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1.  Identification, structure, and characterization of an exopolysaccharide produced by Histophilus somni during biofilm formation 
BMC Microbiology  2011;11:186.
Histophilus somni, a gram-negative coccobacillus, is an obligate inhabitant of bovine and ovine mucosal surfaces, and an opportunistic pathogen responsible for respiratory disease and other systemic infections in cattle and sheep. Capsules are important virulence factors for many pathogenic bacteria, but a capsule has not been identified on H. somni. However, H. somni does form a biofilm in vitro and in vivo, and the biofilm matrix of most bacteria consists of a polysaccharide.
Following incubation of H. somni under growth-restricting stress conditions, such as during anaerobiosis, stationary phase, or in hypertonic salt, a polysaccharide could be isolated from washed cells or culture supernatant. The polysaccharide was present in large amounts in broth culture sediment after H. somni was grown under low oxygen tension for 4-5 days (conditions favorable to biofilm formation), but not from planktonic cells during log phase growth. Immuno-transmission electron microscopy showed that the polysaccharide was not closely associated with the cell surface, and was of heterogeneous high molecular size by gel electrophoresis, indicating it was an exopolysaccharide (EPS). The EPS was a branched mannose polymer containing some galactose, as determined by structural analysis. The mannose-specific Moringa M lectin and antibodies to the EPS bound to the biofilm matrix, demonstrating that the EPS was a component of the biofilm. The addition of N-acetylneuraminic acid to the growth medium resulted in sialylation of the EPS, and increased biofilm formation. Real-time quantitative reverse transcription-polymerase chain reaction analyses indicated that genes previously identified in a putative polysaccharide locus were upregulated when the bacteria were grown under conditions favorable to a biofilm, compared to planktonic cells.
H. somni is capable of producing a branching, mannose-galactose EPS polymer under growth conditions favorable to the biofilm phase of growth, and the EPS is a component of the biofilm matrix. The EPS can be sialylated in strains with sialyltransferase activity, resulting in enhanced density of the biofilm, and suggesting that EPS and biofilm formation may be important to persistence in the bovine host. The EPS may be critical to virulence if the biofilm state is required for H. somni to persist in systemic sites.
PMCID: PMC3224263  PMID: 21854629
2.  Chemical modification of L-glutamine to alpha-amino glutarimide on autoclaving facilitates Agrobacterium infection of host and non-host plants: A new use of a known compound 
BMC Chemical Biology  2011;11:1.
Accidental autoclaving of L-glutamine was found to facilitate the Agrobacterium infection of a non host plant like tea in an earlier study. In the present communication, we elucidate the structural changes in L-glutamine due to autoclaving and also confirm the role of heat transformed L-glutamine in Agrobacterium mediated genetic transformation of host/non host plants.
When autoclaved at 121°C and 15 psi for 20 or 40 min, L-glutamine was structurally modified into 5-oxo proline and 3-amino glutarimide (α-amino glutarimide), respectively. Of the two autoclaved products, only α-amino glutarimide facilitated Agrobacterium infection of a number of resistant to susceptible plants. However, the compound did not have any vir gene inducing property.
We report a one pot autoclave process for the synthesis of 5-oxo proline and α-amino glutarimide from L-glutamine. Xenobiotic detoxifying property of α-amino glutarimide is also proposed.
PMCID: PMC3130638  PMID: 21624145
3.  Characterization and Comparison of Biofilm Development by Pathogenic and Commensal Isolates of Histophilus somni▿  
Journal of Bacteriology  2007;189(22):8179-8185.
Histophilus somni (Haemophilus somnus) is an obligate inhabitant of the mucosal surfaces of bovines and sheep and an opportunistic pathogen responsible for respiratory disease, meningoencephalitis, myocarditis, arthritis, and other systemic infections. The identification of an exopolysaccharide produced by H. somni prompted us to evaluate whether the bacterium was capable of forming a biofilm. After growth in polyvinyl chloride wells a biofilm was formed by all strains examined, although most isolates from systemic sites produced more biofilm than commensal isolates from the prepuce. Biofilms of pneumonia isolate strain 2336 and commensal isolate strain 129Pt were grown in flow cells, followed by analysis by confocal laser scanning microscopy and scanning electron microscopy. Both strains formed biofilms that went through stages of attachment, growth, maturation, and detachment. However, strain 2336 produced a mature biofilm that consisted of thick, homogenous mound-shaped microcolonies encased in an amorphous extracellular matrix with profound water channels. In contrast, strain 129Pt formed a biofilm of cell clusters that were tower-shaped or distinct filamentous structures intertwined with each other by strands of extracellular matrix. The biofilm of strain 2336 had a mass and thickness that was 5- to 10-fold greater than that of strain 129Pt and covered 75 to 82% of the surface area, whereas the biofilm of strain 129Pt covered 35 to 40% of the surface area. Since H. somni is an obligate inhabitant of the bovine and ovine host, the formation of a biofilm may be crucial to its persistence in vivo, and our in vitro evidence suggests that formation of a more robust biofilm may provide a selective advantage for strains that cause systemic disease.
PMCID: PMC2168709  PMID: 17644581

Results 1-3 (3)