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Saturday, 18 February 2017

Biocontrol of Fungi Causing White Nose Syndrome


[photo:] White nose syndrome infected little brown bat.  White nose syndrome appears on the noses, wings, ears and feet of infected bats. It is a deadly disease affecting cave hibernating bats. Photo by Al Hicks, NYSDEC, Bugwood.orgResearch Issue

White nose syndrome is caused by a cold-loving fungus, Pseudogymnoascus destructans, which grows on the bats’ skin. P. destructans is deadly to hibernating bats because it penetrates tissues of the nose and mouth as well as the wings, which are vital to bats’ ability to avoid dehydration and maintain body temperature. Infected bats wake up more frequently than normal, exhausting their limited energy reserves long before spring.
In affected hibernacula, 78 to 100% of bat populations have died; total overall deaths so far are estimated at between 5 and 6 million bats since WNS was first recorded in 2006-2007 in upstate New York. 

Our Research

Dr. Sybill Amelon, a research wildlife biologist with the Northern Research Station (NRS), is collaborating with Gary Knudsen of the University of Idaho, the NRS is evaluating the potential of naturally occurring fungi on bat skin to combat P. destructans and potentially provide protection for susceptible bat species.  Amelon is working with Knudsen and others to evaluate fungi and bacteria normally found on bat skin to determine if they might have usefulness in biological control of WNS fungus.
Fungal disease in vertebrates is relatively rare, perhaps the most similar example is Chytridiomycoses found in amphibians.  Biological control is one approach that has shown promise in that disease for several species.  Animal skin contains a community of ‘normal microbiota' that provides a first line of defense against some pathogens.  Current research on the complex interplay between the skin microbiota and the innate immune system of the skin indicates a beneficial role, much like that of the gut microbiota. It is likely that, under certain conditions, the normal microbiota of bat skin is preemptive or inhibitory to  P. destructans, the degree to which this naturally occurs, and the possibility of augmenting any natural protective microbial communities, remain to be determined. 

The focus of this research is to obtain representative isolates of skin bacteria and fungi from selected bat species, to screen them for antagonistic activity towards P. destructans, and ultimately to test the most promising antagonists as potential biological control agents for WNS in the laboratory. Environmental and nutritional growth requirements of these bacteria or fungi will then be further characterized. 

Expected Outcomes

The goal of this research is to identify potential bio-control agents that will increase the survival of bats exposed to white nose syndrome.

Research Results

Initial tests recovered a large number and variety of fungi from the sampled bats, with a smaller representation of bacterial types. Some phenotypes were common among the bats in each group studied, and some were apparently present in samples from both locations. Additional identification will be needed to confirm this observation. These samples will be evaluated for their potential to help bats resist WNS. This work was a first step in identifying the microbiota found on healthy bats. By comparing and contrasting the microbiota of species highly susceptible to P. destructans to the microbiota of other species that are less susceptible, we hope to find isolates that can safely be used to increase resistance in the most susceptible bat species.  We are collaborating with mycologists to develop an approach that targets the host-pathogen interrelationship. 

Related Research

Amelon has studied resource selection, relative abundance of 3 maternity colonies of Indiana bat before arrival of WNS in Missouri colonies; the second phase of this study will examine these same factors as WNS becomes more prevalent .  Published as 
Womack, K.M.; Amelon, S.K.; Thompson, F.R.. 2013.  Resource selection by Indiana bats during the maternity season. The Journal of Wildlife Management: published online January 2013.
In another study that has not yet been published, Amelon and colleagues found that improved analysis of long-term monitoring data demonstrates marked regional declines of bat populations in the eastern United States before appearance of WNS.

Research Participants


For further details log on website :
https://www.nrs.fs.fed.us/disturbance/invasive_species/wns/control_management/biocontrol_fungus/

DNA-based detection of the fungal pathogen Geomyces destructans in soils from bat hibernacula

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Year Published

2011

Source

Mycologia. 103(2): 241-246.

