Robbin Koenig - Academia.edu (original) (raw)

Papers by Robbin Koenig

Research paper thumbnail of Phosphatase Expression by Chlorella Vulgaris (Chlorophyceae) is Mediated by Internal Phosphorus Levels and External pH

Journal of Phycology, 2002

This study was conducted to evaluate four species of algae (Chromulina freiburgensis Dofl., Chlor... more This study was conducted to evaluate four species of algae (Chromulina freiburgensis Dofl., Chlorella ellipsoidea Gerneck, C. vulgaris Beijerinck, and Chlamydomonas acidophila Negoro) isolated from an acid pit lake for their bioremediative potential. The Berkeley Pit Lake system, located in Butte, MT, USA is an aban-4

Research paper thumbnail of A randomized controlled field trial of a novel trimethoprim-sulfadiazine oral suspension for treatment of Streptococcus equi subsp zooepidemicus infection of the lower respiratory tract in horses

Journal of the American Veterinary Medical Association, Jan 15, 2015

Objective-To evaluate the effectiveness of a novel trimethoprim-sulfadiazine oral suspension for ... more Objective-To evaluate the effectiveness of a novel trimethoprim-sulfadiazine oral suspension for the treatment of naturally acquired Streptococcus equi subsp zooepidemicus infection in horses. Design-Randomized, controlled field trial. Animals-180 horses with S equi subsp zooepidemicus infection. Procedures-Horses with lower respiratory tract infections caused by S equi subsp zooepidemicus were treated with a new formulation of combined trimethoprim-sulfadiazine oral suspension at a dosage of 24 mg/kg (10.9 mg/lb) twice daily for 10 days (treatment group) or with an equivalent volume of saline (0.9% NaCl) solution (placebo group). Response to treatment, including clinical signs and fecal consistency scores, was assessed twice daily. Any adverse effects were recorded. The primary outcome variable was clinical response; the secondary outcome variable was eradication of S equi subsp zooepidemicus on study day 17 as determined by bacteriologic culture of repeated transtracheal-wash spec...

Research paper thumbnail of A Controlled Safety Study of Elevated Dosages of Trimethoprim Plus Sulfadiazine in Mature Horses

Journal of Equine Veterinary Science, 2014

A novel, proprietary, oral suspension of trimethoprim plus sulfadiazine (TMP/SDZ) had no serious ... more A novel, proprietary, oral suspension of trimethoprim plus sulfadiazine (TMP/SDZ) had no serious adverse effects on clinical, laboratory, or pathologic parameters of mature horses when administered at up to five times the intended combined dosage of 24 mg/kg twice daily for 30 consecutive days. Loose feces was the most common observation that was likely related to TMP/SDZ treatment, and the incidence of loose feces was greater in groups treated with higher dosages of TMP/SDZ. However, all episodes were self-limiting, and no horses were treated for loose feces.

Research paper thumbnail of Safety and efficacy of a new formulation of trimethoprim sulfadiazine in horses

Safety and efficacy of a new formulation of trimethoprim sulfadiazine in horses

Research paper thumbnail of tRNA is the source of low-level trans-zeatin production in Methylobacterium spp

Journal of bacteriology, 2002

Pink-pigmented facultatively methylotrophic bacteria (PPFMs), classified as Methylobacterium spp.... more Pink-pigmented facultatively methylotrophic bacteria (PPFMs), classified as Methylobacterium spp., are persistent colonizers of plant leaf surfaces. Reports of PPFM-plant dialogue led us to examine cytokinin production by PPFMs. Using immunoaffinity and high-performance liquid chromatography (HPLC) purification, we obtained 22 to 111 ng of trans-zeatin per liter from culture filtrates of four PPFM leaf isolates (from Arabidopsis, barley, maize, and soybean) and of a Methylobacterium extorquens type culture originally recovered as a soil isolate. We identified the zeatin isolated as the trans isomer by HPLC and by a radioimmunoassay in which monoclonal antibodies specific for trans-hydroxylated cytokinins were used. Smaller and variable amounts of trans-zeatin riboside were also recovered. trans-Zeatin was recovered from tRNA hydrolysates in addition to the culture filtrates, suggesting that secreted trans-zeatin resulted from tRNA turnover rather than from de novo synthesis. The pro...

