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Results point out that the wzz gene is composed of wzy/rol genes and encodes the chain length deterrminant, wbcE gene represents wzy gene encoding the O-antigen polymerase, and galE/lse gene is the gne whose product is UDP-N-acetylglucosamine-4-epimerase.

Research points out that the O polysaccharide moiety of Burkholderia mallei lipopolysaccharide mediates the resistance to the bactericidal action of serum. The gene cluster of O polysaccharide biosynthesis and its structure are discussed.

Byline: Michael Heesen (1), Brunhilde Blomeke (2), Bernhard Schluter (3), Nicole Heussen (4), Rolf Rossaint (1), Dagmar Kunz (5) Keywords: CD14 antigen Genetics: gene polymorphism Molecular biology: real-time PCR Fluorescence-activated cell sorting Cytokines: tumor necrosis factor-[alpha] Abstract: Objective: CD14 is a receptor for endotoxin and binds components of Gram-positive and Gram-negative bacteria. CD14-bearing monocytes respond to stimulation with the increased synthesis and release of cytokines. The recently described --260 C[right arrow]T promoter polymorphism of the CD14 gene has been found to be related to a risk of myocardial infarction. This study evaluated the role of this polymorphism in the expression of monocyte and soluble CD14. Moreover, the effect of the CD14 --260 genotypes for the ex vivo TNF-[alpha] response to endotoxin was analyzed in whole blood. Patients and participants: Ninety-five healthy blood donors were studied. Measurements and results: CD14 --260 genotyping was performed by means of a real-time PCR with fluorescence labeled hybridization probes. CD14 expression on human monocytes (mCD14) was assessed by fluorescence-activated cell sorting analysis with anti-CD14 monoclonal antibodies. Plasma levels of soluble CD14 (sCD14) were measured by ELISA. The TNF-[alpha] synthesis was determined by chemiluminescence in whole blood after endotoxin stimulation. There were no differences in mCD14 density, sCD14 levels, or the tumor necrosis factor-[alpha] concentrations between individuals with the three different CD14 --260 genotypes CC, CT, and TT. Conclusions: The CD14 --260 polymorphism does not affect the CD14 expression of unstimulated circulating monocytes or soluble CD14 plasma levels. Author Affiliation: (1) Department of Anesthesiology and Intensive Care Medicine, University Hospital of Aachen, Pauwelsstrasse 30, 52057 Aachen, Germany (2) Department of Dermatology, University Hospital of Aachen, Pauwelsstrasse 30, 52057 Aachen, Germany (3) Department of Clinical Chemistry, University Hospital of Munster, Albert-Schweitzer-Strasse 33, 48149 Munster, Germany (4) Department of Statistics and Biometry, University Hospital of Aachen, Pauwelsstrasse 30, 52057 Aachen, Germany (5) Department of Clinical Chemistry and Pathobiochemistry, University Hospital of Aachen, Pauwelsstrasse 30, 52057 Aachen, Germany Article History: Received Date: 08/12/2000 Accepted Date: 21/08/2001 Article note: Final revision received: 21 August 2001 Electronic Publication

Research documents occurrence of the flagellum-independent mobility of Vibrio cholerae and Escherichia coli on surfaces of media composed of agarose. Data indicate that this phenotype is an active translocation process negatively affected by mutations in the lipopolysaccharide biosynthesis pathway.

Results show that a recombinat cosmid carries the waa gene cluster mediating Klebsiella pneumoniae C3 core lipopolysaccharide biosynthesis as revealed by genetic complementation analyses of waa mutants.

Results demonstrate that lpsB gene, contained in a 5.5 kilobase chromosomal region of Sinorhizobium meliloti, encodes mannosyltransferase enzyme involved in the lipopolysaccharide core synthesis. Data further point out that the gene is associated with nodulation and ntrogen fixation.

Ligand blotting and vesicle binding research results indicate that domain II loops play a determinant role in Cry toxin specificity within Heliothis virescens, a cotton crop pest. The Cry toxin used was obtained from Bacillus thuringiensis; implications for agricultural bacteriology are also discussed.

Results demonstrate that L-Rha is the receptor of the lipopolysaccharide core involved in the O polysaccharide attachment in Pseudomonas aeruginosa as shown by gene mutation studies.

