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Alpha-, beta II- and gamma-subspecies of protein kinase C localized in the monkey hippocampus: pre- and post-synaptic localization of gamma-subspecies.

Protein kinase C (PKC) has attracted wide attention as a key enzyme for the expression of long-term potentiation in the hippocampus, a basic model for memory. It is of interest to study the detailed localization of PKC subspecies in the monkey hippocampus. We used immunocytochemistry to examine the localization of PKC subspecies in the hippocampus of the monkey, Macaca mulatta. Subspecies of PKC in the monkey could be separated by hydroxyapatite chromatography and the elution profile proved to be similar to that of the rat. Antibodies against each alpha, beta II and gamma-subspecies of the rat specifically reacted with the respective subspecies of monkey PKC. The alpha-, beta II- and gamma-subspecies were distinctly distributed in the hippocampus. The beta I-subspecies was not evident in the hippocampus. While both the alpha- and gamma-subspecies immunoreactive pyramidal cells were distributed throughout the hippocampus (CA1-CA3), the beta II-subspecies immunoreactive cells were scattered only in the CA1 region. The gamma-subspecies was found in granule cells and dendrites in the dentate gyrus, in mossy fibers and in their terminals in the CA3 region. The alpha-subspecies was also present in granule cells and in the dendrites but not in the mossy fibers. Glial cells did not stain with any of the antibodies used. Electron microscopy clearly showed that the gamma-subspecies was localized in both presynaptic terminals and post-synaptic dendrites. These observations suggest that subspecies of PKC in the monkey hippocampus may be involved in distinct functions and that the gamma-subspecies of PKC may act pre- and post-synaptically in pyramidal cells of the hippocampus.

Amino Acid Sequence↗

Mitochondrial sequence diversity within a subspecies of savanna monkeys (Cercopithecus aethiops) is similar to that between subspecies.

Cercopithecus aethiops can be classified into four subspecies by morphology and by geographic distribution. However, the phylogenetic relationship between these subspecies is unclear. We previously found five distinct haplogroups of mitochondrial DNA (mtDNA) in the subspecies C. aethiops aethiops at the restriction fragment length polymorphism (RFLP) level, and found that those haplogroups are parapatrically distributed in their habitat. To determine the relationship between subspeciation and haplogroup formation in a subspecies, we compared mtDNA control region and 12S rRNA gene sequences (approximately 700 bp) in C. a. aethiops, two other subspecies of C. aethiops, and two species of Cercopithecus: The diversity between haplogroups in C. a. aethiops was almost the same as that between subspecies. This similar level of diversification between and within haplogroups may explain why a previously obtained mtDNA tree did not show monophyletic branching according to subspecies.

Animals↗

Rat brain glial cells in primary culture and subculture contain the delta, epsilon and zeta subspecies of protein kinase C as well as the conventional subspecies.

We raised polyclonal antibodies against the C-terminal peptides of protein kinase C (PkC) subspecies alpha, beta 1, beta 2, gamma, delta, epsilon, and zeta and checked their specificity against brain extracts using Western immunoblot analysis. With equal amounts of protein applied to gels PkC subspecies beta 1, delta, epsilon and zeta were detected in primary cultures of mixed glial cells: bands for the alpha and beta 2 subspecies were less prominent. PkC gamma was not detected in primary glial cultures. The epsilon and zeta subspecies of PkC were detected in subcultures of type 1 astrocytes with weaker bands for the alpha, beta 1 and beta 2 subspecies. Blots of O-2A-lineage glia contained PkCs delta and zeta as prominent bands: the alpha, beta 1 and epsilon subspecies were also present. All PkC subspecies including PkC gamma were detected in C6 glioma cells.

Amino Acid Sequence↗

Salmonella virulence plasmid. Modular acquisition of the spv virulence region by an F-plasmid in Salmonella enterica subspecies I and insertion into the chromosome of subspecies II, IIIa, IV and VII isolates.

