Program ASM Alaska Branch Meeting
Friday 4/12/2013
3:30pm Dr. Janice Matthews-Greer from the LSU Health Sciences Center in Shreveport
(LSUHSC-S)
Overview of Molecular Infectious Disease Testing
Reception and Beer Tasting to follow
7 pm No-Host Dinner at Glacier Brew House.
Saturday 4/13/2013
9:00 Business meeting
10:00 am Coffee break
10:15 Presentations
Dr. Janice Matthews-Greer (LSUHSC-S):
Respiratory Viruses Update and Review
12:00 pm Lunch break
1:00 pm Presentations:
Karen Rudolph (CDC)
Molecular Epidemiology of Invasive Group A Streptococcal Disease in Alaska, 2001 –
2011
Samantha Case (APHL/CDC):
Population structure of invasive pneumococcal isolates among Alaskan children in the
post-conjugate vaccine era
Robert A. Burgess (UAF):
Synergistic Degradation of Lignocellulose by Fungi and Bacteria in Boreal Forest Soil
1:50 pm Break
2:00 pm Presentations
James Fish (Alaska DEC):
Overview of Sulfolane as an Emerging Contaminant in the Tanana River Aquifer
Christopher Kasanke (UAF)
Microbial Ecology of Natural and Accelerated Sulfolane Biodegredation in Contaminated
Subarctic Aquifers
Robert Burgess (UAF):
Detection of Sulfolane Degrading Isolates in Contaminated Groundwater using T-RFLP
2:50 Break
3:00 pm Presentations
Jasmine Carroll and Katherine Lange (UAA)
Arctic Bioluminescent Bacteria: Effects of Temperature and pH
Kendall Neilsen and Ashley Mueller (UAA)
Acetic Acid Production by Gluconacetobacter Isolated from Kombucha Community
Molecular Epidemiology of Invasive Group A Streptococcal Disease in Alaska, 2001 – 2011.
Karen Rudolph, Michael Bruce, Dana Bruden, Tammy Zulz, Alisa Reasonover, Debby Hurlburt,
Thomas Hennessy
Arctic Investigations Program, Centers for Disease Control and Prevention, 4055 Tudor Center
Drive, Anchorage, Alaska
Background. Streptococcus pyogenes (groupA streptococci; GAS) is an exclusively human
pathogen that is most commonly associated with pharyngitis and skin infections but can also
cause more serious invasive infections including puerperal sepsis, bacteremia, pneumonia,
meningitis, necrotizing fasciitis, and streptococcal toxic shock syndrome. Since the mid-1980s, a
surge in the incidence and severity of GAS infections has been documented worldwide, resulting
in significant morbidity and mortality. The Arctic Investigations Program (AIP) began
surveillance for invasive GAS infections in Alaska in 2000 as part of the invasive bacterial
diseases population-based laboratory surveillance program.
Methods. A case of invasive GAS infection was defined as the isolation of group A
Streptococcus from a sterile site or from a wound culture accompanied by necrotizing fasciitis or
streptococcal toxic shock syndrome in a resident of Alaska. Isolates were forwarded to the
laboratory at AIP for confirmation, antimicrobial susceptibility testing and emm typing.
Demographic and clinical data on cases were collected by medical record review.
Results. Between 2001 and 2011, there were 416 cases of GAS reported, for an overall annual
incidence of 5.6 cases per 100,000 persons with 46 deaths (case-fatality ratio, 11.2%). Of the
416 confirmed cases of invasive GAS, 329 (79%) had isolates available for laboratory analysis.
The highest proportion of resistance was found for tetracycline (n= 34; 13.6%). A total of 41
emm types were identified among 329 isolates, of which emm1 (12.5%), was the most prevalent
followed by emm82 (10.4%), emm12 and emm49 (7.0% each) and emm108 (6.1%). The emm
types in a proposed 26-valent vaccine accounted for 55% of all cases and 63% of all fatal cases.
Conclusions. GAS remains an important cause of invasive bacterial disease in Alaska.
Continued surveillance of GAS infections will help improve understanding of the epidemiology
of invasive disease, with impact on disease control, notification of outbreaks, and vaccine
development.
Population structure of invasive pneumococcal isolates among Alaskan children in the
post-conjugate vaccine era.
