ABSTRACT
Reports of Staphylococcus aureus including methicillin-resistant S. aureus (MRSA) detected in marine environments have occurred since the early 1990 s. This investigation sought to isolate and characterize S. aureus from marine waters and sand at a subtropical recreational beach, with and without bathers present, in order to investigate possible sources and to identify the risks to bathers of exposure to these organisms. During 40 days over 17 months, 1,001 water and 36 intertidal sand samples were collected by either bathers or investigators at a subtropical recreational beach. Methicillin-sensitive S. aureus (MSSA) and MRSA were isolated and identified using selective growth media and an organism-specific molecular marker. Antimicrobial susceptibility, staphylococcal cassette chromosome mec (SCCmec) type, pulsed-field gel electrophoresis (PFGE) pattern, multi-locus sequence type (MLST), and staphylococcal protein A (spa) type were characterized for all MRSA. S. aureus was isolated from 248 (37 %) bather nearby water samples at a concentration range of <2-780 colony forming units per ml, 102 (31 %) ambient water samples at a concentration range of <2-260 colony forming units per ml, and 9 (25 %) sand samples. Within the sand environment, S. aureus was isolated more often from above the intertidal zone than from intermittently wet or inundated sand. A total of 1334 MSSA were isolated from 37 sampling days and 22 MRSA were isolated from ten sampling days. Seventeen of the 22 MRSA were identified by PFGE as the community-associated MRSA USA300. MRSA isolates were all SCCmec type IVa, encompassed five spa types (t008, t064, t622, t688, and t723), two MLST types (ST8 and ST5), and 21 of 22 isolates carried the genes for Panton-Valentine leukocidin. There was a correlation (r = 0.45; p = 0.05) between the daily average number of bathers and S. aureus in the water; however, no association between exposure to S. aureus in these waters and reported illness was found. This report supports the concept that humans are a potential direct source for S. aureus in marine waters.
Subject(s)
Methicillin-Resistant Staphylococcus aureus/isolation & purification , Seawater/microbiology , Staphylococcal Infections/microbiology , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Bacterial Toxins/genetics , Bacterial Toxins/metabolism , Drug Resistance, Bacterial , Exotoxins/genetics , Exotoxins/metabolism , Humans , Leukocidins/genetics , Leukocidins/metabolism , Methicillin-Resistant Staphylococcus aureus/classification , Methicillin-Resistant Staphylococcus aureus/genetics , Public FacilitiesABSTRACT
In May of 2011, a live mass stranding of 26 short-finned pilot whales (Globicephala macrorhynchus) occurred in the lower Florida Keys. Five surviving whales were transferred from the original stranding site to a nearby marine mammal rehabilitation facility where they were constantly attended to by a team of volunteers. Bacteria cultured during the routine clinical care of the whales and necropsy of a deceased whale included methicillin-sensitive and methicillin-resistant Staphylococcus aureus (MSSA and MRSA). In order to investigate potential sources or reservoirs of MSSA and MRSA, samples were obtained from human volunteers, whales, seawater, and sand from multiple sites at the facility, nearby recreational beaches, and a canal. Samples were collected on 3 days. The second collection day was 2 weeks after the first, and the third collection day was 2 months after the last animal was removed from the facility. MRSA and MSSA were isolated on each day from the facility when animals and volunteers were present. MSSA was found at an adjacent beach on all three collection days. Isolates were characterized by utilizing a combination of quantitative real-time PCR to determine the presence of mecA and genes associated with virulence, staphylococcal protein A typing, staphylococcal cassette chromosome mec typing, multilocus sequence typing, and pulsed field gel electrophoresis (PFGE). Using these methods, clonally related MRSA were isolated from multiple environmental locations as well as from humans and animals. Non-identical but genetically similar MSSA and MRSA were also identified from distinct sources within this sample pool. PFGE indicated that the majority of MRSA isolates were clonally related to the prototype human strain USA300. These studies support the notion that S. aureus may be shed into an environment by humans or pilot whales and subsequently colonize or infect exposed new hosts.