Abstract

White-nose syndrome (WNS) is an emerging disease causing unprecedented morbidity and mortality among bats in eastern North America. The disease is characterized by cutaneous infection of hibernating bats by the psychrophilic fungus Geomyces destructans. Detection of G. destructans in environments occupied by bats will be critical for WNS surveillance, management and characterization of the fungal lifecycle. We initiated an rRNA gene region-based molecular survey to characterize the distribution of G. destructans in soil samples collected from bat hibernacula in the eastern United States with an existing PCR test. Although this test did not specifically detect G. destructans in soil samples based on a presence/absence metric, it did favor amplification of DNA from putative Geomyces species. Cloning and sequencing of PCR products amplified from 24 soil samples revealed 74 unique sequence variants representing 12 clades. Clones with exact sequence matches to G. destructans were identified in three of 19 soil samples from hibernacula in states where WNS is known to occur. Geomyces destructans was not identified in an additional five samples collected outside the region where WNS has been documented. This study highlights the diversity of putative Geomyces spp. in soil from bat hibernacula and indicates that further research is needed to better define the taxonomy of this genus and to develop enhanced diagnostic tests for rapid and specific detection of G. destructans in environmental samples.

Keywords

Citation

Lindner, Daniel L.; Gargas, Andrea; Lorch, Jeffrey M.; Banik, Mark T.; Glaeser, Jessie; Kunz, Thomas H.; Blehert, David S. 2011. DNA-based detection of the fungal pathogen Geomyces destructans in soils from bat hibernacula. Mycologia. 103(2): 241-246.

Last updated on: October 20, 2011

For further details log on website :
https://www.nrs.fs.fed.us/pubs/39451

Bat white-nose syndrome: A real-time TaqMan polymerase chain reaction test targeting the intergenic spacer region of Geomyces destructans

Author
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  • This publication is available only online.
  • Muller, Laura K
  • Lorch, Jeffrey M.
  • Lindner, Daniel L.
  • O'Connor, Michael
  • Gargas, Andrea
  • Blehert, David S.

Year Published

2013

Source

Mycologia. 105(2): 253-259.

Abstract

The fungus Geomyces destructans is the causative agent of white-nose syndrome (WNS), a disease that has killed millions of North American hibernating bats. We describe a real-time TaqMan PCR test that detects DNA from G. destructans by targeting a portion of the multicopy intergenic spacer region of the rRNA gene complex. The test is highly sensitive, consistently detecting as little as 3.3 fg of genomic DNA from G. destructans. The real-time PCR test specifically amplified genomic DNA from G. destructans but did not amplify target sequence from 54 closely related fungal isolates (including 43 Geomyces spp. isolates) associated with bats. The test was further qualified by analyzing DNA extracted from 91 bat wing skin samples, and PCR results matched histopathology findings. These data indicate the real-time TaqMan PCR method described herein is a sensitive, specific, and rapid test to detect DNA from G. destructans and provides a valuable tool for WNS diagnostics and research.

Keywords

Citation

Muller, Laura K; Lorch, Jeffrey M.; Lindner, Daniel L.; O'Connor, Michael; Gargas, Andrea; Blehert, David S. 2013. Bat white-nose syndrome: A real-time TaqMan polymerase chain reaction test targeting the intergenic spacer region of Geomyces destructans. Mycologia. 105(2): 253-259.

Last updated on: April 12, 2013

For further details log on website :
https://www.nrs.fs.fed.us/pubs/42974

Phylogenetic evaluation of Geomyces and allies reveals no close relatives of Pseudogymnoascus destructans, comb nov, in bat hibernacula of eastern North America

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Year Published

2013

Source

Fungal Biology. 117: 638-649.