Research paper thumbnail of Staphylococcus aureus AgrA Binding to the RNAIII-agr Regulatory Region

Journal of Bacteriology, 2004

The control of virulence gene expression in the human pathogen Staphylococcus aureus is under the... more The control of virulence gene expression in the human pathogen Staphylococcus aureus is under the partial control of the two-component quorum-sensing system encoded by genes of the agr locus. The product of the agrA gene has been shown by amino acid sequence similarity to be the putative response regulator; however, binding of AgrA to promoters under its control has not yet been demonstrated. In this study, we isolated and purified soluble AgrA by expression under osmotic shock conditions and ion-exchange chromatography. Purified AgrA showed high-affinity binding to the RNAIII-agr intergenic region by electrophoretic mobility shift assays. Binding was localized by DNase I protection assays to a pair of direct repeats in the P2 and P3 promoter regions of the agr locus. We found that this binding was enhanced by the addition of the small phosphoryl donor, acetyl phosphate. The difference in binding affinity between these two promoters was found to result from a 2-bp difference between the downstream direct repeats of the P2 and P3 sites. Mutation of these base pairs in the P3 site to match those found in the P2 site increased the affinity of AgrA for the P3 site relative to that for the P2 site. These results are consistent with the function of AgrA as a response regulator with recognition sites in the promoter regions of RNAIII and the agr locus.

Research paper thumbnail of Production of pure protein and antibodies and development of immunoassays to detect Ara h 3 levels in peanut varieties

International Journal of Food Science & Technology, 2011

Peanuts are one of the most allergenic foods and are widespread in western food products. Therefo... more Peanuts are one of the most allergenic foods and are widespread in western food products. Therefore, there has been intense research into the allergic nature of the proteins involved. Ara h 3 is one of three immunodominant allergenic proteins. It is a 60-kDa protein, which forms following cleavage of the preprotein and association of the resultant 40-and 20-kDa subunits. The large subunit has been shown to harbour most of the reactive epitopes and has the protein fold likely responsible for its trypsin inhibitor activity. In this work, we have developed a method for the high-level expression and purification of recombinant Ara h 3 40-kDa subunit. Specific antibodies have been produced and applied to the secondary and tertiary screens of hundreds of peanut cultivars. Several of these cultivars were identified that have significantly reduced accumulation of Ara h 3.

Research paper thumbnail of Multicenter clinical evaluation of a multi-dose formulation of propofol in the dog

BMC Veterinary Research, 2013

Background: Propofol is a widely used injectable anesthetic agent for induction and short-term ma... more Background: Propofol is a widely used injectable anesthetic agent for induction and short-term maintenance in dogs. A multi-dose formulation of propofol (MDP) has been developed which includes 2% benzyl alcohol as a preservative. In order to document the use of the product under clinical conditions, MDP was tested in a prospective clinical trial conducted at six sites within the United States. One hundred thirty-eight healthy, client-owned dogs were assigned to one of six treatment groups based on premedicants (none, acepromazine/buprenorphine, midazolam/ buprenorphine, medetomidine/buprenorphine) and maintenance agents (MDP, inhaled anesthetic). Anesthesia was induced by the intravenous administration of MDP given to effect. Physiological indices including heart rate, respiratory rate and blood pressure were monitored prior to and during anesthesia induction, maintenance and recovery. Adverse events, defined for severity by pre-established limits of these physiological values, as well as side effects, defined as any observation outside the normal range, were noted. Results: The mean intubation dose was 7.6 ± 2.1 mg/kg for MDP alone and 4.7 ± 1.3, 4.0 ± 1.0 mg/kg and 3.2 ± 1.4 mg/kg when buprenorphine was used in combination with midazolam, acepromazine and medetomidine, respectively. Of the 32 adverse events, apnea (12 incidents), bradycardia (9 incidents) and hypotension (7 incidents) were most frequently recorded. Emesis, cyanosis and second degree heart block were each noted once and successfully resolved. The cause of a single death 2 days post-anesthesia was assessed as a surgical complication. Conclusions: MDP was found to be acceptable for use in healthy dogs for induction and short term maintenance of anesthesia when used alone and in combination with premedicants and inhaled anesthetics.