Subject(s)
Cetacea/microbiology , Fin Whale/microbiology , Methicillin-Resistant Staphylococcus aureus/isolation & purification , Staphylococcal Infections/microbiology , Staphylococcal Infections/veterinary , Animals , Anti-Bacterial Agents/pharmacology , Florida , Humans , Methicillin-Resistant Staphylococcus aureus/classification , Methicillin-Resistant Staphylococcus aureus/drug effects , Methicillin-Resistant Staphylococcus aureus/genetics , VolunteersABSTRACT
Superinfections from Staphylococcus aureus following influenza are an increasing concern. We assessed several laboratory and clinical strains in a mouse coinfection model with influenza virus. A methicillin-resistant USA300 clone and several recent clinical strains from patients with necrotizing pneumonia caused high mortality following influenza virus infection in mice. Both viral and bacterial lung titers were enhanced during coinfections compared with single infections. However, differences in titers did not correspond with differences in disease outcomes in a comparison of superinfections from a highly pathogenic strain with those from a poorly pathogenic strain. These strains did differ, however, in expression of Panton-Valentine leukocidin and in the degree of inflammatory lung damage each engendered. The viral cytotoxin PB1-F2 contributed to the negative outcomes. These data suggest that additional study of specific bacterial virulence factors involved in the pathogenesis of inflammation and lung damage during coinfections is needed.
Subject(s)
Influenza A virus/pathogenicity , Influenza, Human/complications , Pneumonia, Staphylococcal/microbiology , Staphylococcus aureus/pathogenicity , Superinfection/microbiology , Animals , Cytotoxins , Disease Models, Animal , Female , Humans , Influenza A virus/immunology , Liver/pathology , Liver/virology , Lung/pathology , Lung/virology , Methicillin-Resistant Staphylococcus aureus , Mice , Mice, Inbred BALB C , Pneumonia, Staphylococcal/complications , Polymerase Chain Reaction , Spleen/pathology , Spleen/virology , Superinfection/complications , Survival AnalysisABSTRACT
BACKGROUND: Staphylococcus aureus including methicillin resistant S. aureus, MRSA, are human colonizing bacteria that commonly cause opportunistic infections primarily involving the skin in otherwise healthy individuals. These infections have been linked to close contact and sharing of common facilities such as locker rooms, schools and prisons Waterborne exposure and transmission routes have not been traditionally associated with S. aureus infections. Coastal marine waters and beaches used for recreation are potential locations for the combination of high numbers of people with close contact and therefore could contribute to the exposure to and infection by these organisms. The primary aim of this study was to evaluate the amount and characteristics of the shedding of methicillin sensitive S. aureus, MSSA and MRSA by human bathers in marine waters. RESULTS: Nasal cultures were collected from bathers, and water samples were collected from two sets of pools designed to isolate and quantify MSSA and MRSA shed by adults and toddlers during exposure to marine water. A combination of selective growth media and biochemical and polymerase chain reaction analysis was used to identify and perform limited characterization of the S. aureus isolated from the water and the participants. Twelve of 15 MRSA isolates collected from the water had identical genetic characteristics as the organisms isolated from the participants exposed to that water while the remaining 3 MRSA were without matching nasal isolates from participants. The amount of S. aureus shed per person corresponded to 105 to 106 CFU per person per 15-minute bathing period, with 15 to 20% of this quantity testing positive for MRSA. CONCLUSIONS: This is the first report of a comparison of human colonizing organisms with bacteria from human exposed marine water attempting to confirm that participants shed their own colonizing MSSA and MRSA into their bathing milieu. These findings clearly demonstrate that adults and toddlers shed their colonizing organisms into marine waters and therefore can be sources of potentially pathogenic S. aureus and MRSA in recreational marine waters. Additional research is needed to evaluate recreational beaches and marine waters as potential exposure and transmission pathways for MRSA.
Subject(s)
Methicillin-Resistant Staphylococcus aureus/isolation & purification , Seawater/microbiology , Staphylococcus aureus/isolation & purification , Water Microbiology , Adult , Child, Preschool , Female , Humans , Infant , Male , Nose/microbiologyABSTRACT
Morbidity and mortality estimates due to methicillin-resistant Staphylococcus aureus (MRSA) infections continue to rise. Therapeutic options are limited by antibiotic resistance. Anti-pathogenic compounds, which inhibit quorum sensing (QS) pathways, may be a useful alternative to antibiotics. Staphylococcal QS is encoded by the AGR locus and is responsible for the production of δ-hemolysin. Quantification of δ-hemolysin found in culture supernatants permits the analysis of AGR activity at the translational rather than transcriptional level. We employed reversed phase high performance chromatographic (RP-HPLC) techniques to investigate the anti-QS activity of 168 extracts from 104 Italian plants through quantification of δ-hemolysin. Extracts from three medicinal plants (Ballota nigra, Castanea sativa, and Sambucus ebulus) exhibited a dose-dependent response in the production of δ-hemolysin, indicating anti-QS activity in a pathogenic MRSA isolate.