Abstract

White-nose syndrome (WNS) of bats, caused by the fungus previously known as Geomyces destructans, has decimated populations of insectivorous bats in eastern North America. Recent work on fungi associated with bat hibernacula uncovered a large number of species of Geomyces and allies, far exceeding the number of described species. Communication about these species has been hindered by the lack of a modern taxonomic evaluation, and a phylogenetic framework of the group is needed to understand the origin of G. destructans and to target closely related species and their genomes for the purposes of understanding mechanisms of pathogenicity. We addressed these issues by generating DNA sequence data for the internal transcribed spacer (ITS) region, nuclear large subunit (LSU) rDNA, MCM7, RPB2, and TEF1 from a diverse array of Geomyces and allies that included isolates recovered from bat hibernacula as well as those that represent important type species. Phylogenetic analyses indicate Geomyces and allies should be classified in the family Pseudeurotiaceae, and the genera Geomyces, Gymnostellatospora, and Pseudogymnoascusshould be recognized as distinct. True Geomyces are restricted to a basal lineage based on phylogenetic placement of the type species, Geomyces auratus. Thus, G. destructans is placed in genus Pseudogymnoascus. The closest relatives of Pseudogymnoascus destructans are members of the Pseudogymnoascus roseus species complex,however, the isolated and long branch of P. destructansindicates that none of the species included in this study are closely related, thus providing further support to the hypothesis that this pathogen is non-native and invasive in eastern North America. Several conidia-producing isolates from bat hibernacula previously identified as members of Pseudeurotium are determined to belong to the genus Leuconeurospora, which is widespread, especially in colder regions. Teberdinia hygrophila is transferred to Pseudeurotium as Pseudeurotium hygrophilum, comb. nov., in accordance with the one name per fungus system of classification, and two additional combinations are made in Pseudogymnoascus including Pseudogymnoascus carnis and Pseudogymnoascus pannorum. Additional sampling from other regions of the world is needed to better understand the evolution and biogeography of this important and diverse group of fungi.

Keywords

Citation

Minnis, Andrew M.; Lindner, Daniel L. 2013. Phylogenetic evaluation of Geomyces and allies reveals no close relatives of Pseudogymnoascus destructans, comb. nov., in bat hibernacula of eastern North America. Fungal Biology. 117: 638-649.

Last updated on: September 13, 2013

For further details log on website :
https://www.nrs.fs.fed.us/pubs/44379

A culture-based survey of fungi in soil from bat hibernacula in the eastern United States and its implications for detection of Geomyces destructans, the causal agent of bat white-nose syndrome

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  • This publication is available only online.

Year Published

2013

Source

Mycologia. 105(2): 237-252.

Abstract

The recent emergence of white-nose syndrome (WNS), a fungal disease causing unprecedented mortality among hibernating bats of eastern North America, has revealed a knowledge gap regarding fungal communities associated with bats and their hibernacula. We used culture-based techniques to investigate the diversity of fungi in soil samples collected from 24 bat hibernacula in the eastern United States. Ribosomal RNA regions (internal transcribed spacer and partial intergenic spacer) were sequenced to preliminarily characterize isolates. Geomyces species were one of the most abundant and diverse groups cultured, representing approximately 33% of all isolates. Geomyces destructans was isolated from soil samples from three hibernacula in states where WNS is known to occur, and many of the other cultured Geomyces isolates likely represent undescribed taxa. Further characterization of the diversity of fungi that occur in hibernacula will both facilitate an improved understanding of the ecology of G. destructans within this complex fungal community and provide an opportunity to identify characteristics that differentiate G. destructans from non-pathogenic relatives.

Keywords

Citation

Lorch, Jeffrey M.; Lindner, Daniel L.; Gargas, Andrea; Muller, Laura K; Minnis, Andrew M.; Blehert, David S. 2013. A culture-based survey of fungi in soil from bat hibernacula in the eastern United States and its implications for detection of Geomyces destructans, the causal agent of bat white-nose syndrome. Mycologia. 105(2): 237-252.

Last updated on: April 12, 2013

For further details log on website :
https://www.nrs.fs.fed.us/pubs/42973

Distribution and environmental persistence of the causative agent of white-nose syndrome, Geomyces destructans, in bat hibernacula of the eastern United States

Author
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  • This publication is available only online.
  • Lorch, Jeffrey M.
  • Muller, Laura K.
  • Russell, Robin E.
  • O'Connor, Michael
  • Lindner, Daniel L.
  • Blehert, David S.