Results demonstrate that Escherichia coli O55 and O157 experience an antigenic shift and transform into pathogenic types via cotransfer and recombination processes involving rfb and gnd loci from heterologous bacterial source.

Byline: S. Schroeder (1), M. Reck (1), L. Eric Lehmann (1), M. Book (1), A. Hoeft (1), F. Stuber (1) Keywords: Key words Inducible heat-shock-protein 70; PstI gene polymorphism; Reverse transcription-polymerase chain reaction; Endotoxin; Heat-shock-protein 70 mRNA expression; Clinical outcome Abstract: Objective: To determine whether the human leukocyte antigen linked biallelic heat-shock protein 70--2 (HSP70--2) gene polymorphism is associated with variable HSP70--2 messenger RNA expression. Design: Prospective observational study in consecutive healthy blood donors. Setting: Department of Anesthesiology, laboratory for molecular biology in a university hospital. Participants: 24 healthy blood donors. Interventions: None. Measurements and results: We studied the functional implication of the HSP70--2 (G/A) PstI gene polymorphism in 24 healthy, white blood donors with various HSP70--2 (G/A) genotypes by analyzing the endotoxin-inducible HSP70--2 mRNA expression by means of the reverse transcription--polymerase chain reaction. HSP70 expression was expressed semiquantitatively by calculating the ratio of HSP70--2 mRNA and the constitutively expressed glutaraldehyde 3-phosphate dehydrogenase mRNA. No significant differences in HSP70--2 mRNA expression after lipopolysaccharide (from Salmonella minnesota Re 595) stimulation were detected in individuals homozygous for the allele A (0.68, range 0.38--1, n = 10), in individuals homozygous for the allele G (0.79, range 0.42--1.1, n = 8), and in heterozygotes (HSP70--2 G/A 0.52, range 0.4--0.67, n = 6 p > 0.05). Conclusions: The PstI polymorphism of the endotoxin-inducible HSP70--2 gene is not associated with variable HSP70--2 mRNA expression ex vivo. This finding is in accordance with the observation that HSP70--2 genotypes do not affect clinical outcome in human systemic inflammation. Author Affiliation: (1) Klinik und Poliklinik fur Anasthesiologie und Spezielle Intensivmedizin, Friedrich-Wilhelms-Universitat Bonn, Sigmund-Freud-Strasse 25, 53 105 Bonn, Germany e-mail: schroed@mailer.meb.uni-bonn.de Tel.: + 49-2 28-2 87 60 18 Fax: + 49-2 28-2 87 51 57, DE Article note: Received: 5 October 1999 Final revision received: 18 April 2000 Accepted: 26 April 2000

Results indicate that the lipooligosaccharide galactosyltransferase II gene in Haemophilus ducreyi codes for a 33,400 dalton protein which bears homology to glycosyltransferases of other bacteria. Data show that the biosynthetic defect lgtB in this gene does not influence serum-sensitive phenotype.

Lipopolysaccharide (LPS) is a potent stimulator of monocytes and macrophages, causing secretion of tumor necrosis factor [Alpha] (TNF-[Alpha]) and other inflammatory mediators. Given the deleterious effects to the host of TNF-[Alpha], it has been postulated that TNF-[Alpha] gene expression must be tightly regulated. The nature of the nuclear factor(s) that control TNF-[Alpha] gene transcription in humans remains obscure, although NF-[Kappa]B has been suggested. Our previous studies pertaining to macrophage response to LPS identified a novel DNA-binding domain located from -550 to -487 in the human TNF-[Alpha] promoter that contains transcriptional activity, but lacks any known NF-[Kappa]B-binding sites. We have used this DNA fragment to isolate and purify a 60-kDa protein binding to this fragment and obtained its amino-terminal sequence, which was used to design degenerate probes to screen a cDNA library from THP-1 cells. A novel cDNA clone (1.8 kb) was isolated and fully sequenced. Characterization of this cDNA clone revealed that its induction was dependent on LPS activation of THP-1 cells; hence, the name LPS-induced TNF-alpha factor (LITAF). Inhibition of LITAF mRNA expression in THP-1 cells resulted in a reduction of TNF-[Alpha] transcripts. In addition, high level of expression of LITAF mRNA was observed predominantly in the placenta, peripheral blood leukocytes, lymph nodes, and the spleen. Finally, chromosomal localization using fluorescence in situ hybridization revealed that LITAF mapped to chromosome 16p12-16p13.3. Together, these findings suggest that LITAF plays an important role in the activation of the human TNF-[Alpha] gene and proposes a new mechanism to control TNF-[Alpha] gene expression.