Subject(s)
Bacterial Proteins/drug effects , Hemolysin Proteins/drug effects , Methicillin-Resistant Staphylococcus aureus/drug effects , Plant Extracts/pharmacology , Plants, Medicinal/chemistry , Quorum Sensing/drug effects , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/antagonists & inhibitors , Bacterial Proteins/metabolism , Dose-Response Relationship, Drug , Fagaceae/chemistry , Hemolysin Proteins/antagonists & inhibitors , Hemolysin Proteins/metabolism , Italy , Lamiaceae/chemistry , Methicillin-Resistant Staphylococcus aureus/physiology , Microbial Sensitivity Tests , Sambucus/chemistryABSTRACT
Studies evaluating the relationship between microbes and human health at non-point source beaches are necessary for establishing criteria which would protect public health while minimizing economic burdens. The objective of this study was to evaluate water quality and daily cumulative health effects (gastrointestinal, skin, and respiratory illnesses) for bathers at a non-point source subtropical marine recreational beach in order to better understand the inter-relationships between these factors and hence improve monitoring and pollution prevention techniques. Daily composite samples were collected, during the Oceans and Human Health Beach Exposure Assessment and Characterization Health Epidemiologic Study conducted in Miami (Florida, USA) at a non-point source beach, and analyzed for several pathogens, microbial source tracking markers, indicator microbes, and environmental parameters. Analysis demonstrated that rainfall and tide were more influential, when compared to other environmental factors and source tracking markers, in determining the presence of both indicator microbes and pathogens. Antecedent rainfall and F+ coliphage detection in water should be further assessed to confirm their possible association with skin and gastrointestinal (GI) illness outcomes, respectively. The results of this research illustrate the potential complexity of beach systems characterized by non-point sources, and how more novel and comprehensive approaches are needed to assess beach water quality for the purpose of protecting bather health.
Subject(s)
Bathing Beaches , Gastrointestinal Diseases/microbiology , Respiratory Tract Infections/microbiology , Seawater/microbiology , Water Microbiology , Coliphages/isolation & purification , Enterococcus/isolation & purification , Enterovirus/isolation & purification , Environmental Exposure/adverse effects , Environmental Monitoring/methods , Epidemiological Monitoring , Florida/epidemiology , Gastrointestinal Diseases/epidemiology , Humans , Rain , Respiratory Tract Infections/epidemiology , Respiratory Tract Infections/transmissionABSTRACT
Swimming in ocean water, including ocean water at beaches not impacted by known point sources of pollution, is an increasing health concern. This study was an initial evaluation of the presence of indicator microbes and pathogens and the association among the indicator microbes, pathogens, and environmental conditions at a subtropical, recreational marine beach in south Florida impacted by non-point sources of pollution. Twelve water and eight sand samples were collected during four sampling events at high or low tide under elevated or reduced solar insolation conditions. The analyses performed included analyses of fecal indicator bacteria (FIB) (fecal coliforms, Escherichia coli, enterococci, and Clostridium perfringens), human-associated microbial source tracking (MST) markers (human polyomaviruses [HPyVs] and Enterococcus faecium esp gene), and pathogens (Vibrio vulnificus, Staphylococcus aureus, enterovirus, norovirus, hepatitis A virus, Cryptosporidium spp., and Giardia spp.). The enterococcus concentrations in water and sand determined by quantitative PCR were greater than the concentrations determined by membrane filtration measurement. The FIB concentrations in water were below the recreational water quality standards for three of the four sampling events, when pathogens and MST markers were also generally undetectable. The FIB levels exceeded regulatory guidelines during one event, and this was accompanied by detection of HPyVs and pathogens, including detection of the autochthonous bacterium V. vulnificus in sand and water, detection of the allochthonous protozoans Giardia spp. in water, and detection of Cryptosporidium spp. in sand samples. The elevated microbial levels were detected at high tide and under low-solar-insolation conditions. Additional sampling should be conducted to further explore the relationships between tidal and solar insolation conditions and between indicator microbes and pathogens in subtropical recreational marine waters impacted by non-point source pollution.