Samantha Case
APHL/CDC Emerging Infectious Diseases Training Fellow
CDC Arctic Investigations Program
Introduction of conjugate vaccines against Streptococcus pneumoniae have led to a significant
decrease in overall rates of invasive pneumococcal disease (IPD), particularly in children less
than five years of age. However, cases of IPD caused by nonvaccine serotypes have increased.
Thus, it is critical to continue monitoring the molecular epidemiology of IPD. Sterile site isolates
received at AIP from children under five years of age from 2001-2012 have been serotyped and
tested for antibiotic resistance. Multilocus sequence typing is ongoing and will be used to
characterize the genetic background of the isolates. Using the data produced by MLST, a
population snapshot of the IPD isolates will be generated to determine if there have been changes
in the clonal composition of the isolates since the introduction of the two conjugate vaccines:
PCV7 in 2001 and PCV13 in 2010.
Synergistic Degradation of Lignocellulose by Fungi and Bacteria in Boreal Forest Soil
Robert A. Burgess, D. Lee Taylor, Kelsie Stone, Mary Beth Leigh
University of Alaska Fairbanks
Boreal forests contain an estimated 28% of the world’s soil carbon, and currently act as a
significant global carbon sink. Plant-derived lignocellulose is a major component of soil carbon,
and its decomposition is mediated by soil microbes (bacteria and fungi). In order to predict the
fate of this soil carbon and its potential feedbacks to climate change, the identities and
interactions of soil microbial decomposer communities must be better understood. This study
uses stable isotope probing (SIP) with 13C-labeled lignocellulose and two of its constituents,
cellulose and vanillin, to evaluate the hypothesis that lignocellulose decomposition occurs
synergistically in soil. Results indicate that multiple taxa are involved in lignocellulose
degradation, and that certain taxa preferentially target specific portions of the lignocellulose
macromolecule. These data support the synergistic degradation hypothesis, provide some of the
strongest evidence to date that different “guilds” of microbes preferentially target different
components of plant litter, and provide information on the identity of members of different
guilds. This information provides knowledge of small-scale microbial processes that dictate
ecosystem-level carbon cycling, and can assist in predictions of the fate of boreal forest carbon
stocks.
Overview of Sulfolane as an Emerging Contaminant in the Tanana River Aquifer.
James Fish, Environmental Project Specialist, Alaska Department of Environmental
Conservation, Contaminated Sites Program
In 2009, the compound sulfolane was detected in drinking water wells of North Pole, Alaska
residents, located down-gradient from the Flint Hills Resources North Pole Refinery. This
presentation will provide a brief overview on refinery operations, a background on the spill
history of sulfolane, and a synopsis of the Alaska Department of Environmental Conservation’s
efforts to understand the biodegradation and fate and transport of this emerging contaminant in
subsurface soil and groundwater of the Tanana River aquifer.
Microbial Ecology of Natural and Accelerated Sulfolane Biodegradation in Contaminated
Subarctic Aquifers
C. P. Kasanke, M.B. Leigh
University of Alaska Fairbanks
Sulfolane is a molecule that is used to de-acidify natural gas and selectively remove lighter
aromatics from petroleum. Sulfolane has a high affinity for water and has been found in the
aquifers surrounding sites where it has been improperly stored or disposed. To date there are
only three contaminated regions where sulfolane degradation has been studied, sour gas
treatment plants in western Canada, a sulfinol waste sludge disposal site in Brisbane, Australia,
and a pesticide manufacturing plant in Lathrop, California. Little is known about sulfolane
biodegradation in aquifers containing permafrost. There are also many knowledge gaps
regarding the microbial community responsible for sulfolane degradation. A sulfolane plume
located in North Pole, Alaska provides a unique opportunity to answer these important
fundamental questions. The goals of this project are to: 1) Assess the potential for aerobic and
anaerobic microbial degradation of sulfolane in sub-arctic soil and groundwater. 2) Identify
products and intermediates of biodegradation. 3) Indentify members of the microbial community
involved in sulfolane degradation using stable isotope probing with 13C labeled sulfolane.
