate of 0.8 mL/min with a 30 min linear gradient from 50 or 70 to 100% acetonitrile, followed by a 10 min wash with 100% acetonitrile. Each collected fraction was lyophilized and stored at −20 ◦ C for further studies. Synthetic non-formylated peptides, warnericin RK, and H2U, were purchased from Genscript (Piscataway, USA). PSM was provided by Dr. T. Jouenne from the PBS laboratory (CNRS UMR6270, Rouen, France). 2.3. Mass spectrometry For molecular weight determination, the HPLC fractions were analyzed by mass spectrometry, on a Perkin Elmer Sciex API 165 mass spectrometer equipped with an ion-spray source. Each sample was resuspended in 50% acetonitrile/0.1% formic acid and was analyzed by infusion at a flow rate of 5 L/min. The instrument scale for the mass-to-charge (m/z) ratio was calibrated with the ions of the ammonium adduct of polypropylene glycol. Scan data were obtained with LC2-Tune and mass calculation was done with Biomultiview 1.2 (Software package Sciex). 2.4. Peptide titration Peptide concentration was determined by the bicinchoninic acid assay as described by the supplier (Sigma) with bovine serum albumin as a standard. 25 L of each sample were mixed with 200 L of

A. Marchand et al. / Peptides 32 (2011) 845–851


a solution of bicinchoninic acid and copper sulfate 50:1 (v/v). The preparation was incubated at 37 ◦ C for 30 min and the absorbance at 595 nm was measured by a microtiter plate reader OPSYS MR (Thermo Labsystems). 2.5. Anti-Legionella activity assays 2.5.1. Spot on lawn assays Anti-Legionella activity was determined using spot on lawn assays as described below. A 107 colony forming units (CFU)/mL suspension of L. pneumophila was spread onto a BCYE agar plate. Samples to be tested (5 L of staphylococcal culture or 50 L of staphylococcal CE) were spotted onto the surface of the agar plate. The plate was then incubated at 37 ◦ C for 96 h. Anti-Legionella activity was revealed by a zone of inhibition around the test samples. Quantitative analysis was carried out by measuring diameters of the zone of inhibition. 2.5.2. Minimal inhibitory concentration Minimal inhibitory concentration (MIC) of the different peptides was determined using microtiter plate assays as described previously [20]. MIC represents the lowest concentration of peptide which totally inhibits the growth of L. pneumophila. 2.5.3. Membrane permeabilization assays and cultivability evaluation Exponentially growing L. pneumophila (OD600 = 0.4–0.8) were washed and resuspended in BYE at a concentration of 106 CFU/mL. The bacterial suspension was treated with various concentrations, ranging from 0 to 10 M, of peptides during 45 min at 37 ◦ C. The generation time, in these conditions, was evaluated and corresponded to 2.5 h (data not shown) (i) One half of the cell suspensions were then analyzed using a flow cytometric approach after bacterial staining with a couple of fluorochromes (SYTO9 and propidium iodide (PI)), as described previously [21]. Flow cytometric measurements were performed on a FACSCanto II flow cytometer (BD Biosciences, Le Pont de Claix, France) with a 488 nm argon excitation laser. A total of 50,000 events were analyzed in each sample, using BD FACSDiVa 6 software (BD Biosciences) for data acquisition and analysis. Optical filters were set up such that PI fluorescence was measured at 670 LP nm and SYTO 9 fluorescence was measured at 530/30 BP nm. A SYTO 9+/PI-gate was drawn to determine the percentage of non permeabilized cells in the control without peptide. The minimal permeabilization concentration (MPC) of peptides corresponds to the concentration leading to 90% of permeabilization of L. pneumophila. (ii) The other half of the bacteria suspensions were diluted to 1/10, 1/100 or 1/1000 depending on the peptide and then spread on a BCYE agar plate using an automatic spiral plater (Whitley Automatic Spiral Plater, Don Whitley Scientific Ltd). Plates were incubated 96 h at 37 ◦ C before colonies numeration using tables provided by the supplier. The results were expressed by the decrease in the L. pneumophila cultivability estimated at the maximum peptide concentration. 2.6. Hemolytic activity assays Hemolytic activity of the peptides was determined by measuring the released hemoglobin from human erythrocytes as described previously [20]. 100% hemolysis was given by adding 0.1% Triton X 100 to the reaction mixture instead of peptide solutions.