Subject(s)
Bacteria/isolation & purification , Bathing Beaches , Parasites/isolation & purification , Seawater/microbiology , Viruses/isolation & purification , Water Microbiology , Animals , Bathing Beaches/standards , Clostridium perfringens/isolation & purification , Cryptosporidium/isolation & purification , Enterococcus/isolation & purification , Enterococcus faecium/isolation & purification , Environmental Monitoring , Environmental Pollutants/isolation & purification , Escherichia coli/isolation & purification , Florida , Fresh Water/microbiology , Humans , Polyomavirus/isolation & purification , Recreation , Seawater/parasitology , Seawater/virology , Silicon Dioxide , Viruses/genetics , Water SupplyABSTRACT
The objectives of this work were to compare enterococci (ENT) measurements based on the membrane filter, ENT(MF) with alternatives that can provide faster results including alternative enterococci methods (e.g., chromogenic substrate (CS), and quantitative polymerase chain reaction (qPCR)), and results from regression models based upon environmental parameters that can be measured in real-time. ENT(MF) were also compared to source tracking markers (Staphylococcus aureus, Bacteroidales human and dog markers, and Catellicoccus gull marker) in an effort to interpret the variability of the signal. Results showed that concentrations of enterococci based upon MF (<2 to 3320 CFU/100 mL) were significantly different from the CS and qPCR methods (p < 0.01). The correlations between MF and CS (r = 0.58, p < 0.01) were stronger than between MF and qPCR (r ≤ 0.36, p < 0.01). Enterococci levels by MF, CS, and qPCR methods were positively correlated with turbidity and tidal height. Enterococci by MF and CS were also inversely correlated with solar radiation but enterococci by qPCR was not. The regression model based on environmental variables provided fair qualitative predictions of enterococci by MF in real-time, for daily geometric mean levels, but not for individual samples. Overall, ENT(MF) was not significantly correlated with source tracking markers with the exception of samples collected during one storm event. The inability of the regression model to predict ENT(MF) levels for individual samples is likely due to the different sources of ENT impacting the beach at any given time, making it particularly difficult to to predict short-term variability of ENT(MF) for environmental parameters.
Subject(s)
Bathing Beaches , Environmental Monitoring/methods , Sewage/analysis , Water Pollutants/analysis , Enterococcus/isolation & purification , Seawater/chemistry , Seawater/microbiology , Staphylococcus aureus/isolation & purification , Water Pollution/statistics & numerical dataABSTRACT
AIM OF STUDY: One-third of botanical remedies from southern Italy are used to treat skin and soft tissue infection (SSTI). Staphylococcus aureus, a common cause of SSTI, has generated increasing concern due to drug resistance. Many plants possess antimicrobial agents and provide effective remedies for SSTI. Our aim was to investigate plants from different ethnobotanical usage groups for inhibition of growth and biofilms in methicillin-resistant Staphylococcus aureus (MRSA). MATERIALS AND METHODS: Three groups were assessed: plant remedies for SSTI, plant remedies not involving the skin, and plants with no ethnomedical application. We screened 168 extracts, representing 104 botanical species, for activity against MRSA (ATCC 33593). We employed broth dilution methods to determine the MIC after 18 h growth using an optical density (OD 600 nm) reading. Anti-biofilm effects were assessed by growing biofilms for 40 h, then fixing and staining with crystal violet. After washing, 10% Tween 80 was added and OD 570 nm readings were taken. RESULTS: Extracts from 10 plants exhibited an IC50Subject(s)
Bacterial Adhesion/drug effects
, Biofilms/drug effects
, Methicillin-Resistant Staphylococcus aureus/drug effects
, Plankton/drug effects
, Plant Extracts/pharmacology
, Plants, Medicinal
, Biofilms/growth & development
, Italy
, Methicillin-Resistant Staphylococcus aureus/growth & development
, Plankton/growth & development
ABSTRACT
OBJECTIVES: To evaluate Staphylococcus aureus isolates from infected skin lesions for their potential to produce immune system-modulating toxins and to correlate these with white blood cell (WBC) counts associated with these lesions. DESIGN: Specimens were obtained for bacterial culture and gram staining from 105 infected skin lesions, and the number of WBCs per low-power field (LPF) was determined. Chromosomal DNA was prepared from 84 bacterial isolates and subjected to real-time polymerase chain reaction analysis to determine the presence of genes encoding potential immunomodulating toxins. Bacterial populations were divided into 2 groups: those associated with low WBC counts (0-5 WBCs/LPF) and those with high WBC counts (> 5 WBCs/LPF). We applied chi(2) statistical analyses to compare the toxin gene profiles associated with WBC counts on initial swab for culture. PATIENTS: Samples were obtained from patients at a single geographic location. RESULTS: A higher than expected percentage of bacteria capable of producing the exfoliative toxins A and/or B (ETA and/or ETB) and Panton-Valentine leukocidin (PVL) was seen in all skin lesions infected with S aureus without regard to WBC count with initial cultures. Comparison of the toxins associated with the low WBC group vs the high WBC group showed that low WBC counts were associated with ETA and ETB, while high WBC counts were associated with PVL and toxic shock syndrome toxin. There were no differences in the clinical appearance of the lesions between groups. CONCLUSIONS: Staphylococcus aureus virulence factors ETA, ETB, and PVL are associated with WBC counts from infected skin lesions. The exact role they play in affecting the WBC counts remains to be determined.