The cloning of an alpha-endotoxin from Bacillus thuringiensis subsp. jegathesan is presented. The toxin can be used as biological control agent against mosquitos. The cloned endotoxin does not react to the antibody raised against the Cyt1Aa protein. The amino acid sequence is 31% identical to the Cyt1Aa protein and 66% identical to the Cyt2Aa protein. Bacillus thuringiensis is a gram-positive bacterium that is used in controlling insects harmful to man and plants.

Genes implicated in core biosynthesis of Pseudomonas aeruginosa were cloned and characterized to determine the function of lipopolysaccharide (LPS) in its virulence. A P. aeruginosa PAO1 library was electrotransformed into Salmonella waaC and waaF mutants to isolate the homologous heptosyltransferase I and II genes of P. aeruginosa. Two plasmids, pCOREc1 and pCOREc2, which restored smooth LPS production in the waaC mutant, were isolated. Plasmid pCOREf1 was able to complement the Salmonella waaF mutant.

Analysis of the 4.2-kb region of the vacuolating cytotoxin alleles (vacA) of Helicobacter pylori (Hp) indicated the presence of an open reading frame (ORF) that is related to the virulence-associated protein D allele (vapD). Sequence analysis of the ORF indicated the presence of the vapD gene in 61.3% of Hp strains. However, the vapD gene is not a part of the pathogenicity island in Hp and the presence of the gene was not related to vacA alleles.

The mechanisms that regulate lipopolysaccharide (LPS)-mediated induction of the serum amyloid A (SAA) gene in THP-1 monocyte/macrophage cells were characterized via DNA-binding electromobility shift assay. Analysis of THP-1 monocyte/macrophage cells expressing SAA genes indicated the role of LPS in increasing the levels of SAA messenger RNA and inducing the DNA binding activity of the protein. Furthermore, LPS-mediated SAA gene induction in the monocyte/macrophage cells was regulated by the SAS element within the -280 and -226 bases.

Substitutions or deletions of domain II loop residues of Bacillus thuringiensis [Delta]-endotoxin CryIAb were constructed using site-directed mutagenesis techniques to investigate their functional roles in receptor binding and toxicity toward gypsy moth (Lymantria dispar). Substitution of loop 2 residue N372 with Ala or Gly (N372A, N372G) increased the toxicity against gypsy moth larvae 8-fold and enhanced binding affinity to gypsy moth midgut brush border membrane vesicles (BBMV) [approximately equal to]4-fold. Deletion of N372 (D3), however, substantially reduced toxicity (>21 times) as well as binding affinity, suggesting that residue N372 is involved in receptor binding. Interestingly, a triple mutant, DF-1 (N372A, A282G and L283S), has a 36-fold increase in toxicity to gypsy moth neonates compared with wild-type toxin. The enhanced activity of DF-1 was correlated with higher binding affinity (18-fold) and binding site concentrations. Dissociation binding assays suggested that the off-rate of the BBMV-bound mutant toxins was similar to that of the wild type. However, DF-1 toxin bound 4 times more than the wild-type and N372A toxins, and it was directly correlated with binding affinity and potency. Protein blots of gypsy moth BBMV probed with labeled N372A, DF-1, and CryIAb toxins recognized a common 210-kDa protein, indicating that the increased activity of the mutants was not caused by binding to additional receptor(s). The improved binding affinity of N372A and DF-1 suggest that a shorter side chain at these loops may fit the toxin more efficiently to the binding pockets. These results offer an excellent model system for engineering [Delta]-endotoxins with higher potency and wider spectra of target pests by improving receptor binding interactions.