Table 2 Anti-Legionella activity of 15 staphylococcal strains and culture supernatants. Staphylococcal strain Anti-Legionella activitya Culture S. warneri RK S. aureus 2850 S. lugdunensis 967 S. saprophyticus 715 S. hominis 373 S. epidermidis 567 S. xylosus 700404 S. cohnii 898 S. haemolyticus 2259 S. lentus 982 S. equorum 4057C S. simulans 4334 S. chromogenes AM1 S. carnosus 4251 S. caprae 2534D + ++ + +++ +++ + ++ +++ +++ ++ + + − − − Crude extract ++ +++ +++ + ++ +++ + + ++ − − − − − − Hechard et al. [8] Laboratory collection Donvito et al. [5] IMI collection IMI collection IMI collection ATCC IMI collection Laboratory collection IMI collection Laboratory collection Laboratory collection Laboratory collection Laboratory collection Laboratory collection Source/referenceb

a “+++” corresponds to an inhibition zone with a diameter superior to 1.1 cm, “++” to a diameter between 0.8 cm and 1.1 cm and “+” to a diameter between 0.6 cm and 0.8 cm. “−” was affected when no inhibition zone was observed. b IMI: Institut für Molekular Infektionsbiologie, Universität Würzburg, Germany.

3. Results 3.1. Screening Staphylococci for the anti-Legionella activity Staphylococcus strains representative of different species, virulent or not, are listed in Table 2. In a first step, anti-Legionella activity of the selected strains was checked by spotting colonies on an agar medium seeded with L. pneumophila Lens. Inhibitory activity was detected when a clear zone appeared around the colonies after the growth of Legionella. From the 15 selected strains, 12 displayed an inhibition zone (Table 2). In order to verify the relationship of this apparent anti-Legionella activity and the secretion of an antibacterial factor, the cell-free culture supernatants, named crude extracts (CEs), of the strains were assayed against L. pneumophila Lens. Nine from the 15 crude extracts displayed activity halos on the agar test revealing the action of secreted anti-Legionella factors (Table 2). From the 15 tested strains, 12 Staphylococci showed inhibition areas around the colonies but only 9 displayed activity in their culture supernatant. For the non active crude extracts prepared from the 3 strains which showed inhibition halos around colonies, the concentration of the inhibiting factor in the supernatant could be too small to be detected. Indeed, as indicated in Table 2 diameters of the inhibition zones varied from a crude extract to another, showing either that the amount of the antibacterial agents varied depending on the species or that the various species produced different factors that differed in their specific activity. Alternatively, the secreted antimicrobials could be denatured during the incubation at 70 ◦ C of the crude extracts. This step was conducted in order to inhibit proteases. In this case, the antimicrobial agents could be proteins but not peptides. This could be verified by using protease inhibitors instead of heat treatment for example. Alternatively, the antimicrobials could be directly subjected to chromatographic separation without heat treatment in order to obtain antibacterial agents separated from proteases. Almost all the peptides described in the literature (Table 1), secreted by the Staphylococci species selected in our study, were successfully purified by the two steps process developed in this work. However, the SLUSH B from S. lugdunensis, as well as hemolysin I from S. warneri RK were detected by mass spectrometry but in too low amounts to be useful for activity assays. In the same way, GGI III from S. haemolyticus was never detected. Our S. aureus strain did not produce any detectable PSM, as it was demonstrated for hospital associated strains contrarily to the community asso-