Subject(s)
Bacterial Toxins/biosynthesis , Staphylococcal Skin Infections/immunology , Staphylococcus aureus/metabolism , Virulence Factors/biosynthesis , Antibody Formation , Enterotoxins/biosynthesis , Exfoliatins/biosynthesis , Exotoxins/biosynthesis , Humans , Leukocidins/biosynthesis , Leukocyte Count , Protein Isoforms/biosynthesis , Staphylococcal Skin Infections/blood , Staphylococcal Skin Infections/pathology , Superantigens/biosynthesisABSTRACT
Fetomaternal hemorrhage is a rare, potentially catastrophic event for a fetus. Leakage of the fetus's blood into the mother's circulation can cause fetal anemia, hydrops, and even death. The prevailing symptom is decreased fetal movement, and signs can include a sinusoidal electronic fetal monitor pattern, a positive Kleihauer-Betke test, or changes in fetal Doppler blood flow. A mother's report or perception of decreased fetal movement coupled with a nonreactive nonstress test or abnormal ultrasound findings should prompt an investigation into underlying causes.
Subject(s)
Blood Transfusion/methods , Fetal Movement , Fetomaternal Transfusion/therapy , Pregnancy Outcome , Ultrasonography, Prenatal/methods , Adult , Female , Fetal Monitoring/methods , Fetomaternal Transfusion/diagnostic imaging , Follow-Up Studies , Humans , Postnatal Care/methods , Pregnancy , Pregnancy Trimester, Third , Risk Assessment , Treatment OutcomeABSTRACT
Many skin infections are caused by Staphylococcus aureus, a bacterial pathogen that produces virulence factors associated with these conditions such as exfoliative toxins A and B (ETA, ETB) and the leukotoxin Panton-Valentine leukocidin (PVL). Herein, we examine the potential of skin-infecting S. aureus to produce virulence factors and their impact on the local immune response. Toxin gene profiles were generated from 188 S. aureus isolated as single infecting organisms from skin lesions and demonstrated a higher potential to express ETA, ETB, and PVL than community isolates (p < 0.001). Within the study isolate group, the prevalence of genes encoding PVL was higher among methicillin-resistant S. aureus (MRSA; n = 49), while genes encoding ETs were more prevalent in methicillin-susceptible S. aureus (MSSA; n = 139). When lesion-associated white blood cell (WBC) counts were dichotomized into high- or low-WBC-count-associated bacteria, the gene for ETA was found to be associated with a low WBC count among MSSA (p = 0.001). The ETA-induced mouse model of staphylococcal scalded skin syndrome was used to investigate the link between ETA and cytokine production. Elevated IL-6 levels in the serum and increased expression of IL-6 mRNA in the skin were detected in response to ETA exposure. These findings were recapitulated in vitro using primary human keratinocytes. Thus, S. aureus may influence the local immune response via ETA cleavage of desmoglein 1 and the induction of cutaneous IL-6 expression.