A 2-kb region that complements the Tn5-derived lipopolysaccharide (LPS) rough mutant Rhizobium leguminosarum RU301 was sequenced. Two open reading frames (ORFs) were observed. The first ORF (lpcA) is homologous to a family of bacterial sugar transferases involved in LPS core tetrasaccharide biosynthesis. ORF2 (lpcB), in which Tn5 transposed, has no significant homology to any DNA in the GenBank-EMBL databases. The 3-deoxy-D-manno-2-octulosonic acid (Kdo) residue which normally attaches the core tetrasaccharide to the O chain was missing, indicating that lpcB may encode a CMP-Kdo:LPS Kdo transferase.

The rfb gene clusters of the Klebsiella species are necessary for the synthesis of the D-galactan I of O antigens. The O1 polysaccharides of the bacteria are composed of two repeat units of D-galactan I which is altered by serotype specificity. DNA homology indicated that the rfb clusters of Klebsiella serotype O8 and O9 belong to different clonal groups. The Kliebsiella species also exhibits clonal diversity with functionally identical rfb loci.

Mammals continually confront microbes at mucosal surfaces. A current model suggests that epithelial cells contribute to defense at these sites, in part through the production of broad-spectrum antibiotic peptides. Previous studies have shown that invertebrates can mount a host defense response characterized by the induction in epithelial cells of a variety of antibiotic proteins and peptides when they are challenged with microorganisms, bacterial cell wall/membrane components, or traumatic injury [Boman, H.G. & Hultmark, D. (1987) Annu. Rev. Microbiol. 41, 103-126]. However, factors that govern the expression of similar defense molecules in mammalian epithelial cells are poorly understood. Here, a 13-fold induction of the endogenous gene encoding tracheal antimicrobial peptide was found to characterize a host response of tracheal epithelial cells (TECs) exposed to bacterial lipopolysaccharide (LPS). Northern blot data indicated that TECs express CD14, a well-characterized LPS-binding protein known to mediate many LPS responses. A monoclonal antibody to CD14 blocked the observed tracheal antimicrobial peptide induction by LPS under serum-free conditions. Together the data support that CD14 of epithelial cell origin mediates the LPS induction of an antibiotic peptide gene in TECs, providing evidence for the active participation of epithelial cells in the host's local defense response to bacteria. Furthermore, the data allude to a conservation of this host response in evolution and suggest that a similar inducible pathway of host defense is prevalent at mucosal surfaces of mammals.

Bacillus thuringiensis AB88 expresses a CryV-type protein (CGCryV) in the early stationary phase. This is a protein secreted into the media. Examination of the deduced protein sequence from the cryV-type gene indicated the presence of an N-terminal domain that possibly functions as a signal peptide. This CGCryV protein is the first reported case of a delta-endotoxin being a secreted protein.

A hybrid strain of Escherichia coli with a deep rough rfa defect was developed which indicated that the K40 (group IB) K(sub LPS) antigen exists mostly as long chains. The typical chain length distribution of the K40 antigen was changed by introducing multicopy rol, indicating that the K40 group IB K antigen is equivalent to a Rol-dependent O antigen. Findings revealed that group IA K(sub LPS) is also synthesized by a Rol-dependent pathway and that short oligosaccharide K(sub LPS) results from the absence of Rol activity in these strains.

Mutations in the isn gene are responsible for the I69 phenotype in Haemophilus influenzae, in which the lipopoplysaccharide is composed of only lipid A and a single phosphorylated 2-keto-3-deoxyoctulosonic acid molecule. The gene can be found between loci with a high degree of homology to the dpp and art loci from Escherichia coli. Results suggest the isn plays a role in the biosynthesis of lopopolysaccharides.

The rfb locus in Escherichia coli is involved in the biosynthesis of O-specific lipopolysaccharide (LPS). Localization of the E. coli gene encoding gluconate-6-phosphate dehydrogenase established the 3' endpoint of the rfb locus. Further sequence analysis of this region revealed the presence of two genes coding for phosphomannomutase and GDP-mannose pyrophosphorylase, enzymes involved in the conversion of mannose-6-phosphate to GDP-mannose, a nucleotide sugar precursor of the O7-specific repeating unit of LPS.

The rol (regulator of O length) genes of Escherichia coli O75 and Salmonella typhimurium LT2 were sequenced and compared. Transcription of the E. coli and S. typhimurium rol genes proceeded counter to the histidine operon. The deduced amino acid sequences revealed 72 percent homology. The S. typhimurium Rol protein was able to regulate expression of the O-polysaccharide antigen of E. coli in a complementation study. The regulation of size distribution of the O-antigen chain appeared to be independent of the subunit structure.