A. Marchand et al. / Peptides 32 (2011) 845–851

ciated strains [24]. Finally, no hemolytic peptide was recovered, using our methodology, from the culture supernatant of the S. cohnii strain. This could be due to the strain used, which either do not produce these particular peptides or in too weak amounts. Indeed, the anti-Legionella activity of the crude extract prepared from this strain was low (Table 2). S. epidermidis, S. haemolyticus, S. lugdunensis and S. aureus, known to produce hemolytic peptides, were of particular interest because they appeared as active as S. warneri RK. 3.2. Purification and identification of anti-Legionella peptides Because we postulated that the anti-Legionella factors were proteinaceous, as found for S. warneri RK, the process was first optimized for the purification of warnericin RK and S. warneri hemolysins I and II. The purification method of these peptides previously described and based on the affinity chromatography on hydroxyapatite was time consuming [20]. Thus, taking account of the procedure described for the purification of the -toxin of S. epidermidis [16], we developed a two steps chromatographic strategy. Briefly, crude extracts (CEs) from all the tested Staphylococci were directly applied on a hydrophobic interaction phase column. The adsorbed anti-Legionella peptides were then eluted from the column by using a rapid gradient of acetonitrile. The fractions displaying an anti-Legionella activity were then injected on an analytical C8 HPLC column. The chromatograms obtained from the CE of S. warneri RK (Fig. 1A) were quite different than the one previously described [20]. Particularly, the relative intensities of the peptides peaks were different. Moreover, all the different antiLegionella peptides previously described were found but only in their N-formyl forms. The same protocol was applied to other antiLegionella CE and the observed HPLC profiles varied depending on the strains (Fig. 1). However in all cases, at least one major peak was detected. The corresponding fractions were tested for antiLegionella activity and analyzed by electrospray ionization mass spectrometry (ESI-MS). Molecular masses of the analyzed peptides were compared to those of peptides known to be secreted by the corresponding Staphylococci. All the peptides were thus identified except one, which was named Haemo 3. This peptide, produced by S. haemolyticus 2259, has a molecular mass of 2259.78 Da. Interestingly, all the purified peptides were formylated on their N-terminal methionine. We failed to purify any peptide from the CE of S. cohnii 898. However, the S. cohnii peptide H2U sequence displayed similarities with the warnericin RK one, thus we decided to acquire the non-formylated synthetic peptide. In the same way, in order to evaluate the impact of the formylation on the peptides activities, synthetic warnericin RK and S. epidermidis PSM were tested. 3.3. Hemolytic and anti-Legionella activities of the peptides The purified peptides were assayed for anti-Legionella and hemolytic activities. The antibacterial activity was first determined by measuring the minimal inhibitory concentration (MIC) which corresponds to the capacity of a peptide to inhibit the growth of Legionella (Table 3). Among all the tested peptides, the formylated forms of warnericin RK, -hemolysins I and II from S. warneri and PSM from S. epidermidis showed the highest inhibitory activities towards Legionella (MIC < 1 M). On the other hand, the SLUSH A and C from S. lugdunensis, and GGI II from S. haemolyticus displayed the lowest inhibitory activity (MIC > 5 M). Interestingly, all the latter peptides displayed a longer amino-acids sequence as compared to the first peptides group. Besides, analysis of the growth of L. pneumophila as a function of the concentration of added peptide, showed two opposite types of observed curve shape. In the first case (Fig. 2A), with nonformylated warnericin RK, the peptide seemed to act through an “all or nothing” mechanism with no inhibition of L. pneumophila growth above the MIC. In the second case (Fig. 2B), with non-formylated PSM , the inhibition seemed “progressive” and dependent of the peptide dose. In an attempt to explain this result, we hypothesized that the different shapes would be related to different modes of action. So, in order to verify such hypothesis, the minimal permeabilization concentration (MPC) of peptides, which corresponds to the concentration leading to 90% of permeabilization of the target bacteria, was determined (Table 3). Briefly, the percentage of permeabilized cells, corresponding to those stained with propidium iodide (IP), was measured using flow cytometry. As pointed out for the MIC, the formylated forms of the peptides produced by S.

Fig. 1. Reverse-phase elution profiles at 220-nm of active fractions obtained from hydrophobic interaction chromatography from Staphylococcus crude extracts. (A) S. lugdunensis 967, (B) S. aureus 2850, (C) S. epidermidis 567, (D) S. haemolyticus 2259 and (E) S. warneri RK.