Subject(s)
Desmoglein 1/metabolism , Exfoliatins/metabolism , Interleukin-6/biosynthesis , Keratinocytes/metabolism , Staphylococcal Skin Infections/metabolism , Staphylococcal Skin Infections/microbiology , Staphylococcus aureus/metabolism , Animals , Cell Line , Epidermis/metabolism , Epidermis/microbiology , Humans , Interleukin-6/blood , Keratinocytes/microbiology , Leukocytes/immunology , Leukocytes/pathology , Methicillin-Resistant Staphylococcus aureus/genetics , Methicillin-Resistant Staphylococcus aureus/metabolism , Proteolysis , Staphylococcal Skin Infections/pathology , Staphylococcus aureus/genetics , Virulence Factors/genetics , Virulence Factors/metabolismABSTRACT
Understanding the pathology resulting from Staphylococcus aureus and Pseudomonas aeruginosa polymicrobial wound infections is of great importance due to their ubiquitous nature, increasing prevalence, growing resistance to antimicrobial agents, and ability to delay healing. Methicillin-resistant S. aureus USA300 is the leading cause of community-associated bacterial infections resulting in increased morbidity and mortality. We utilized a well-established porcine partial thickness wound healing model to study the synergistic effects of USA300 and P. aeruginosa on wound healing. Wound re-epithelialization was significantly delayed by mixed-species biofilms through suppression of keratinocyte growth factor 1. Pseudomonas showed an inhibitory effect on USA300 growth in vitro while both species co-existed in cutaneous wounds in vivo. Polymicrobial wound infection in the presence of P. aeruginosa resulted in induced expression of USA300 virulence factors Panton-Valentine leukocidin and α-hemolysin. These results provide evidence for the interaction of bacterial species within mixed-species biofilms in vivo and for the first time, the contribution of virulence factors to the severity of polymicrobial wound infections.
Subject(s)
Methicillin-Resistant Staphylococcus aureus/pathogenicity , Pseudomonas aeruginosa/pathogenicity , Wound Infection/microbiology , Animals , Biofilms/growth & development , Female , Immunohistochemistry , Real-Time Polymerase Chain Reaction , Staphylococcal Infections/metabolism , Swine , Virulence Factors/genetics , Virulence Factors/metabolism , Wound Infection/metabolismABSTRACT
Fecal indicator microbes, such as enterococci, are often used to assess potential health risks caused by pathogens at recreational beaches. Microbe levels often vary based on collection time and sampling location. The primary goal of this study was to assess how spatial and temporal variations in sample collection, which are driven by environmental parameters, impact enterococci measurements and beach management decisions. A secondary goal was to assess whether enterococci levels can be predictive of the presence of Staphylococcus aureus, a skin pathogen. Over a ten-day period, hydrometeorologic data, hydrodynamic data, bather densities, enterococci levels, and S. aureus levels including methicillin-resistant S. aureus (MRSA) were measured in both water and sand. Samples were collected hourly for both water and sediment at knee-depth, and every 6 h for water at waist-depth, supratidal sand, intertidal sand, and waterline sand. Results showed that solar radiation, tides, and rainfall events were major environmental factors that impacted enterococci levels. S. aureus levels were associated with bathing load, but did not correlate with enterococci levels or any other measured parameters. The results imply that frequencies of advisories depend heavily upon sample collection policies due to spatial and temporal variation of enterococci levels in response to environmental parameters. Thus, sampling at different times of the day and at different depths can significantly impact beach management decisions. Additionally, the lack of correlation between S. aureus and enterococci suggests that use of fecal indicators may not accurately assess risk for some pathogens.
Subject(s)
Bathing Beaches/standards , Enterococcus , Geologic Sediments/microbiology , Public Health Practice/standards , Seawater/microbiology , Staphylococcus aureus/isolation & purification , Population Density , Rain , Species Specificity , Time Factors , Water MovementsABSTRACT
Alopecia areata (AA) is a common autoimmune disease characterized by non-scarring hair loss. Previous studies have demonstrated an association between AA and physiological/psychological stress. In this study, we investigated the effects of heat treatment, a physiological stress, on AA development in C3H/HeJ mice. Whereas this strain of mice are predisposed to AA at low incidence by 18 months of age, we observed a significant increase in the incidence of hair loss in heat-treated 8-month-old C3H/HeJ mice compared with sham-treated mice. Histological analysis detected mononuclear cell infiltration in anagen hair follicles, a characteristic of AA, in heat-treated mouse skin. As expected, increased expression of induced HSPA1A/B (formerly called HSP70i) was detected in skin samples from heat-treated mice. Importantly, increased HSPA1A/B expression was also detected in skin samples from C3H/HeJ mice that developed AA spontaneously. Our results suggest that induction of HSPA1A/B may precipitate the development of AA in C3H/HeJ mice. For future studies, the C3H/HeJ mice with heat treatment may prove a useful model to investigate stress response in AA.