The lipopolysaccharide (LPS) O antigens of Actinobacillus pleuropneumoniae capsular serotypes 1, 9 and 11 were studied using chemical and spectroscopic methods. The serotype 1 LPS O-PS had branched tetrasaccharide and linear, deaminated trisaccharide repeating units while serotype 11 LPS O-PS had identical tetrasaccharide repeating unit. The serotype 9 LPS O-PS, on the other hand, had only about 25 and 75 percent of the same tetrasaccharide and trisaccharide repeating units, respectively.

Mini-Mu vector was utilized in cloning the domain of the genes encoding the lipopolysaccaride core of the outer membrane of Escherichia coli. Genetic analysis of this region using recombination and complementation techniques showed 17 open reading frames and three different operons transcribed in opposite directions. Protein mapping was done and a homology between the K-12 strain of E. coli and Salmonella typhimurium was observed.

The first three genes involved in the synthesis of the lipopolysaccharide core in Escherichia coli were identified and characterized. These genes were preliminarily identified by their complementation to rfa mutants in Salmonella typhimurium. Sequence data revealed three open reading frames corresponding rfaQ, rfaP and rfaG. In vitro synthesized proteins showed them to be basic and soluble. Sequence comparisons showed some homologies between RfaQ and RfaC and RfaF, which are involved in the synthesis of the heptose region of the core. Sequence homologies were also observed between RfaB, RfaG and RfaK, which are speculated to be hexose transferases.

Cloning of the gmhA gene from Escherichia coli showed that it shares a significant homology to the product of the gene HI#1181 of Haemophilus influenzae. Expression of this gene showed that it encodes the enzyme phosphoheptose isomerase which catalyzes the first step in the formation of ADP-L-glycero-D-mannoheptose. Mutations to this gene cause alterations in the permeability of the E. coli outer membrane to hydrophobic compounds. Sequence analysis of the gmhA gene product showed that it belongs to a novel family of phosphosugar aldo-keto isomerases.

The authors have studied insecticidal protein Vip3A secreted by Bacillus thuringiensis during the growth. The protein's mode of action has been investigated via the use of Cry1Ab protein for comparison, and the results have demonstrated that Vip3A shares the same sigma-endotoxin with Cry1Ab but utilizes different molecular targets and forms distinct ion channels.

We tested the hypothesis that endotoxemia and fasting are associated with increased gut apoptotic activity, gut permeability, and inflammation in a distant organ. Fed or fasted CD-1 mice were studied 6 h after intraperitoneal injection of either saline (sham) or endotoxin (4 mg/kg of 0111:B4 Escherichia coli lipopolysaccharide). We found that endotoxin increased gut caspase-3 and -6 activity by 4.9 [+ or -] 0.6- and 4.5 [+ or -] 0.5-fold, respectively (P [is less than] 0.001), and increased terminal deoxynucleotidyltransferasemediated dUTP nick-end labeling (TUNEL) staining of mucosal cells (P [is less than] 0.05). Feeding decreased caspase-3 activity by 40% (P [is less than] 0.05) and decreased endotoxin-induced TUNEL staining (P [is less than] 0.05). Endotoxin increased gut poly(ADP-ribose) polymerase activity by 15% (P [is less than] 0.05). Endotoxin increased gut permeability by 44% (P [is less than] 0.05), an effect reduced 36% by feeding (P [is less than] 0.05). Similarly, endotoxin increased pulmonary neutrophil infiltration (6.0 [+ or -] 1.0-fold, P [is less than] 0.001) and increased lung interleukin (IL)-6 (5.9 [+ or -] 0. l-fold, P [is less than] 0.001) and macrophage inflammatory protein (MIP)-2 expression (290 [+ or -] 40-fold, P [is less than] 0.001), whereas feeding decreased this effect by 43% for neutrophils, 40% for IL-6 (P [is less than] 0.05), and 35% for MIP-2 (P [is less than] 0.05). Thus endotoxin increases gut apoptotic activity, gut permeability, and pulmonary inflammation. Enteral feeding may decrease the distant organ inflammation by reducing gut apoptosis, thereby maintaining gut mucosal function during endotoxemia. sepsis; endotoxin; caspase; poly(ADP-ribose) polymerase