A. Marchand et al. / Peptides 32 (2011) 845–851 Table 3 Activities of peptides towards L. pneumophila and red blood cells. Peptidea MIC curve shape M Group 1 f-Warnericin RK f- -hemolysin II f- -hemolysinb -hemolysin II Warnericin RK f-Ggi I f-SLUSH C f-SLUSH A f-PSM Haemo 3 f- -hemolysinc PSM f-PSM H2U f-Ggi II 0.30 0.54 1.05 1.09 1.22 4.15 5.16 11.28 0.3 < MIC < 12 0.63 1.38 1.59 1.90 2.69 3.04 13.23 0.6 < MIC < 14 AoN AoN AoN AoN AoN Pro Pro Pro AoN AoN Pro Pro Pro Pro Pro MIC MPC M 0.6 0.5 0.3 0.4 1.22 1.37 0.7 2.0 11.0 >8.4 >10.1 >5.4 4.5 >5.2 >3 Bacterial cultivability decrease log 1.0 1.7 1.7 1.3 3.1 0.3 1.2 1.0 >0.9 0.3 0.1 0.1 0.7 0.5 0.6 0.0

Similar Documents

Free Essay

Anti Money

...Anti-Money Laundering Act 2010 2. Definitions.-In this Ordinance, unless there is anything repugnant in the subject or context,- (a) “attachment” means prohibition of transfer, conversion, disposition or movement of property by an order issued under section 8; (c) “CTR” means report on currency transactions exceeding such amount as may be specified by the National Executive Committee; (d) “Court” means the Court specified under section 20 (e) “Director General” means the Director General of FMU appointed under section 6; (f) “financial institutions” includes any institution carrying on any or more of the as listed in section 2(f). (g) “fiscal offence” means an offence punishable under the Income Tax Ordinance, 2001 (XLIX of 2001), the Federal Excise Act, 2005, the Customs Act, 1969 (IV of 1969), the Sales Tax Act, 1990 and any other law as the Federal Government may notify in this behalf; (h) “FMU” means the Financial Monitoring Unit established under section 6; (i) “foreign serious offence” means an offence – (i) against the law of a foreign State stated in a certificate issued by, or on behalf of, the government of that foreign State; and (ii) which, had it occurred in Pakistan, would have constituted a predicate offence; (j) “investigating or prosecuting agency” means the National Accountability Bureau (NAB), Federal Investigation Agency (FIA), Anti-Narcotics Force (ANF) or any other law enforcement agency as may be notified by the Federal Government for the......

Words: 2488 - Pages: 10

Premium Essay

Anti Fed

...equally in the House, regardless of the state in which they lived—unlike the Articles of Confederation, according to which the Continental Congress equally represented the states. In other words, the proposed Constitution would make the United States a nation of one people rather than a loose confederation of states. The proposed Constitution, and the change it wrought in the nature of the American Union, spawned one of the greatest political debates of all time. In addition to the state ratifying conventions, the debates also took the form of a public conversation, mostly through newspaper editorials, with Anti-federalists on one side objecting to the Constitution, and Federalists on the other supporting it. Writers from both sides tried to persuade the public that precious liberty and self-government, hard-earned during the late Revolution, were at stake in the question. Anti-federalists such as the Federal Farmer, Centinel, and Brutus argued that the new Constitution would eventually lead to the dissolution of the state governments, the consolidation of the Union into “one great republic” under an unchecked national government, and as a result the loss of free, self-government. Brutus especially believed that in such an extensive and diverse nation, nothing short of despotism “could bind so great a country under one government.” Federalists such as James Madison (writing as Publius) countered that it was precisely a large nation, in conjunction with a......