Subject(s)
Alopecia Areata/epidemiology , Hot Temperature , Alopecia Areata/pathology , Animals , Disease Models, Animal , HSP70 Heat-Shock Proteins/metabolism , Incidence , Mice , Mice, Inbred C3H , Stress, PhysiologicalABSTRACT
The use of enterococci as the primary fecal indicator bacteria (FIB) for the determination of recreational water safety has been questioned, particularly in sub/tropical marine waters without known point sources of sewage. Alternative FIB (such as the Bacteroidales group) and alternative measurement methods (such as rapid molecular testing) have been proposed to supplement or replace current marine water quality testing methods which require culturing enterococci. Moreover, environmental parameters have also been proposed to supplement current monitoring programs. The objective of this study was to evaluate the health risks to humans from exposure to subtropical recreational marine waters with no known point source. The study reported symptoms between one set of human subjects randomly assigned to marine water exposure with intensive environmental monitoring compared with other subjects who did not have exposure. In addition, illness outcomes among the exposed bathers were compared to levels of traditional and alternative FIB (as measured by culture-based and molecular-based methods), and compared to easily measured environmental parameters. Results demonstrated an increase in self-reported gastrointestinal, respiratory and skin illnesses among bathers vs. non-bathers. Among the bathers, a dose-response relationship by logistic regression modeling was observed for skin illness, where illness was positively related to enterococci enumeration by membrane filtration (odds ratio = 1.46 [95% confidence interval = 0.97-2.21] per increasing log10 unit of enterococci exposure) and positively related to 24 h antecedent rain fall (1.04 [1.01-1.07] per increasing millimeters of rain). Acute febrile respiratory illness was inversely related to water temperature (0.74 [0.56-0.98] per increasing degree of water temperature). There were no significant dose-response relationships between report of human illness and any of the other FIB or environmental measures. Therefore, for non-point source subtropical recreational marine waters, this study suggests that humans may be at increased risk of reported illness, and that the currently recommended and investigational FIB may not track gastrointestinal illness under these conditions; the relationship between other human illness and environmental measures is less clear.
Subject(s)
Bathing Beaches , Enterococcus/isolation & purification , Feces/microbiology , Recreation , Seawater/microbiology , Tropical Climate , Water Microbiology , Adult , Humans , Logistic Models , Multivariate Analysis , Respiratory Tract Diseases/microbiology , Skin/microbiology , Skin/pathologySubject(s)
Exfoliatins/metabolism , Staphylococcal Skin Infections/physiopathology , Staphylococcus aureus/metabolism , Exfoliatins/genetics , Humans , Staphylococcal Skin Infections/diagnosis , Staphylococcal Skin Infections/microbiology , Staphylococcus aureus/isolation & purification , Staphylococcus aureus/pathogenicity , VirulenceABSTRACT
Traditionally, the use of enterococci has been recommended as the fecal indicator bacteria of choice for testing marine recreational water quality, and prior studies have shown that bathers shed large numbers of enterococci into the water. The current study expands upon prior research by evaluating shedding from both toddlers and adults, and by the expansion of measurements to include enterococci shedding via three different methods (membrane filter (MF), chromogenic substrate (CS), and quantitative polymerase chain reaction (qPCR)) and shedding of alternative fecal indicator bacteria (Bacteroidales human markers UCD and HF8 via qPCR). Two sets of experiments were conducted. The first experiment consisted of two groups of 10 adults who bathed together in a large pool. The second study consisted of 14 toddlers who bathed individually in a small pool which allowed for sand recovery. Sand recovery was used to estimate the amount of sand transported on the bodies of toddlers and to estimate the number of fecal indicator bacteria released from this sand. The numbers of estimated enterococci shed per adult ranged from 1.8 x 10(4) to 2.8 x 10(6) CFU, from 1.9 x 10(3) to 4.5 x 10(6) MPN, and from 3.8 x 10(5) to 5.5 x 10(6) GEU based on the MF, CS, and qPCR methods, respectively. The estimated numbers of Bacteroidales human markers ranged from 1.8 x 10(4) to 1.3 x 10(6) for UCD, and ranged from the below detection limit to 1.6 x 10(5) for HF8. The estimated amount of sand transported per toddler (n=14) into the water column after sand exposure was 8+/-6g on average. When normalizing the numbers of enterococci shed from toddlers via sand by the 3.9 body surface area ratio, the differences between toddlers and adults were insignificant. Contributions of sands to the total enterococci (MF) shed per toddler was 3.7+/-4.4% on average. Although shedding via beach sand may contribute a small fraction of the microbial load during initial bathing, it may have a significant role if bathers go to water repetitively after sand exposure.