Source and Description of Primers. Degenerate primers (set A) were designed from published human and murine lipopolysaccharide-binding protein (LBP) exon 4 and exon 5 gene sequences (Genbank accession no. AF105067 and X00347, respectively). These primers were used to amplify porcine LBP genomic DNA. Porcine-specific primers (set B) were designed based on the pig DNA sequence generated using set A primers. Primer Sequences. Set A primers were as follows: forward 5'-GTC AAG GGC ATC ASC ATT TC and reverse 5'-TCT GGT TGT GGA AGA GRT TCA3'. Set B primers were as follows: forward 5'-TGC AGC AGC CAC ATC GAC 3' and reverse 5'-AGA GGC ACA GTG AGG CAC AG. Method of Detection. The PCR with set A primers was performed in 10-[micro]L PCR reactions including 0.38 U Taq Gold DNA polymerase (Perkin-Elmer, Branchburg, NJ), 1x PCR buffer, 2.0 mM Mg[Cl.sub.2], 0.2 mM of each dNTP, 3 pmol of each primer, and 12.5 ng genomic DNA. Thermocycling was carried out in a Robocycler (Stratagene, La Jolla, CA) or an MJ Research PTC-200 instrument (Watertown, MA) and included initial denaturation at 95 [degrees] C for 2 min, followed by 40 cycles at 95 [degrees] C for 45 s, 56 [degrees] C for 1 min, 72 [degrees] C for 1.5 min, and a final extension step at 72 [degrees] C for 5 min. The PCR product generated with set A primers was directly sequenced using dye terminators and an ABI 377 sequencer (Perkin-Elmer, Foster City, CA) at the Iowa State University DNA Sequencing and Synthesis Facility. The set B primers were used to amplify an approximately 1,200-bp fragment. This product was used for physical and linkage mapping. Set B PCR was performed using the MJ Research PTC-100 instrument and conditions as for set A except that the Mg[Cl.sub.2] concentration was 1.5 mM and TaqDNA polymerase (Promega, Madison, WI) was used. A PCR-RFLP marker was identified by using the Fnu4HI enzyme, and this PCR-RFLP was used for conducting linkage mapping and estimating allele frequencies in different populations of pigs. In the PCR-RFLP assays, 4-[micro]L PCR products were digested with 10 U Fnu4HI in 1x buffer solution. After digestion at 37 [degrees] C for 8 h, digested products were loaded in a 2.5% agarose gel containing ethidium bromide, electrophoresed, and photographed. Sequencing and Polymorphisms. The PCR product (approximately 1,300 bp in length) amplified with set A primers was sequenced from both directions. Because a polythymidine repeat region exists within the fragment that proved difficult to accurately sequence, we obtained only a 323-bp sequence with the exon 4 end and a 656-bp sequence with the exon 5 end and submitted these to GenBank (accession no. AF252875 and AF252876). A total of 127 bp out of the 979 bp total available sequence data shared 78.0% identity with sections of exon 4 (102bp) and exon 5 (25bp) of the human LBP gene at the nucleotide level. Comparison of the 979-bp sequence from Yorkshire and Meishan breeds revealed four single nucleotide polymorphisms, one of which occurred in the coding region with no amino acid change. Two single nucleotide polymorphisms GC to TT substitution in noncoding region at position 567 and 568 of AF252876 affecting an Fnu4HI restriction site were found to segregate in the PiGMaP families. The PCR-RFLP (Fnu4HI) with set B products included two allelic fragments of sizes 295 bp and 225 bp for alleles 1 and 2, respectively (Figure 1). Inheritance Pattern. The segregation of the PCR-RFLP (Fnu4HI) in the PiGMaP families (Archibald et al., 1995) was consistent with autosomal Mendelian inheritance. Allele Frequencies. Eighty-four unrelated DNA samples from six pig breeds and two European wild boars as well as a commercial population were amplified with set B primers and genotyped with Fnu4HI digestion. Allele 2 was observed with a frequency of 100% in Hampshire (n = 6), Landrace (n = 8), Large White (n = 11), Wild boar (n = 2), and a commercial population based on Large White and Landrace (n = 24). Allele 2 frequencies were estimated at 94.4% in Yorkshire (n = 9), 81.3% in Duroc (n = 8), and 34.4% in Meishan (n = 