Year Published

2013

Source

Applied and Environmental Microbiology. 79(4): 1293-1301

Abstract

>White-nose syndrome (WNS) is an emerging disease of hibernating bats caused by the recently described fungus Geomyces destructans. First isolated in 2008, the origins of this fungus in North America and its ability to persist in the environment remain undefined. To investigate the correlation between manifestation of WNS and distribution of G. destructans in the United States, we analyzed sediment samples collected from 55 bat hibernacula (caves and mines) both within and outside the known range of WNS using a newly developed real-time PCR assay. Geomyces destructans was detected in 17 of 21 sites within the known range of WNS at the time when the samples were collected; the fungus was not found in 28 sites beyond the known range of the disease at the time when environmental samples were collected. These data indicate that the distribution of G. destructans is correlated with disease in hibernating bats and support the hypothesis that the fungus is likely an exotic species in North America. Additionally, we examined whether G. destructans persists in infested bat hibernacula when bats are absent. Sediment samples were collected from 14 WNS-positive hibernacula, and the samples were screened for viable fungus by using a culture technique. Viable  G. destructans was cultivated from 7 of the 14 sites sampled during late summer, when bats were no longer in hibernation, suggesting that the fungus can persist in the environment in the absence of bat hosts for long periods of time.

Keywords

Citation

Lorch, Jeffrey M.; Muller, Laura K.; Russell, Robin E.; O’Connor, Michael; Lindner, Daniel L.; Blehert, David S. 2013. Distribution and environmental persistence of the causative agent of white-nose syndrome, Geomyces destructans, in bat hibernacula of the eastern United States. Applied and Environmental Microbiology. 79(4): 1293-1301. doi: 10.1128/​AEM.02939-12

Last updated on: April 18, 2013

For further details log on website :
https://www.nrs.fs.fed.us/pubs/43280

Molecular characterization of a heterothallic mating system in Pseudogymnoascus destructans, the fungus causing white-nose syndrome of bats

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Year Published

2014

Source

Genes|Genomes|Genetics. 4(9): 1755-1763.

Abstract

White-nose syndrome (WNS) of bats has devastated bat populations in eastern North America since its discovery in 2006. WNS, caused by the fungus Pseudogymnoascus destructans, has spread quickly in North America and has become one of the most severe wildlife epidemics of our time. While P. destructans is spreading rapidly in North America, nothing is known about the sexual capacity of this fungus. To gain insight into the genes involved in sexual reproduction, we characterized the mating-type locus (MAT) of two Pseudogymnoascus spp. that are closely related to P. destructans and homothallic (self-fertile). As with other homothallic Ascomycota, the MAT locus of these two species encodes a conserved a-box protein (MAT1-1-1) as well as two high-mobility group (HMG) box proteins (MAT1-1-3 and MAT1-2-1). Comparisons with the MATlocus of the North American isolate of P. destructans (the ex-type isolate) revealed that this isolate of P. destructans was missing a clear homolog of the conserved HMG box protein (MAT1-2-1). These data prompted the discovery and molecular characterization of a heterothallic mating system in isolates of P. destructans from the Czech Republic. Both mating types of P. destructans were found to coexist within hibernacula, suggesting the presence of mating populations in Europe. Although populations of P. destructansin North America are thought to be clonal and of one mating type, the potential for sexual recombination indicates that continued vigilance is needed regarding introductions of additional isolates of this pathogen.

Keywords

Citation

Palmer, Jonathan M.; Kubatova, Alena; Novakova, Alena.; Minnis, Andrew M.; Kolarik, Miroslav; Lindner, Daniel L. 2014. Molecular characterization of a heterothallic mating system in Pseudogymnoascus destructans, the fungus causing white-nose syndrome of bats. Genes|Genomes|Genetics. 4(9): 1755-1763.

Last updated on: September 26, 2014

For further details log on website :
https://www.nrs.fs.fed.us/pubs/46717

The fungus Trichophyton redellii sp. nov. causes skin infections that resemble white-nose syndrome of hibernating bats

Author
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  • This publication is available only online.
  • Lorch, Jeffrey M.
  • Minnis, Andrew M.
  • Meteyer, Carol U.
  • Redell, Jennifer A.
  • White, J. Paul
  • Kaarakka, Heather M.
  • Muller, Laura K.
  • Lindner, Daniel L.
  • Verant, Michelle L.
  • Shearn-Bochsler, Valerie
  • Blehert, David S.

Year Published

2015

Source

Journal of Wildlife Diseases. 51(1): 36-47.

