Detailed Research Information

Detailed project information for
Study Plan Number 01071-01

Branch : Fish Health Branch

Study Plan Number : 01071-01

Study Title : Determination of bacterial species distributions in water and biofilms by molecular genetic assay

Starting Date : 06/01/1996

Completion Date : 09/30/2002

Principal Investigator(s) : Schill, Bane (William)

Primary PI : Schill, Bane (William)

Telephone Number : (304) 724-4438

Email Address : bane_schill@usgs.gov

SIS Number : 5002177

Primary Program Element : Fisheries and Aquatic Resources

Second Program Element : Fish and Aquatic Habitats

Status : Completed

Abstract : The application of molecular biological techniques to the study of microbial diversity has resulted in the first comprehensive phylogeny of microorganisms (Woese 1987). Investigations of rRNA diversity generally have found that sequences derived from a particular environmental type (ecotypes) have clustered into a few distinct phylogenetic groups (phylotypes). Many precedents exist in macroecology for the connection of particular phylotypes with ecotypes. The relationships, however, between phylotypes and ecotypes at the microbiological level are currently not well understood (Stahl 1995). Because bacteria react quickly to environmental insults via heat-shock mechanisms and genetic information is rather easily dispersed among microorganisms via conjugation, plasmid transfer, and transposon mediated mechanisms, we speculate that resistance elements may be widely dispersed among bacterial residents of environments chronically impacted by pollutants or physical stressors.

Research Summary:

The application of molecular biological techniques to the study of microbial diversity has resulted in the first comprehensive phylogeny of microorganisms. Investigations of rRNA diversity generally have found that sequences derived from a particular environmental type (ecotypes) have clustered into a few distinct phylogenetic groups (phylotypes). Many precedents exist in macroecology for the connection of particular phylotypes with ecotypes. The relationships, however, between phylotypes and ecotypes at the microbiological level are currently not well understood. Because bacteria react quickly to environmental insults via heat-shock mechanisms and genetic information is rather easily dispersed among microorganisms via conjugation, plasmid transfer, and transposon mediated mechanisms, we speculated that resistance elements may be widely dispersed among bacterial residents of environments chronically impacted by pollutants or physical stressors.

Rainbow trout cultured in a recycle system and fed oxytetracycline (OTC) at 2.5 gm/ 100 lbs of fish for 14 days developed resistant gut flora, but similar resistance did not develop among heterotrophic bacteria found in the fluidized sand biofilters. Additional studies with feces and sediment (composed of feces and uneaten food) were carried out. Heterotrophic counts were performed on R2A and Mueller Hinton media with and without 100 ug /ml OTC on individual trout feces (50-55 trout each sample period), on 12 fecal pools (5 trout/pool), and on sediment. Feces and sediment were sampled before OTC feeding; on days 3, 7, 10 and 14 OTC feeding; and 3, 7, 14, 21, 28, 35, and 42 day post OTC feeding. During the first cycle of OTC feeding, resistance (expressed as average % of total heterotrophic counts on both media) in feces of single and pooled trout increased from 11% before OTC feeding to 73% by day 14. Upon completion of OTC feeding resistance dropped to 35.9% by day three, increased slightly on day 14 post OTC feeding and dropped to 1.26% by day 21. The second cycle of OTC feeding was begun immediately after completion of the first cycle. The same general pattern of resistance was seen using fecal samples of individual fish as had been observed with pooled samples, and sediment resistance generally reflected that found in feces. Surprisingly, sediment resistance was no higher than that found in feces despite the fact that OTC levels in sediment were determined to be as high as 1900 ppm. A definite bimodal pattern of resistance was seen with the first peak on day 14 of the second OTC feeding cycle and the second about 14 days after OTC was withdrawn. A second population of rainbow trout was obtained to determine if these patterns of OTC resistance in feces were universal or population-specific. Two cycles of OTC feeding were given and the same sampling days of pooled feces and sediment were used. Results from two such trials showed that resistance patterns during the first feeding cycle were similar to those observed during the two trials of the first fish population. We did not, however, observe the bimodal response during the second OTC feedings that we had detected with the first fish population. Additionally, sediment resistance levels were generally not reflective of those obtained with feces in these trials.

Products:

Schill, W.B. & G.L. Bullock. 2002. Development of Oxytetracycline Resistance in Rainbow Trout (Oncorhynchus mykiss) Gut Flora and Sediment in a Recirculating Culture System 27th Annual Eastern Fish Health Workshop, Charleston, SC.

Schill, W.B., G.L. Bullock & E. Burchard. 2001. Development Of Oxytetracycline Resistance In Rainbow Trout (Oncorhynchus mykiss) Gut Flora And Biofilter Flora In A Recirculating System 26th Annual Eastern Fish Health Workshop, Shepherdstown, West Virginia.

Schill, W.B., G.L. Bullock, and V. Blazer. 2001. Development of Microbial Tetracycline Resistance in a Recirculating Fish Culture System: Study of the Potential Impacts of Aquaculture on the Natural Environment. Eastern Region Workshop on Emerging Issues in Water Quality, Orlando, FL.

Bullock, G.L., B. Schill & E. Burchard. 2000. Transmission of Aeromonas salmonicida and Yersinia ruckeri in a salmonid recycle system by means of fluidized sand biofilters. Proceedings of the 25th Annual Eastern Fish Health Workshop, John Carver Inn, Plymouth, Massachusetts.