4) Determine if microbial degradation contributes to the effectiveness of two engineered
remediation systems being actively used at a contaminated site: air sparging and activated carbon
water treatment systems. This research will provide necessary insights into the environmental
fate of sulfolane as well as its impacts on the microbial communities associated with the aquifer.
Detection of Sulfolane Degrading Isolates in Contaminated Groundwater using T-RFLP
Robert Burgess1, Michael Ortego1, Kaitlynn McKirgan1, Jacob Howell1, Christopher Kasanke 1,
Jim Fish2, Mary Beth Leigh1
1
University of Alaska Fairbanks
2
Alaska Department of Environmental Conservation
Sulfolane, a chemical used in refining petroleum, is a recently identified (emerging)
environmental contaminant. While previous studies have demonstrated that sulfolane can be
degraded by microbes, very little is known about the identity and distribution of sulfolane
degraders in the environment or in sulfolane contaminated groundwater. This study examined the
identity and distribution of sulfolane degraders in the Flint Hills Refinery (FHR) sulfolane plume
located in North Pole, Alaska. Twenty-two sulfolane degrading bacteria were isolated from
plume groundwater by using previously developed differential media followed by growth on
agar plates containing sulfolane as the sole carbon source (SOCS). Sulfolane degradation was
confirmed by inoculating liquid SOCS media with each isolate and enumerating bacteria with
plate counts over a time course. Terminal restriction fragment length polymorphism (T-RFLP)
analysis was performed on a subset of isolates to determine the length of their terminal
restriction fragments (T-RFs). T-RFLP was also performed on groundwater samples from within
the FHR plume, including from an area influenced by an air sparging pilot system. Microbial
communities within the plume appear to be affected by sulfolane concentration, the presence of
petroleum co-contaminants, and the air sparge system. Additionally, all groundwater samples
from within the plume contain a T-RF of the same size as the isolate T-RF in high abundance.
Air sparging increases the relative abundance of this T-RF in the microbial community. These
results suggest that sulfolane degrading bacteria are present in the plume and that air sparging
may increase their abundance.
Arctic Bioluminescent Bacteria: Effects of Temperature and pH
Authors: Jasmine Carroll and Katherine Lange
University of Alaska Anchorage
Bioluminescent bacteria were isolated from the gills of an Alaskan Rockfish. These bacteria
exhibited growth on synthetic seawater agar as well as Thiosulfate Citrate Bile salts Sucrose
(TCBS) agar. Colonies exhibited blue-green bioluminescence in a dark room after 3 days of
incubation at 15˚C, fading after 5 days. DNA from isolated organisms was sequenced using 16S
primers for identification. The isolates were identified as Photobacterium phosphorium. Acid
and temperature tolerance of isolates will be evaluated in order to establish whether free-living
arctic bioluminescent bacteria can withstand changing ocean temperatures and acidification.
Broth cultures will be grown at temperatures from 0-28˚C as well as pH levels from 8.0-5.0. The
pH conditions will be tested concurrently with temperature conditions so that each possible
scenario is analyzed. The focus of our research is therefore to complement the existing
understanding of the effects of temperature and pH variation on the growth rates of
bioluminescent bacteria.
Acetic Acid Production by Gluconacetobacter spp. Isolated from Kombucha Community
Kendall Neilsen and Ashley Mueller
University of Alaska Anchorage
Kombucha is a popular fermented beverage originating in China that is prepared by the
fermentation of sweetened tea by a symbiosis of acetic bacteria and yeast strains. Currently
several strains within the Gluconacetobacter and Acetobacter genera have been identified in
kombucha samples with Gluconacetobacter xylinus being the dominant strain. This goal of this
research was to isolate G. xylinus from a kombucha sample and investigate the ethanol tolerance
and qualitatively measure acetic acid production. The formation of a cellulose pellicle suggested
that the isolates were G. xylinus. Glucose yeast calcium carbonate (GYCA) enrichment agar and
Frateur’s medium was used to select and differentiate between genera, while an antifungal agent
was used to inhibit any yeast growth. Further biochemical tests including catalase and oxidase
tests, and the presence of a capsule confirmed that the isolated strain was G. xylinus. After the
strain was isolated, it was cultured in various ethanol concentrations to determine the optimal
ethanol concentration required to produce the maximum amount of acetic acid.