Research paper thumbnail of tRNA is the source of low-level trans-Zeatin production in Methylobacterium spp

Journal of bacteriology, 2002

Pink-pigmented facultatively methylotrophic bacteria (PPFMs), classified as Methylobacterium spp.... more Pink-pigmented facultatively methylotrophic bacteria (PPFMs), classified as Methylobacterium spp., are persistent colonizers of plant leaf surfaces. Reports of PPFM-plant dialogue led us to examine cytokinin production by PPFMs. Using immunoaffinity and high-performance liquid chromatography (HPLC) purification, we obtained 22 to 111 ng of trans-zeatin per liter from culture filtrates of four PPFM leaf isolates (from Arabidopsis, barley, maize, and soybean) and of a Methylobacterium extorquens type culture originally recovered as a soil isolate. We identified the zeatin isolated as the trans isomer by HPLC and by a radioimmunoassay in which monoclonal antibodies specific for trans-hydroxylated cytokinins were used. Smaller and variable amounts of trans-zeatin riboside were also recovered. trans-Zeatin was recovered from tRNA hydrolysates in addition to the culture filtrates, suggesting that secreted trans-zeatin resulted from tRNA turnover rather than from de novo synthesis. The product of the miaA gene is responsible for isopentenylation of a specific adenine in some tRNAs. To confirm that the secreted zeatin originated from tRNA, we mutated the miaA gene of M. extorquens by single exchange of an internal miaA fragment into the chromosomal gene. Mutant exconjugants, confirmed by PCR, did not contain zeatin in their tRNAs and did not secrete zeatin into the medium, findings which are consistent with the hypothesis that all zeatin is tRNA derived rather than synthesized de novo. In germination studies performed with heat-treated soybean seeds, cytokinin-null (miaA) mutants stimulated germination as well as wild-type bacteria. While cytokinin production may play a role in the plant-PPFM interaction, it is not responsible for stimulation of germination by PPFMs.

Research paper thumbnail of Phosphatase Expression by Chlorella Vulgaris (Chlorophyceae) is Mediated by Internal Phosphorus Levels and External pH

Journal of Phycology, 2002

This study was conducted to evaluate four species of algae (Chromulina freiburgensis Dofl., Chlor... more This study was conducted to evaluate four species of algae (Chromulina freiburgensis Dofl., Chlorella ellipsoidea Gerneck, C. vulgaris Beijerinck, and Chlamydomonas acidophila Negoro) isolated from an acid pit lake for their bioremediative potential. The Berkeley Pit Lake system, located in Butte, MT, USA is an aban-4

Research paper thumbnail of A randomized controlled field trial of a novel trimethoprim-sulfadiazine oral suspension for treatment of Streptococcus equi subsp zooepidemicus infection of the lower respiratory tract in horses

Journal of the American Veterinary Medical Association, Jan 15, 2015

Objective-To evaluate the effectiveness of a novel trimethoprim-sulfadiazine oral suspension for ... more Objective-To evaluate the effectiveness of a novel trimethoprim-sulfadiazine oral suspension for the treatment of naturally acquired Streptococcus equi subsp zooepidemicus infection in horses. Design-Randomized, controlled field trial. Animals-180 horses with S equi subsp zooepidemicus infection. Procedures-Horses with lower respiratory tract infections caused by S equi subsp zooepidemicus were treated with a new formulation of combined trimethoprim-sulfadiazine oral suspension at a dosage of 24 mg/kg (10.9 mg/lb) twice daily for 10 days (treatment group) or with an equivalent volume of saline (0.9% NaCl) solution (placebo group). Response to treatment, including clinical signs and fecal consistency scores, was assessed twice daily. Any adverse effects were recorded. The primary outcome variable was clinical response; the secondary outcome variable was eradication of S equi subsp zooepidemicus on study day 17 as determined by bacteriologic culture of repeated transtracheal-wash spec...