Words: 548 - Pages: 3

Premium Essay

The Anti Gym

...The Anti-Gym “Meanwhile fitness never came naturally to me. Growing up I wasn’t an athlete, was relatively inactive and led a sedentary lifestyle.” said John, one of many victims of obesity. At first, he was not sure to share his story but after sharing he narrated as was his experience when he started gaining weight as a teenager: “I was 12 years old when I started putting the weight on and like many children today, I spent long hours in front of screens, TV, video games, movies and the computer, coupled with sugary treats, pop, chips and other snacks. Became my after school past-time. At the age of, 14 I woke up one morning, had a shower like I did every morning, but this morning was different. I normally avoided mirrors whenever I could, but not that day. As I stepped from the shower I caught myself in the mirror. I saw my protruding belly, chubby cheeks and multiple chins, my gaze wandering until eventually ending in a standoff, staring face-to-face with what I had become. As I looks into my own eyes, I started to sob, shedding uncontainable tears. It was that exact moment, my turning point, when I came to a stark realization that something had to change.” He thought “Enough is enough!” and that was in that moment he never looked back and his life changed forever. Nowadays, obesity in adolescents is a serious issue because they eat more and more junk food and this occurs because adolescents have a sedentary lifestyle, lack of adequate physical activity and poor......

Words: 1836 - Pages: 8

Free Essay

Diabetes/Alternative Therapy Activity

...Diabetes/Alternative Therapy Activity 1). Identification and description of the health promotion/alternative therapy activity; including any historical information The alternative therapy activity for diabetes is the acupuncture therapy. Acupuncture is one of the ancient ways of treatment which is around 800 years old. In those years, the Chinese used sharp stones or bones as their surgical instruments. Acupuncture continued to develop since then and has now become one of the leading therapies in the world. The treatment of diabetes by acupuncture therapy is best known for reducing blood glucose levels mostly in the patients with type 2 which is non–insulin diabetes and the most common. The process has identified around 20 body points which are very effective in reducing the blood glucose levels. The main points of acupuncture therapy in the treatment of diabetes conditions are sanyinjiao, zusanli, quchii and the yishu. These points are chosen on the bases of the diabetes history, and the specific stage of the diabetes. The acupuncture points and the treatment process are different from person to person. This diabetes therapy is treated using the combination of acupuncture and specific Chinese herbal medicines which have a hypoglycemic effect. One of the herbs called ginseng plays a major in lowering the blood glucose levels (Oleson, 2012). In a single session, acupuncture may involve the application of dozen points for the treatment. The course for the treatment of......

Words: 1575 - Pages: 7

Free Essay


...| Team building Activities | | 1. Blind Wine Waiter Team Building Task Overview Teams of six must successfully find, uncork and pour a bottle of wine into five glasses. Each team manner must carry out no more than one element of the task and at least five of the team must wear blindfolds. Pre-Work None required Equipment and Layout One bottle of wine per team, one wineglass per team, blindfolds for 5 members of each team, one corkscrew per team. Running The Activity 1. Introduce this as a light-hearted activity that will improve communication across teams. 2. Divide the group into teams of 6 and ask each team to elect a leader. 3. Hand out blindfolds to all team members other than the leader, instruct all team members other than the leader to put on their blindfold. 4. Ask the team leader to take a seat somewhere close to his/her team and ask him/her to sit on her hands. 5. For each team, position one bottle of wine, one wineglass and one corkscrew in various locations around the room. Take care to ensure that nothing is positioned where it might easily fall or break (eg make sure the wine bottle(s) and glass(es) are placed on the floor against a wall, or in the centre of a table). 6. Tell all participants that their task is to find a bottle of wine, a corkscrew and a wine glass, open the bottle and pour their leader a glass of wine. 7. Tell the participants the rules: - the team leader cannot move from his/her position and......

Words: 6849 - Pages: 28

Premium Essay


...will identify the personal benefits he/she gains from participating in leisure activities 2x per week, for 2months. Young Adult In the first session of each week, the group will discuss the personal benefits of every possible leisure activities. Then at the 2nd session will actually experience some activity that related to the discussion in first session. Each 2nd session of the week will have different activities. After the activity the group will going the benefits which will gain from what they are participating again. Elderly Goal: participant will increase their knowledge of available recreational resource and keep the isolation away. Object participant will participate in a variety activities 1x per week. The activities or programs that provided is to meet the ever change needs and interests of the group for example like tai chi,ballroom dancing,art and craft , table games, fitness instruction is offered on the weekly activities calendar 4)All supplies will be ready before the group comes. Clear explanation of each activity will be provide and make sure that every participants in the group are understanding what will be going on before the activity start. One on one assistance if one's needed. 5) group size 6~10participants, each session is holding 1hr 30Mins. Meeting once a week for both target group. 6)both group will need a standard gymnasium, depening on the activities, or else need large open room, chair ,table, papers,color pencils,......