Subject(s)
Bacteroidetes/isolation & purification , Baths , Enterococcus/isolation & purification , Water Microbiology , Adult , Bacteroidetes/genetics , Bathing Beaches , DNA, Bacterial/genetics , Enterococcus/genetics , Fresh Water/microbiology , Humans , Infant , Polymerase Chain Reaction , Seawater/microbiology , Silicon Dioxide , Water Purification/instrumentation , Water Purification/methodsABSTRACT
Numerous bacterial pathogens use type III secretion systems (T3SSs) or T4SSs to inject or translocate virulence proteins into eukaryotic cells. Several different reporter systems have been developed to measure the translocation of these proteins. In this study, a peptide tag-based reporter system was developed and used to monitor the injection of T3S and T4S substrates. The glycogen synthase kinase (GSK) tag is a 13-residue phosphorylatable peptide tag derived from the human GSK-3beta kinase. Translocation of a GSK-tagged protein into a eukaryotic cell results in host cell protein kinase-dependent phosphorylation of the tag, which can be detected with phosphospecific GSK-3beta antibodies. A series of expression plasmids encoding Yop-GSK fusion proteins were constructed to evaluate the ability of the GSK tag to measure the injection of Yops by the Yersinia pestis T3SS. GSK-tagged YopE, YopH, LcrQ, YopK, YopN, and YopJ were efficiently phosphorylated when translocated into HeLa cells. Similarly, the injection of GSK-CagA by the Helicobacter pylori T4SS into different cell types was measured via phosphorylation of the GSK tag. The GSK tag provides a simple method to monitor the translocation of T3S and T4S substrates.
Subject(s)
Bacterial Outer Membrane Proteins/metabolism , Bacterial Proteins/metabolism , Glycogen Synthase Kinases/analysis , Glycogen Synthase Kinases/metabolism , Virulence Factors/analysis , Virulence Factors/metabolism , Amino Acid Sequence , Bacterial Outer Membrane Proteins/genetics , Bacterial Proteins/genetics , Genetic Vectors/genetics , Glycogen Synthase Kinases/genetics , HeLa Cells , Helicobacter pylori/metabolism , Humans , Molecular Sequence Data , Phosphorylation , Plasmids/genetics , Protein Transport , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolism , Virulence Factors/genetics , Yersinia pestis/metabolismABSTRACT
UNLABELLED: A previously well, spontaneously breathing premature infant (gestational age 25 weeks, birth weight 364 g, age 74 days) developed staphylococcal scalded skin syndrome (SSSS). A methicillin-sensitive strain of Staphylococcus aureus producing exfoliative toxins A and B (ETA, ETB) was isolated from a gastric aspirate and a pharyngeal swab. The disease recurred with a milder clinical picture 4 weeks later in the same patient while under steroid treatment. Cultures obtained from conjunctiva and pharynx were again positive for S. aureus. A second premature infant in an adjacent ward developed SSSS 2 weeks after the recurrence in the first patient. No other cases were observed thereafter. A total of 25 individuals who had contact with the first patient were screened for staphylococcal colonisation. S.aureus was isolated from the posterior part of the nasal cavity in 8 of the 25 contacts. These strains and the strain of the first patient were evaluated by PCR for the presence of genes encoding ETA and ETB. Expression of toxins was confirmed by gel electrophoresis and Western blot analysis. Purified toxins were injected into newborn mice to confirm toxin activity. Besides the strain isolated from the first patient, only one isolate from the medical staff was positive for the genes encoding ETA and ETB. CONCLUSION: the carrier of this strain had contact with both patients, suggesting that this individual was the vector between the two patients but not necessarily the source of the original infection. Strict infection control measures were implemented and no further spread of the disease occurred.