Research paper thumbnail of A Controlled Safety Study of Elevated Dosages of Trimethoprim Plus Sulfadiazine in Mature Horses

Journal of Equine Veterinary Science, 2014

A novel, proprietary, oral suspension of trimethoprim plus sulfadiazine (TMP/SDZ) had no serious ... more A novel, proprietary, oral suspension of trimethoprim plus sulfadiazine (TMP/SDZ) had no serious adverse effects on clinical, laboratory, or pathologic parameters of mature horses when administered at up to five times the intended combined dosage of 24 mg/kg twice daily for 30 consecutive days. Loose feces was the most common observation that was likely related to TMP/SDZ treatment, and the incidence of loose feces was greater in groups treated with higher dosages of TMP/SDZ. However, all episodes were self-limiting, and no horses were treated for loose feces.

Research paper thumbnail of Safety and efficacy of a new formulation of trimethoprim sulfadiazine in horses

Safety and efficacy of a new formulation of trimethoprim sulfadiazine in horses

Research paper thumbnail of tRNA is the source of low-level trans-zeatin production in Methylobacterium spp

Journal of bacteriology, 2002

Pink-pigmented facultatively methylotrophic bacteria (PPFMs), classified as Methylobacterium spp.... more Pink-pigmented facultatively methylotrophic bacteria (PPFMs), classified as Methylobacterium spp., are persistent colonizers of plant leaf surfaces. Reports of PPFM-plant dialogue led us to examine cytokinin production by PPFMs. Using immunoaffinity and high-performance liquid chromatography (HPLC) purification, we obtained 22 to 111 ng of trans-zeatin per liter from culture filtrates of four PPFM leaf isolates (from Arabidopsis, barley, maize, and soybean) and of a Methylobacterium extorquens type culture originally recovered as a soil isolate. We identified the zeatin isolated as the trans isomer by HPLC and by a radioimmunoassay in which monoclonal antibodies specific for trans-hydroxylated cytokinins were used. Smaller and variable amounts of trans-zeatin riboside were also recovered. trans-Zeatin was recovered from tRNA hydrolysates in addition to the culture filtrates, suggesting that secreted trans-zeatin resulted from tRNA turnover rather than from de novo synthesis. The pro...

Research paper thumbnail of Staphylococcus aureus AgrA Binding to the RNAIII-agr Regulatory Region

Journal of Bacteriology, 2004

The control of virulence gene expression in the human pathogen Staphylococcus aureus is under the... more The control of virulence gene expression in the human pathogen Staphylococcus aureus is under the partial control of the two-component quorum-sensing system encoded by genes of the agr locus. The product of the agrA gene has been shown by amino acid sequence similarity to be the putative response regulator; however, binding of AgrA to promoters under its control has not yet been demonstrated. In this study, we isolated and purified soluble AgrA by expression under osmotic shock conditions and ion-exchange chromatography. Purified AgrA showed high-affinity binding to the RNAIII-agr intergenic region by electrophoretic mobility shift assays. Binding was localized by DNase I protection assays to a pair of direct repeats in the P2 and P3 promoter regions of the agr locus. We found that this binding was enhanced by the addition of the small phosphoryl donor, acetyl phosphate. The difference in binding affinity between these two promoters was found to result from a 2-bp difference between the downstream direct repeats of the P2 and P3 sites. Mutation of these base pairs in the P3 site to match those found in the P2 site increased the affinity of AgrA for the P3 site relative to that for the P2 site. These results are consistent with the function of AgrA as a response regulator with recognition sites in the promoter regions of RNAIII and the agr locus.

Research paper thumbnail of Production of pure protein and antibodies and development of immunoassays to detect Ara h 3 levels in peanut varieties

International Journal of Food Science & Technology, 2011

Peanuts are one of the most allergenic foods and are widespread in western food products. Therefo... more Peanuts are one of the most allergenic foods and are widespread in western food products. Therefore, there has been intense research into the allergic nature of the proteins involved. Ara h 3 is one of three immunodominant allergenic proteins. It is a 60-kDa protein, which forms following cleavage of the preprotein and association of the resultant 40-and 20-kDa subunits. The large subunit has been shown to harbour most of the reactive epitopes and has the protein fold likely responsible for its trypsin inhibitor activity. In this work, we have developed a method for the high-level expression and purification of recombinant Ara h 3 40-kDa subunit. Specific antibodies have been produced and applied to the secondary and tertiary screens of hundreds of peanut cultivars. Several of these cultivars were identified that have significantly reduced accumulation of Ara h 3.