Words: 266 - Pages: 2

Premium Essay


...By: Normandie Lovince I am a supporter of the anti-federalist party. The anti-federalist took some of the ideas that the federalist had into consideration. Instead of abolishing or ignoring these ideas, they wanted to improve them. The anti-federalist and the federalist share two very opposing views. As you read this essay, you will gradually start to see just how my ideas are being supported as to why I've chosen to become an anti-federalist. The anti-federalist party was the first out of two political parties of the U.S. This party was led by Henry, George Mason and Samuel Adams alongside Richard Henry Lee who wanted the president and the senate to have the entire executive and 2/3 of the legislative power. As an anti-federalist, I believe that the constitution should not be ratified. I feel like the best way, that the U.S citizens should be protected is by being kept safe from the Government and the bill of rights will do that because of the freedom and liberty that it gives us. "The greatest importance for Freemen to retain themselves are the liberties given to us in the bill of rights", which is why it's so important that we'd add it to the constitution. In order to get the bill of rights to be in the constitution we'd need to sacrifice part of our natural rights, for the good of others around us. The anti-federalist believed that the constitution should have a bill of rights. The Anti-federalist opposed the constitution, while the federalist themselves......

Words: 836 - Pages: 4

Free Essay

Anti Piracy

...accuracy of any information or advice given in the Booklet or any omission from the Booklet or for any consequence whatsoever resulting directly or indirectly from compliance with, adoption of or reliance on guidance contained in the Booklet even if caused by a failure to exercise reasonable care on the part of any of the aforementioned parties. Printed & bound in Great Britain by Bell & Bain Ltd. Glasgow Published in 2010 by Witherby Seamanship International Ltd, 4 Dunlop Square, Livingston, Edinburgh, EH54 8SB, Scotland, UK Tel No: +44(0)1506 463 227 Email: ii Contents Section 1 Section 2 Section 3 Section 4 Section 5 Section 6 Section 7 Section 8 Section 9 Introduction Somali Pirate Activity Risk Assessment Typical Pirate Attacks Implementing BMP Company Planning Masters’ Planning Prior to Transit – Voyage Planning Prior to Transit – Self Protection Measures 1 3 5 9 11 13 15 17 21 35 39 41 43 45 47 Section 10 If a Pirate Attack is Imminent Section 11 If Boarded by Pirates Section 12 In the Event of Military Action Section 13 Post Incident Reporting Section 14 Updating Best Management Practices ANNEX A ANNEX B Useful Contact Details UKMTO Vessel Position Reporting Form 48 iii ANNEX C ANNEX D ANNEX E Piracy Definitions Follow-up Report Additional Guidance for Vessels Engaged in Fishing, in the Gulf of Aden and off the Coast of Somalia Organisations 49 52 55 59 ANNEX F iv Section......

Words: 9889 - Pages: 40

Premium Essay

Anti Bias

...Early childhood children can identify different color, language, gender, and physical. Children watch the how people are different and yet the same. They understand their native language and recognizes the differences of another language. An anti-bias curriculum help children to embrace as teachers nurture the development of each child’s ability respect issues of diversity and be fear in the classroom. Teachers can create an Anti-bias classroom and the classroom environment should be Multicultural friendly that implement the education which have an active approach to challenge all prejudices, stereotypes or bias. In my opinion an Anti-Bias education is relevant because it reflects on education as a whole that is not based on if people are comfortable with the topics. Educators should have a specific goal for children and give them a clear cut educational experience. It explains how teachers should allow children to talk about what is going on in their world. Even in preschool children are expose to racism and prejudice, by implementing an Anti-bias curriculum children will have an opportunity to defect discrimination. For example, a child told another child that she do not want to play with her because the other child hair wasn’t done. This situation needed to be address right away because you would not want the any child to feel bad about him or herself, because the other children do not want to play with them. Perception by children that they are not good because of......

