Production of 2-Ketogluconic Acid by Serratia marcescens

Production of 2-ketogluconic acid from glucose by fermentation with Serratia marcescens NRRL B-486 was studied in 20-liter stainless-steel fermentors. Conditions for 2-ketogluconic acid production included the following: glucose-salt medium, aeration rate of 0.75 volumes per volume per minute, agitation rate of 400 rev/min, temperature of 30 C, CaCO₃ to neutralize the acid formed, and a 5% (v/v) inoculum. Foaming was controlled with an antifoam agent added at intervals during the fermentation. When 120 g per liter of glucose were supplied, 95 to 100% yields of 2-ketogluconic acid were obtained in 16 hr. Larger amounts of glucose could be used in the fermentation provided that the carbohydrate was fed continuously. Continuous feeding of glucose to a total amount of 180 g per liter gave 95 to 100% yields of 2-ketogluconic acid in 24 hr; feeding glucose to a total amount of 240 g per liter gave 85 to 90% yields in 32 to 40 hr.     

Allescheria boydii and Aspergillus fumigatus Skin Test Antigens

May, Lewis K. (Woman's Medical College of Pennsylvania, Philadelphia), Ralph A. Knight, and H. William Harris. Allescheria boydii and Aspergillus fumigatus skin test antigens. J. Bacteriol. 91:2155-2157. 1966.-Protein and polysaccharide fractions were extracted from culture filtrates of Allescheria boydii and Aspergillus fumigatus by the methods of Seibert and of Heidelberger, and injected intradermally into guinea pigs previously infected with these fungi. The diameter of erythema and induration was determined at 8, 24, and 48 hr. The protein and polysaccharide antigens yielded specific skin reactions in homologously infected guinea pigs. Erythema appeared at 8 hr with both the protein and polysaccharide antigens. At this time, the polysaccharide skin tests showed erythema and a central blanched wheal. A similar wheal was not observed with the protein. The erythema of the polysaccharide reaction began fading at 24 hr, whereas the protein reaction remained unchanged through 48 hr with both antigens. In guinea pigs, the area of erythema was more constant and thus easier to measure than was induration.     

Relationships between abdominal and diaphragmatic volume displacements

We investigated the relationship between the volumes displaced by the diaphragm and the abdominal wall during spontaneous breathing in supine anesthetized dogs. Diaphragmatic volume displacement (Vdi) was calculated from measurements taken from anteroposterior fluoroscopic images employing a previously described geometric model. The volume displacement of the abdominal wall (Vabd) was measured with a calibrated Respitrace. Shortening of single diaphragm muscle bundles in costal and crural regions was measured as the distance between radiopaque beads sutured to the peritoneal surface of the muscle. We found that Vdi always exceeded Vabd, but Vabd/Vdi was larger in animals in which the abdominal wall was more compliant. In this preparation, Vdi is better correlated with costal than with crural shortening. Vabd did not correlate with either costal or crural shortening. We infer that the difference between Vdi and Vabd reflects the volume displacement of the lower rib cage caused by diaphragm contraction. This volume difference was tightly correlated with costal shortening. We conclude from these data that coupling between Vdi and Vabd is influenced by the relative compliances of the chest wall and abdomen. Shortening of regions of the diaphragm may have variable relationships to the measured volume displacement, but costal shortening is intimately related to expansion of the lower rib cage.     

Rearrangement of VHa1-encoding Ig gene segment to the a2 chromosome in an a1/a2 heterozygous rabbit. Evidence for trans recombination

VDJ genes were cloned from leukemic B cells of an a1/a2 heterozygous Emu-cmyc transgenic rabbit. Restriction mapping and nucleotide sequence analysis indicated that one clone, 5C3, had a VHa1-encoding gene segment functionally rearranged to a JH gene segment from the a2 chromosome. This VDJ gene may be the result of a trans recombination between a VH gene on the a1 chromosome and a JH gene segment on the a2 chromosome or, it may be the result of a cis recombination if the a2 chromosome contains VHa1-encoding gene segments.     

Identification of sites within gp41 that serve as targets for antibody-dependent cellular cytotoxicity by using human monoclonal antibodies.

In an effort to determine the functional activity of anti-HIV-1 human mAb and to define the epitopes against which they are directed, supernatants from 10 EBV-transformed lymphoblastoid cell lines producing mAb to HIV were tested. Five clones producing mAb to gp41 and five producing mAb to p24 were identified. The anti-HIV-1 human mAb were tested in neutralization and cell fusion assays in the form of cell culture supernatants at concentrations ranging from 1.7 to 22.0 micrograms/ml. None of the human mAb were found either to inhibit HIV-1-(IIIB or RF) associated cell fusion or to neutralize HIV-1 (IIIB) infection of AA5 cells. All human mAb were additionally tested in 6 h 51Cr release assays for their ability to direct HIV-1 specific antibody-dependent cellular cytotoxicity (ADCC). For ADCC assays, PBMC were isolated from healthy seronegative donors and used as effector cells. HIV-1 infected (IIIB, RF, and MN) CEM.NKR cells as well as CEM.NKR cells with purified gp120 adsorbed onto their surface served as targets. None of the anti-p24 mAb mediated ADCC. In contrast, three of the anti-gp41 mAb were able to direct a significant level of ADCC against each of the infected targets, but as expected, failed to lyse gp120 adsorbed cells. To define the specific epitopes against which the anti-gp41 mAb were directed, seven small peptides homologous to regions within the extracellular domain of gp41 were synthesized. Using RIA, two of the mAb could be mapped. The most effective ADCC-directing human mAb bound to a peptide comprising amino acids 644-663, whereas the least effective ADCC directing anti-gp41 human mAb bound to a region within the immunodominant portion of gp41 outlined by amino acids 579-604. Together, these results for the first time assign a functional activity to human mAb directed at specific regions within gp41 by demonstrating that areas within this molecule can serve as targets for ADCC.