Research paper thumbnail of Multicenter clinical evaluation of a multi-dose formulation of propofol in the dog

BMC Veterinary Research, 2013

Background: Propofol is a widely used injectable anesthetic agent for induction and short-term ma... more Background: Propofol is a widely used injectable anesthetic agent for induction and short-term maintenance in dogs. A multi-dose formulation of propofol (MDP) has been developed which includes 2% benzyl alcohol as a preservative. In order to document the use of the product under clinical conditions, MDP was tested in a prospective clinical trial conducted at six sites within the United States. One hundred thirty-eight healthy, client-owned dogs were assigned to one of six treatment groups based on premedicants (none, acepromazine/buprenorphine, midazolam/ buprenorphine, medetomidine/buprenorphine) and maintenance agents (MDP, inhaled anesthetic). Anesthesia was induced by the intravenous administration of MDP given to effect. Physiological indices including heart rate, respiratory rate and blood pressure were monitored prior to and during anesthesia induction, maintenance and recovery. Adverse events, defined for severity by pre-established limits of these physiological values, as well as side effects, defined as any observation outside the normal range, were noted. Results: The mean intubation dose was 7.6 ± 2.1 mg/kg for MDP alone and 4.7 ± 1.3, 4.0 ± 1.0 mg/kg and 3.2 ± 1.4 mg/kg when buprenorphine was used in combination with midazolam, acepromazine and medetomidine, respectively. Of the 32 adverse events, apnea (12 incidents), bradycardia (9 incidents) and hypotension (7 incidents) were most frequently recorded. Emesis, cyanosis and second degree heart block were each noted once and successfully resolved. The cause of a single death 2 days post-anesthesia was assessed as a surgical complication. Conclusions: MDP was found to be acceptable for use in healthy dogs for induction and short term maintenance of anesthesia when used alone and in combination with premedicants and inhaled anesthetics.

Research paper thumbnail of tRNA is the source of low-level trans-Zeatin production in Methylobacterium spp

Journal of bacteriology, 2002

Pink-pigmented facultatively methylotrophic bacteria (PPFMs), classified as Methylobacterium spp.... more Pink-pigmented facultatively methylotrophic bacteria (PPFMs), classified as Methylobacterium spp., are persistent colonizers of plant leaf surfaces. Reports of PPFM-plant dialogue led us to examine cytokinin production by PPFMs. Using immunoaffinity and high-performance liquid chromatography (HPLC) purification, we obtained 22 to 111 ng of trans-zeatin per liter from culture filtrates of four PPFM leaf isolates (from Arabidopsis, barley, maize, and soybean) and of a Methylobacterium extorquens type culture originally recovered as a soil isolate. We identified the zeatin isolated as the trans isomer by HPLC and by a radioimmunoassay in which monoclonal antibodies specific for trans-hydroxylated cytokinins were used. Smaller and variable amounts of trans-zeatin riboside were also recovered. trans-Zeatin was recovered from tRNA hydrolysates in addition to the culture filtrates, suggesting that secreted trans-zeatin resulted from tRNA turnover rather than from de novo synthesis. The product of the miaA gene is responsible for isopentenylation of a specific adenine in some tRNAs. To confirm that the secreted zeatin originated from tRNA, we mutated the miaA gene of M. extorquens by single exchange of an internal miaA fragment into the chromosomal gene. Mutant exconjugants, confirmed by PCR, did not contain zeatin in their tRNAs and did not secrete zeatin into the medium, findings which are consistent with the hypothesis that all zeatin is tRNA derived rather than synthesized de novo. In germination studies performed with heat-treated soybean seeds, cytokinin-null (miaA) mutants stimulated germination as well as wild-type bacteria. While cytokinin production may play a role in the plant-PPFM interaction, it is not responsible for stimulation of germination by PPFMs.