16). Chromosomal Location. To physically map LBP, the set B primers and a pig/rodent somatic cell hybrid panel comprising 27 cell lines (Yerle et al., 1996) was used. The results showed that LBP is located on pig chromosome 17 with 98% probability and in region (1/ 2q21)-q23 with 100% concordance, with less than 0.5 % error. The PCR data analysis was performed as described (http://www.toulouse.inra.fr/lgc/pig/hybrid/ chromo17/chromo17.htm). Linkage mapping was performed by using CRI-MAP (Green et al., 1990) analysis of PCR-RFLP (Fnu4HI) genotypes of the three-generation PiGMaP families (Archibald et al. 1995). The linkage results showed that the porcine LBP was linked to genes/markers previously mapped to pig chromosome 17, confirming the physical mapping. Markers linked to LBP with two-point recombination frequencies and LOD scores in parentheses were as follows: S0204 (0.13, 5.07), S0359 (0.18, 6.87), and Sw1031 (0.19, 6.51). Multipoint analysis using data for loci in this region indicated that LBP is most likely located between S0204 and S0359 (overall [log.sub.10]-1ikelihood = -235.71), rather than between S0359 and SW1031 (overall [log.sub.10]-likelihood = -237.21). Comments. Lipopolysaccharide-binding protein (LBP) is an acute-phase protein that strongly modulates the response to endotoxins (Schumann et al., 1990). Increased LBP decreases endotoxin activity and also limits septic shock due to Gram-negative bacterial infection (Lamping et al., 1998). In the present study, LBP was mapped as a candidate gene for immunogenetic traits in pigs. Literature Cited Archibald, A., C. Haley, J. Brown, et al. 1995. The PiGMaP Consortium Linkage Map of the Pig (Sus scrofa). Mamm. Genome 6:157-175. Green, P., K. Falls, and S. Crooks. 1990. Documentation for CRI-MAP. version 2.4. Washington University School of Medicine, St. Louis, MO. Lamping, N., R. Dettmer, N. W. J. Schroder, D. Pfeil, W. Hallatschek, R. Burger, and R. R. Schumann. 1998. LPS-binding protein protects mice from septic shock caused by LPS or gram-negative bacteria. J. Clin. Investig. 101:2065-2071. Schumann, R. R., S. R. Leong, G. W. Flaggs, P. W. Gray, S. D. Wright, J. C. Mathison, P. S. Tobias, and R. J. Ulevitch. 1990. Structure and function of lipopolysaccharide binding protein. Science (Wash D.C.) 249:1429-1431. Yerle, M., G. Echard, A. Robic, A. Mairal, C. Dubut-Fontana, J. Riquet, P. Pinton, D. Milan, Y. Lahbib-Mansis, and J. Gellin. 1996. A somatic hybrid panel for pig regional gene mapping characterized by molecular cytogenetics. Cytogenet. Cell. Genet. 73:194-202. Key Words: Binding Proteins, Gene Mapping, Genetic Polymorphism, Lipopolysaccharides, Pigs

The widespread planting of corn genetically modified to produce Bacillus thuringiensis endotoxin has led to speculation that pollen from these fields might adversely affect nearby nontarget lepidopterans. A previous study of Bt corn engineered with Monsanto event 810 failed to detect an effect of pollen exposure on the black swallowtail, Papilio polyxenes, in either the field or the laboratory. Here, we report results of a field study investigating the impact of exposure to pollen from a Bt corn hybrid containing Novartis event 176 on two species of Lepidoptera, black swallowtails and monarch butterflies, Danaus plexippus. Nearly half of the 600 monarch larvae died within the first 24 h; this and subsequent mortality was not associated with proximity to Bt corn and may have been due in part to predation. Survivorship of black swallowtails was much higher than that of the monarchs and was also independent of proximity to the transgenic corn. However, despite five rainfall events that removed much of the pollen from the leaves of their host plants during the experiment, we observed a significant reduction in growth rates of black swallowtail larvae that was likely caused by pollen exposure. These results suggest that Bt corn incorporating event 176 can have adverse sublethal effects on black swallowtails in the field and underscore the importance of event selection in reducing environmental impacts of transgenic plants.