Words: 1433 - Pages: 6

Premium Essay


...Anti Globalization Michael Albert said that globalization leads to "...a thug's economy, a heartless economy, a base and vile and largely boring economy. It is the antithesis of human fulfillment and development. It mocks equity and justice. It enshrines greed (Globalization Quotes.)" Globalization is a harmful process and it has created more havoc and negative effects than good outcomes. Furthermore, some argue that globalization will help the world deal with crises like poverty and unemployment; this however, is no true. The International Forum on Globalization stated that globalization led to a "sharp increase in unemployment in both the North and the South as corporate activity becomes more mobile, unrestricted, opaque, and unaccountable (Globalization Quotes.)" This process does, in fact, lead to massive unemployment and high poverty levels. The growth in trade has, in general, created more jobs but the equivalent growth in competition has made many companies fire their workers so that they may cut costs, boost efficiency and increase profits. The less developed countries had some negative effects to deal with. For example, China, while experiencing economic growth, has also begun to struggle with unemployment. Studies show that increased trade between the North and South has lead to a decrease in the income equality between semiskilled and skilled workers in the South. On the other hand, the inequality grew among such workers in the North. The manufactured goods......

Words: 544 - Pages: 3

Free Essay


...Community Service Department Activity Guide Summer 2015 REGISTER ONLINE BEGINNING MONDAY, MAY 4 Create Your User Account – See Page 2 Registration (520) 421-8677 • Summer is upon us and the temperatures will really begin to warm up. Within our summer activity guide, you will find several pages of cool opportunities to combat the heat. The newly renovated Palm Island Family Aquatics Center offers open swimming, special splash events, swimming lessons, a swim team, aerobics and Zumba classes. If you would like to use the facility for a private party or corporate event - those rental opportunities are available as well. Enjoy your summer evenings playing in a few outdoor sports leagues such as softball and pickle ball or indoor activities such as basketball and volleyball. The Len Colla Recreation Center and the Dorothy Powell Senior Adult Center both provide daily exercise, arts and educational programming in a very friendly and comfortable setting. Not to be outdone - both the Main and Vista Public Libraries offer a tremendous assortment of youth, teen and adult programming. The Summer Reading Program kicks off on Saturday, May 30th at the Main Library, please circle that date and stop by and enjoy the festivities. Both libraries will also be hosting free technology classes throughout the summer for those 18 years of age and older. Learning is a very cool thing! You will also notice several enhancements being made this summer......

Words: 6060 - Pages: 25

Premium Essay

Infectious Diseases

...can replicate freely until overgrowth or antibiotic administration occurs * Meningitis, inflammation within the sub-arachnoid spaces and associated neurological damage are not a direct result of bacteria, but of the host’s inflammatory response to pathogens and constituents * White pus is evident in the sub-arachnoid space and over the surface of the brain, base of the brain and this can impede flow of the CSF resulting in an increased intra-cranial pressure ------------------------------------------------- CVS INFECTIONS Bacteraemia Pathogenesis * Bacteraemia is when bacteria enter and circulate the blood * It is not always associated with fever and illness * Bacteria are controlled by the bactericidal and anti-inflammatory properties of blood components and vascular endothelium * Poor oral hygiene can lead to transient bacteraemia * Bacteria can enter the blood via a range of ways including: direct inoculation, release from a focus of interaction, by natural flora escaping sites of colonisation * Some organisms may invade through the epithelium – pneumococcal, meningococcal, H. influenzae bacteria Rheumatic Fever Causes: Streptococcus pyogenes of the throat – gram-positive Prevention: Penicillin (beta lactam) 9 days after sore throat Treatment: * Aspirin or NSAIDs Prophylaxis of Recurrent Strep Throat (until individual leaves school): * Phenoxymethylpenicillin (beta lactam) 250mg BD * (OR Cephalosporin (beta......