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Branch :	Fish Health Branch
Study Plan Number :	01071-01
Study Title :	Determination of bacterial species distributions in water and biofilms by molecular genetic assay
Starting Date :	06/01/1996
Completion Date :	09/30/2002
Principal Investigator(s) :	Schill, Bane (William)
Primary PI :	Schill, Bane (William)
Telephone Number :	(304) 724-4438
Email Address :	bane_schill@usgs.gov
SIS Number :	5002177
Primary Program Element :	Fisheries and Aquatic Resources
Second Program Element :	Fish and Aquatic Habitats
Status :	Completed
Abstract :	The application of molecular biological techniques to the study of microbial diversity has resulted in the first comprehensive phylogeny of microorganisms (Woese 1987). Investigations of rRNA diversity generally have found that sequences derived from a particular environmental type (ecotypes) have clustered into a few distinct phylogenetic groups (phylotypes). Many precedents exist in macroecology for the connection of particular phylotypes with ecotypes. The relationships, however, between phylotypes and ecotypes at the microbiological level are currently not well understood (Stahl 1995). Because bacteria react quickly to environmental insults via heat-shock mechanisms and genetic information is rather easily dispersed among microorganisms via conjugation, plasmid transfer, and transposon mediated mechanisms, we speculate that resistance elements may be widely dispersed among bacterial residents of environments chronically impacted by pollutants or physical stressors. Research Summary: The application of molecular biological techniques to the study of microbial diversity has resulted in the first comprehensive phylogeny of microorganisms. Investigations of rRNA diversity generally have found that sequences derived from a particular environmental type (ecotypes) have clustered into a few distinct phylogenetic groups (phylotypes). Many precedents exist in macroecology for the connection of particular phylotypes with ecotypes. The relationships, however, between phylotypes and ecotypes at the microbiological level are currently not well understood. Because bacteria react quickly to environmental insults via heat-shock mechanisms and genetic information is rather easily dispersed among microorganisms via conjugation, plasmid transfer, and transposon mediated mechanisms, we speculated that resistance elements may be widely dispersed among bacterial residents of environments chronically impacted by pollutants or physical stressors. Rainbow trout cultured in a recycle system and fed oxytetracycline (OTC) at 2.5 gm/ 100 lbs of fish for 14 days developed resistant gut flora, but similar resistance did not develop among heterotrophic bacteria found in the fluidized sand biofilters. Additional studies with feces and sediment (composed of feces and uneaten food) were carried out. Heterotrophic counts were performed on R2A and Mueller Hinton media with and without 100 ug /ml OTC on individual trout feces (50-55 trout each sample period), on 12 fecal pools (5 trout/pool), and on sediment. Feces and sediment were sampled before OTC feeding; on days 3, 7, 10 and 14 OTC feeding; and 3, 7, 14, 21, 28, 35, and 42 day post OTC feeding. During the first cycle of OTC feeding, resistance (expressed as average % of total heterotrophic counts on both media) in feces of single and pooled trout increased from 11% before OTC feeding to 73% by day 14. Upon completion of OTC feeding resistance dropped to 35.9% by day three, increased slightly on day 14 post OTC feeding and dropped to 1.26% by day 21. The second cycle of OTC feeding was begun immediately after completion of the first cycle. The same general pattern of resistance was seen using fecal samples of individual fish as had been observed with pooled samples, and sediment resistance generally reflected that found in feces. Surprisingly, sediment resistance was no higher than that found in feces despite the fact that OTC levels in sediment were determined to be as high as 1900 ppm. A definite bimodal pattern of resistance was seen with the first peak on day 14 of the second OTC feeding cycle and the second about 14 days after OTC was withdrawn. A second population of rainbow trout was obtained to determine if these patterns of OTC resistance in feces were universal or population-specific. Two cycles of OTC feeding were given and the same sampling days of pooled feces and sediment were used. Results from two such trials showed that resistance patterns during the first feeding cycle were similar to those observed during the two trials of the first fish population. We did not, however, observe the bimodal response during the second OTC feedings that we had detected with the first fish population. Additionally, sediment resistance levels were generally not reflective of those obtained with feces in these trials. Products: Schill, W.B. & G.L. Bullock. 2002. Development of Oxytetracycline Resistance in Rainbow Trout (Oncorhynchus mykiss) Gut Flora and Sediment in a Recirculating Culture System 27th Annual Eastern Fish Health Workshop, Charleston, SC. Schill, W.B., G.L. Bullock & E. Burchard. 2001. Development Of Oxytetracycline Resistance In Rainbow Trout (Oncorhynchus mykiss) Gut Flora And Biofilter Flora In A Recirculating System 26th Annual Eastern Fish Health Workshop, Shepherdstown, West Virginia. Schill, W.B., G.L. Bullock, and V. Blazer. 2001. Development of Microbial Tetracycline Resistance in a Recirculating Fish Culture System: Study of the Potential Impacts of Aquaculture on the Natural Environment. Eastern Region Workshop on Emerging Issues in Water Quality, Orlando, FL. Bullock, G.L., B. Schill & E. Burchard. 2000. Transmission of Aeromonas salmonicida and Yersinia ruckeri in a salmonid recycle system by means of fluidized sand biofilters. Proceedings of the 25th Annual Eastern Fish Health Workshop, John Carver Inn, Plymouth, Massachusetts.
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U.S. Department of the Interior \|\| U.S. Geological Survey 11700 Leetown Road, Kearneysville, WV 25430, USA URL: http://www.lsc.usgs.gov Maintainer: lsc_webmaster@usgs.gov Last Modified: October 21, 2002 dwn Privacy Policy and Disclaimers \|\| FOIA \|\| Accessibility