The spv operon is common to all Salmonella virulence plasmids. DNA hybridization analysis indicates that the spv region is limited in distribution to serovars of Salmonella enterica subspecies I, II, IIIa, IV, and VII and is absent from Salmonella bongori isolates. Among strains of subspecies II, IIIa, and VII, all isolates examined contained sequences that hybridized with the spv region. However, among isolates of subspecies I, DNA sequences capable of hybridizing with the spv region were found in some isolates of certain serovars. Furthermore, in isolates of subspecies I, the virulence plasmid was found in the same set of isolates as an F-related plasmid, as determined by the presence of the spv region of the virulence plasmid and the finO, traD, and repA sequences of the F-plasmid. The concordance of the virulence plasmid and all three F-plasmid sequences in subspecies I serovar Choleraesuis, Paratyphi, and Typhimurium is most easily explained if the spv region is carried in an F-related plasmid in these isolates. In contrast, among S. enterica subspecies II, IIIa, IV, and VII, the isolates that contain spv sequences did not hybridize with an F-related plasmid or any other identifiable plasmid. With the use of pulse-field gel electrophoresis, the spv region in subspecies II, IIIa, and VII was found to be encoded on the chromosome. Analysis of the phylogenetic distribution of spv among Salmonella isolates and comparative nucleotide sequence analysis of spvA and spvC suggests that the spv region was acquired very recently, after speciation of the salmonellae.

Base Sequence↗

Quantitation of human apolipoprotein C-III and its subspecie by radioimmunoassay and analytical isoelectric focusing: abnormal plasma triglyceride-rich lipoprotein apolipoprotein C-III subspecie concentrations in hypertriglyceridemia.

A specific, accurate, and sensitive double antibody radioimmunoassay for measuring human apolipoprotein (apo) C-III has been developed. Anti-apoC-III(1) developed in rabbits cross-reacted completely with apoC-III subspecies. Analytical isoelectric focusing of delipidated triglyceride-rich lipoproteins (TRL) was done to assess the percentage of total apoC-III mass comprised by apoC-III(0), C-III(1), and C-III(2), and the data were used to compute the absolute plasma TRL apoC-III subspecie concentration. Total plasma apoC-III was 11.1 +/- 0.9 mg/dl (mean +/- SEM) in 29 normolipidemic healthy subjects; 21.3 +/- 4.9, 27.5 +/- 2.2, and 53.6 +/- 7 mg/dl in 3, 16, and 13 patients with primary types III, IV, and V hyperlipoproteinemia, respectively, and significantly (P < 0.01) higher than normal. Total plasma triglycerides (TG) correlated positively with total plasma apoC-III (r = 0.88; P = 0.0001) and TRL apoc-III (r = 0.88; P = 0.0001). Progressive hypertriglyceridemia was associated with a rise in the percent of total apoC-III in TRL isolated at d < 1.006 g/ml (r = 0.78; P < 0.0001; n = 43) and a reciprocal decline in the TRL-free plasma fraction (d > 1.006 g/ml). ApoC-III comprised significantly more of HDL(2) than HDL(3) protein (7.3 +/- 0.2 versus 1.6 +/- 0.2%, respectively, P < 0.01). HDL(2) and HDL(3) isolated from patients with type IV hyperlipoproteinemia had subnormal apoC-III as percent of total protein (2.4 +/- 0.5 and 0.6 +/- 0.1, respectively). Total plasma TG correlated negatively with i) apoC-III as percent of total HDL protein (r = -0.67; P = 0.002, n = 20); ii) apoC-III as percent of total HDL(2) protein (r = -0.52; P = 0.019); and iii) apoC-III as percent of total HDL(3) protein (r = -0.72; P = 0.0004). Plasma TRL apoC-III subspecie concentrations were significantly higher in the three hypertriglyceridemic groups (primary types III, IV, and V) compared to normals. TRL apoC-III(0) levels in patients with type IV and V were comparable (2.4 +/- 0.3 and 2.2 +/- 0.6 mg/dl, respectively). However, TRL apoC-III(1) and C-III(2) in patients with type V hyperlipoproteinemia were significantly higher (P < 0.01) than in patients with types IV or III hyperlipoproteinemia. Total plasma TG correlated positively with TRL apoC-III(0) (r = 0.56; P = 0.0004), TRL apoC-III(1) (r = 0.82; P = 0.0001) and TRL apoC-III(2) (r = 0.76; P = 0.0001). The slope of regression line relating total plasma TG with TRL apoC-III(1) was significantly steeper (P < 0.0001) than that for apoC-III(0). Thus, for a given interval of plasma TG, the change in concentration of TRL apoC-III(1) was much greater than that in TRL apoC-III(0). The development of the RIA and its combined use with analytical isoelectric focusing thus allows quantitation of this important glycopeptide and its subspecies in human plasma and its subfractions. Because apoC-III inhibits not only tissue lipoprotein lipase but also the hepatic uptake of triglyceride-rich lipoproteins and remnants, the data support the possibility that an abnormal metabolism of apoC-III subspecies may be linked pathogenetically to elevated plasma TG levels.-Kashyap, M. L., L. S. Srivastava, B. A. Hynd, P. S. Gartside, and G. Perisutti. Quantitation of human apolipoprotein C-III and its subspecies by radioimmunoassay and analytical isoelectric focusing: abnormal plasma triglyceride-rich lipoprotein apolipoprotein C-III subspecie concentrations in hypertriglyceridemia.

