Transcript Document

Abstract #
GRC 2010
Exploratory Study of Bacteria, Protozoa, Yeast, Helminthes, and
Viruses of Health Significance at a Recreational Beach
Ashish H. Shah
[email protected]
University of Miami
Environmental Engineering
Ashish H. Shah1,2, Amir M. Abdelzaher1,2, Helena M. Solo-Gabriele1,2, Matthew Phillips1,2 , Rafael Hernandez1,2, Jonathan Kish1,9 , Gloria Scorzetti1,3, Jack W. Fell1,3,
Mara R. Diaz1,3, Troy M. Scott1,4 , Jerzy Lukasik 5, Valerie J. Harwood6, Shannon McQuaig6, Chris D. Sinigalliano1,7, Maribeth L. Gidley 1,7 , David Wanless1,7,
Arba Agar1,8, Jenbon Lui1,8, Jill R. Stewart9, Lisa R.W. Plano1,10 Lora E. Fleming1,3,11
1University of
Miami, NSF NIEHS OHHCenter , 2Department of Civil, Arch., and Env. Eng.-University of Miami , 3Rosentiel School of Marine and Atmospheric Science-University of Miami, 4BCS Laboratories Inc.- Miami, 5BCS
Laboratories Inc.- Gainesville, 6Dept. of Biology,-University of South Florida, 7National Oceanic and Atmospheric Administration –Miami, 8Dept.of Microbiology and Immunology- University of Miami, 9University of North
Carolina- Chapel Hill, 10Dept .of Microbiology and Immunology and Dept. of Pediatrics- University of Miami,, 11Dept. of Epidemiology & Public Health and Marine Biology & Fisheries -University of Miami
ABSTRACT
An evaluation of the relationship between indicator microbes, pathogens, and the
environment is rare and necessary for establishing a criterion that would allow
for a better understanding of sand pathogens at recreational beach sites, while
concurrently protecting public health. The objective of this study was to
determine the presence, concentrations, and distributions of pathogens and
indicator microbes in sand and their association with environmental parameters.
In this exploratory Sand Pathogens study in Miami (Fl, USA), beach sand samples
were collected and analyzed along three sampling transects (Dry, Wet, and
Inundated) over the course of six days. Each sample was analyzed for several
pathogens, microbial source tracking markers, indicator microbes and
environmental parameters. Preliminary analysis demonstrated an inverse
correlation between moisture content and indicator microbes with no significant
correlation between pathogens and environmental parameters (except for
methicillin resistant S. aureus, MRSA, and C. perfringens). Significant associations
were identified between some indicator microbes and pathogens i.e. yeasts in the
genus Candida., enterococci, E. coli, MRSA, fecal coliforms, enterovirus. The study
elucidated some relationships between indicator microbes, pathogens and the
environment; however due to the rarity of pathogenic incidence, there is still
remains a need to develop more novel and comprehensive approaches to assess
the prevalence of sand pathogens and their potential impacts on human health
from sand exposure at recreational beaches.
OBJECTIVES
 Determine the presence, quantity and distribution of pathogenic
microbes in beach sand (in supratidal, intertidal, and subtidal zones)
 Assess their relationship with environmental parameters and indicator
microbes.
 Interpret results in the context of risk-based human health impacts and
beach monitoring practices.
RESULTS & DISCUSSION
MATERIALS & METHODS
Sand Sampling
800 cm.
Dry sand
Enterococci (MF) vs. Moisture Content
STATIONARY
OBJECT
800 cm.
500 cm.
Dry Sampling Line
Date
Time
Tide
8/2/2009
8:30 AM
1.39 m
8/3/2009
8:45 AM
1.57 m
8/4/2009
9:40 AM
1.77 m
8/17/2009
7:50 AM
2.17 m
8/18/2009
8:50 AM
2.22 m
8/19/2009
9:40 AM
2.41 m
6 Sampling Events
Wet Sampling Line
Microbial Analysis of Water and Sand
Tidal Zone
 Enterococci
 EPA recommended marine bacterial indicator
 Analyzed by MF, chromogenic substrate, and qPCR.
Inundated Sampling Line
 Fecal coliforms, E. coli, and C. perfringens
 Analyzed by MF
Three Sampling Lines
 1 dry, wet, and inundated sand
sample (each with 160 cores)
 Each Sand Core taken every 10 cm.
Mixed thoroughly and homogenized
Added to 10 L of PBS and mixed
vigorously for 10 minutes
 allowed to settle for 10 minutes
Microbe Levels in Sand and Aqueous Samples
Indicator
Bacteria
 HPyV & ESP gene for enterococci
 Analyzed by PCR
 Bacteroidales (Dog, Human HF8 & UCD, gull)
 Analyzed by PCR
 S. aureus & Vibrio vulnificus
Analyzed by culture and PCR
Sample Cores
 Yeasts (Candida spp.)
 Analyzed by culture and PCR
 Cryptosporidium & Giardia
 Analyzed by microscopy & qPCR (bilayer conc.)
STUDY SITE
Microbial
Source
Tracking
Pathogens
 Enterovirus, Norovirus and HAV
 Analyzed by qPCR
 Helminthes (Nematode eggs and larvae)
 Analyzed by microscopy
 Virginia Key within MiamiDade County, Florida, USA
Organism
 Pets are allowed
 Water samples taken at waist deep water
 Beach admission is free
 Wet and Inundated Sampling lines varied with tide
 Subtropical beach
Dry Sampling line
 No known pt source of
pollution
Wet Sampling line
Knee depth (varies based on tide)
Inundated Sampling line
 Frequently exceeds
enterococci standard
(104/100 ml) for single
samples.
Enterococci (MF)
Enterococci (CS)
Environmental Measurements
Enterococci(qPCR)
Fecal Coliform
 Physical-chemical
parameters
C. perfringens