Words: 5531 - Pages: 23

Premium Essay


...Staphylococcal Staphylococcal food poisoning is the intoxication from the indigestion of contaminated food. Staphylococcal is the name of the condition caused by the enterotoxins that some strains of Staphylococcus Aureus produce (BBB Staphylococcus Aureus, 2012). Foods related with staphylococcus food poisoning include poultry and eggs; meat and meat products; salads such as tuna, chicken, or egg; baked goods such as cream-filled pies; milk and dairy products. Mishandled food during preparation and kept in a considerably high temperature known as the temperature danger zone 41 degrees to 141 degrees are involved in staphylococcal food poisoning. Also staphylococcal can be spread by hand contact, coughing, and sneezing. Food handlers are the number one cause of food poisoning outbreaks unclean food surfaces can also cause staphylococcus aureus. According to the United States Food and Drug Administration 1,364 children became ill who ate lunch that was served to 16 elementary schools in Texas. The food was prepared in an outside kitchen and delivered to the schools. The epidemiological studies revealed that the children who became ill ate chicken salad. The chicken was prepared by the kitchen; after boiling the chicken it was cooled at room temperature using fans. After preparation of the chicken salad it was kept refrigerated overnight between 42 degrees and 45 degrees, placed in thermal containers and delivered to the schools and was kept at room temperature until......

Words: 581 - Pages: 3

Premium Essay


...The IUCN Anti-Fraud Policy February 2008 – Version 1.0 Office of the Director General The World Conservation Union Rue Mauverney 28 1196 Gland, Switzerland Tel: +41 22 999 0296 Fax: +41 22 999 0029 Policy Version Control and Document History: The IUCN Anti-Fraud Policy Title Version Source language Published in French under the title Published in Spanish under the title Responsible Unit Developed by Subject (Taxonomy) Date approved Approved by Applicable to Purpose IUCN Anti – Fraud Policy 1.0 released February 2008 English Politique de l’UICN de lutte contre la fraude Política para la Prevención de Fraudes de la UICN Office of the Director General IUCN Oversight Unit Fraud, Internal Control, Risk Management November 2007 Director General and Global Management Team All IUCN Staff Members world-wide The aim of the IUCN Anti-Fraud Policy is to safeguard the reputation and financial viability of IUCN through improved management of fraud risk. It sets out explicit steps to be taken in response to reported or suspected fraud, as well as measures that will be taken to prevent or minimize the risk of fraud. IUCN Internal Control Policy Framework COSO Standards IUCN Code of Conduct and Professional Ethics for the Secretariat Sent to all staff members world-wide, available on the IUCN Knowledge Network (intranet), provided for information to all partner organizations and suppliers with contracts with IUCN, and available publicly on request. Is part of......

Words: 7003 - Pages: 29

Premium Essay

Anti Dumping

...materially injure or threaten material injury to, the domestic industry, the investigations will continue, and Commerce will be scheduled to make its Countervailing duties and antidumping duties preliminary determinations in March and June 2012, respectively, unless extended. The U.S. government recently decided to no longer collect antidumping duties on imports of stainless steel sheet and strip in coils (SSSSC) from Germany, Italy and Mexico. This action follows from a recent vote of the U.S. International Trade Commission (ITC) in connection with a “sunset review” of existing antidumping duty orders on imports of SSSSC from these three countries, as well as from Japan, Korea and Taiwan. The United States on Friday set hefty preliminary anti-dumping duties on large power transformers made in its future free-trade partner South Korea and used in the electric utility industry. Antidumping duty investigations: Large residential washers are automatic clothes washing machines with a cabinet width (measured from its widest point) of at least 24.5 inches (62.23 cm) and no more than 32 inches (81.28 cm). The merchandise covered by these investigations is all large residential washers and certain subassemblies thereof from Korea and Mexico. For purposes of these investigations, the term “large residential washers” denotes all automatic clothes washing machines, regardless of the orientation of the rotational axis, with a cabinet width (measured from its widest point) of at......

Words: 1430 - Pages: 6