Abstract

Before the discovery of white-nose syndrome (WNS), a fungal disease caused by Pseudogymnoascus destructans, there were no reports of fungal skin infections in bats during hibernation. In 2011, bats with grossly visible fungal skin infections similar in appearance to WNS were reported from multiple sites in Wisconsin, US, a state outside the known range of P. destructans and WNS at that time. Tape impressions or swab samples were collected from affected areas of skin from bats with these fungal infections in 2012 and analyzed by microscopy, culture, or direct DNA amplification and sequencing of the fungal internal transcribed spacer region (ITS). A psychrophilic species of Trichophyton was isolated in culture, detected by direct DNA amplification and sequencing, and observed on tape impressions. Deoxyribonucleic acid indicative of the same fungus was also detected on three of five bat carcasses collected in 2011 and 2012 from Wisconsin, Indiana, and Texas, US. Superficial fungal skin infections caused by Trichophyton sp. were observed in histopathology for all three bats. Sequencing of the ITS of Trichophyton sp., along with its inability to grow at 25 C, indicated that it represented a previously unknown species, described herein as Trichophyton redellii sp. nov. Genetic diversity present within T. redellii suggests it is native to North America but that it had been overlooked before enhanced efforts to study fungi associated with bats in response to the emergence of WNS.

Keywords

Citation

Lorch, Jeffrey M.; Minnis, Andrew M.; Meteyer, Carol U.; Redell, Jennifer A.; White, J. Paul; Kaarakka, Heather M.; Muller, Laura K.; Lindner, Daniel L.; Verant, Michelle L.; Shearn-Bochsler, Valerie; Blehert, David S. 2015. The fungus Trichophyton redellii sp. nov. causes skin infections that resemble white-nose syndrome of hibernating bats. Journal of Wildlife Diseases. 51(1): 36-47. 

Last updated on: January 15, 2015

For further details log on website :
https://www.nrs.fs.fed.us/pubs/47364

Rapid, Reliable Detection of the White Nose Syndrome Pathogen


[photo:] Soil sample in the lab.Research Issue

The recent emergence of white-nose syndrome (WNS) in hibernating bats of eastern North America has revealed a knowledge gap regarding fungal communities associated with bats and their hibernacula. At best, only 5 to 10 percent of fungal species on Earth have been named and scientifically described. Developing a specific test for this fungus was difficult because every sample from bats and caves contained a huge diversity of unidentified, unnamed fungi, and these species interfered with detection.

Our Research

Drs. Daniel Lindner and Jessie Glaeser investigated the diversity of fungi in soil samples collected from 24 bat hibernacula in the eastern United States. Ribosomal RNA regions were sequenced for preliminary characterizations of the isolates. 

Expected Outcomes

[photo:] Geomyces destructans grown in the lab.  Photo by Dan Lindner, USFS NRS.Further characterization of the diversity of fungi that occur in hibernacula will both facilitate an improved understanding of the ecology of Pseudogymnoascus destructans(formerly known as Geomyces destructans) within this complex fungal community and provide an opportunity to identify characteristics that differentiate P. destructans from non-pathogenic relatives.  This work also lays the groundwork for finding virulent genes and potentially preventing them from being expressed.
Geomyces species were one of the most abundant and diverse groups cultured, representing about one-third of all isolates. P. destructans was isolated from soil samples from three hibernacula in states where WNS is known to occur, and many of the other cultured Geomyces isolates likely represent undescribed taxa. 
This work was a first step in developing a highly sensitive DNA-based technique for early identification of P. destructans on bats as well as in soils and on cave walls. The DNA-based technique is suitable for detecting P. destructans in almost any kind of sample, and the test is 100-fold more sensitive than the next best test. 