 Volatile organic compounds
& moisture in sand

 Tidal height
 Beach conditions
 # bathers, dogs, & birds
 Wind speed & direction
Correlated with:
r
Moisture Content(%)
-0.822
Enterococci (CS)
0.939
Fecal Coliform
Organism
Correlated with:
r
Moisture Content
-0.815
White Yeasts
0.690
0.831
Nematode Larvae
0.701
Red Yeasts
0.758
Moisture Content(%)
-0.543
White Yeasts
0.748
48 hr. rainfall
0.503
Nematode Larvae
0.800
Nematode Larvae
0.607
Moisture Content(%)
-0.755
Turbidity
-0.626
White Yeasts
0.819
Wind Speed
-0.532
Nematode Larvae
0.740
Moisture Content(%)
-0.761
Moisture Content(%)
-0.558
Nematode Eggs
0.619
Total Bacteroidales
0.822
pH*
-0.693
Moisture Content
-0.816
Temperature**
-0.651
Red Yeasts
0.852
48 hr. rainfall
0.620
White Yeasts
0.782
Nematode Larvae
Moisture Content(%)
0.789
0.504
Red Yeast
White Yeast
S. aureus
Nematode Larvae
Nematode Eggs
SUMMARY & RECOMMENDATIONS
 pH, temperature, salinity,
turbidity in water
 Solar radiation
Hobie
Cat
Beach
Pathogens and Yeasts
Indicator Microbes
 Irregularly narrow beach (5
m).

ACKNOWLEDGEMENTS
This research was supported by NSF-NIEHS Oceans and Human Health
Program (NIEHS P50 ES12736 and NSF OCE 0432368/0911373) and the
NSF REU program (OCE 0432368)



Enterococci
 Analysis method comparisons: MF&CS directly related .
 Correlated with fecal coliforms, red and white yeasts and nematode larvae
Higher levels of indicator and pathogenic microbes during high tide and rain events
 Enterococci exceeded health standard
 Vibrio sp., enterovirus, helminthes & Cryptosporidium spp. were detected.
 More microbes were detected on last three days than first three days.
Indicators seem to work in detecting some pathogen presence, however, other pathogens seem to display
variable unrelated trends.
 Nematode Larvae, white yeasts, and S. aureus follow indicator microbe trends
 Cryptsporidium, enterovirus, and Candida spp. do not follow such trends.
Source points towards multiple non-pt sources
 Bathers, animals, re-growth in sand, etc…
Additional sampling needed to:
 Confirm the tidal and rain effects
 Establish a correlation between indicators & pathogens
Determine correlation between pathogen/indicator presence and human health through epidemiology
studies and QMRA.