Research Results

Drees, K.P.; Palmer, J.M.; Sebra, R.; Lorch, J.M.; Chen, C.; Wu, C.C.; Bok, J.W.; Keller, N.P.; Blehert, D.S.; Cuomo, C.A.; Lindner, D.L.; Foster, J.T. 2016. Use of multiple sequencing technologies to produce a high-quality genome of the fungus Pseudogymnoascus destructans, the causative agent of bat white-nose syndrome. Genome Announcements, 4(3), pp.e00445-16.
Lorch, J.; Minnis, A.; Meteyer, C.; Redell, J.; White, J.; Kaarakka, H.; Muller, L.; Lindner, D.L.; Verant, M.; Shearn-Bochsler, V.; Blehert, D. 2015. The Fungus Trichophyton redellii sp. nov. Causes Skin Infections that Resemble White-nose Syndrome of Hibernating Bats. Journal of Wildlife Diseases 51(1):36-47. 
Shuey, M.M.; Drees, K.P.; Lindner, D.L.; Keim, P.; Foster, J.T. 2014. A highly sensitive qPCR for the detection and differentiation of Pseudogymnoascus destructans and Pseudogymnoascus species. Applied and Environmental Microbiology 80(5):1726-1731. 
Luna, T.; Lindner, D.L.; Dumroese, R.K. 2014. Roost trees—growing hickories (Carya spp.) to assist recovery of bat populations. Native Plants Journal 15: 66–74.
Palmer, J.M.; Kubatova, A.; Novakova, A.; Minnis, A.M.; Kolarik, M.; Lindner, D.L. 2014. Molecular characterization of a heterothallic mating system in Pseudogymnoascus destructans, the fungus causing white-nose syndrome of bats. G3: Genes, Genomes, Genetics 4(9):1755-1763. 
Lorch, J.M.; Muller, L.K.; Russell, R.E.; O’Connor, M.; Lindner, D..L; Blehert, D.S. 2013. Distribution and environmental persistence of the causative agent of white-nose syndrome, Geomyces destructans, in bat hibernacula of the eastern United States.  Applied and Environmental Microbiology 79(4): 1293-1301. 
Muller, Laura K; Lorch, Jeffrey M.; Lindner, Daniel L.; O'Connor, Michael; Gargas, Andrea; Blehert, David S. 2013. Bat white-nose syndrome: A real-time TaqMan polymerase chain reaction test targeting the intergenic spacer region of Geomyces destructans. Mycologia. 105(2): 253-259.
Lindner, Daniel L.; Gargas, Andrea; Lorch, Jeffrey M.; Banik, Mark T.; Glaeser, Jessie A.; Kunz, Thomas H.; Blehert, David S. 2011. DNA-based detection of the fungal pathogen Geomyces destructans in soils from bat hibernacula. Mycologia. 103(2): 241-246.

Research Participants

  • Daniel Lindner &  Jessie Glaeser, Research Plant Pathologists, US Forest Service, Northern Research Station
  • David Blehert, US Geological Survey, National Wildlife Health Center
  • Laura Muller, US Geological Survey, National Wildlife Health Center 
  • Jeffrey Lorch, University of Wisconsin 
  • Andrea Gargas, Symbiology LLC 
  • Michael O’Conner, Wisconsin Veterinary Diagnostics Lab

For further details log on website :
https://www.nrs.fs.fed.us/disturbance/invasive_species/wns/risk_detection_spread/rapid_detection_fungus/

Evaluation of eggshell membrane-based bio-adsorbent for solid-phase extraction of linear alkylbenzene sulfonates coupled with high-performance liquid chromatography.

Author

Related Articles Hover help
The potential of eggshell membrane (ESM) as a novel solid-phase extraction bio-adsorbent was investigated in the present study. The ESM with a unique structure of intricate lattice network showed a predominant ability to capture linear alkylbenzene sulfonates (LAS) as a model of organic pollutants by the hydrophobic interactions between ESM and LAS molecular at pH very close to the isoelectric point of ESM, which was similar to the most widely used trapping mechanism for SPE. Under the optimal conditions, the breakthrough capacities of the ESM packed cartridge for C10-C13 LAS homologues were found to be 30, 53, 50, and 43microgg(-1), respectively. On the basis of high-performance liquid chromatography separation and UV detection of LAS homologues, the proposed system could respond down to 0.027ngmL(-1) of LAS with a linear calibration range from 0.2 to 100ngmL(-1), showing a good LAS enrichment ability of eggshell membrane biomaterial with high sensitivity, and could be successfully used for the detection of residual LAS in environmental water samples. The reproducibility among columns was satisfactory (RSD among columns is less than 10%). A comparison study with ESM, C8 and C18 as adsorbents for LAS demonstrated that ESM-based bio-adsorbent was advantageous over C8 and C18, the widely used traditional adsorbents.


For further details log on website :
http://europepmc.org/abstract/med/20674925

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