Apolipoprotein C-III↗

Identification of a subspecies-specific capsular antigen from Bacteroides melaninogenicus subspecies asaccharolyticus by immunofluorescence and electron microscopy.

An indirect fluorescent antibody test was developed with the use of hyperimmune rabbit antiserum to a purified capsular polysaccharide of Bacteroides melaninogenicus subspecies asaccharolyticus. All of 23 strains of B. melaninogenicus subspecies asaccharolyticus were fluorescence-positive in this test. All 11 strains of Bacteroides melaninogenicus subspecies intermedius tested and three strains of Bacteroides melaninogenicus subspecies melaninogenicus were fluorescence-negative. Thirty-one strains of other bacterial species were also fluorescence-negative. The indirect fluorescent antibody test demonstrated the presence of a subspecies-specific capsular antigen from B. melaninogenicus subspecies asaccharolyticus. The capsular antigen was further demonstrated by electron microscopy with ruthenium red, a polysaccharide-staining material.

Animals↗

Differentiation of Lactococcus lactis subspecies lactis and subspecies cremoris strains by their adaptive response to stresses.

Lactococcus lactis subspecies lactis (L. lactis ssp. lactis) and Lactococcus lactis subspecies cremoris (L. lactis ssp. cremoris) were investigated in respect to their response to acid, bile-salt and freezing stresses. First, the sublethal and lethal levels of each stress were determined for both subspecies. For acid stress, the levels were pH 4.5 and 2.5, respectively, for L. lactis ssp. lactis, and pH 5.0 and 3.0, respectively, for L. lactis ssp. cremoris. For bile-salt stress, the levels were 0.03 and 0.1%, respectively, for L. lactis ssp. lactis, and 0.01 and 0.04%, respectively, for L. lactis ssp. cremoris. For freezing stress, 10 degrees C was used as the sublethal temperature and -20 degrees C was used as the lethal temperature for both subspecies. To evaluate the effect of each stress at log phase, a log-phase culture was challenged directly with the appropriate lethal level (control culture) and a second log-phase culture was pre-exposed to the appropriate sublethal level prior to testing survival under normally lethal conditions (test culture). Some, if not most, of the cells were killed in the control cultures for all three stresses. However, in the test cultures, the viability was significantly improved for all of the L. lactis ssp. lactis strains tested, but not for the L. lactis ssp. cremoris strains. It appears, therefore, that L. lactis ssp. lactis is capable of displaying adaptive response to stresses, whereas L. lactis ssp. cremoris seems to lack this phenotype or the response is much weaker in this subspecies. The effect of each stress on stationary-phase cultures was also investigated. Unlike the log-phase cultures, the stationary-phase cultures of both subspecies, challenged directly with the lethal levels, were highly resistant to each of the three stresses tested.

Adaptation, Physiological↗

Evaluation of PCR-based methods for discrimination of Francisella species and subspecies and development of a specific PCR that distinguishes the two major subspecies of Francisella tularensis.

Previous studies have demonstrated that the four subspecies of the human pathogen Francisella tularensis, despite showing marked variations in their virulence for mammals and originating from different regions in the Northern Hemisphere, display a very close phylogenetic relationship. This property has hampered the development of generally applicable typing methods. To overcome this problem, we evaluated the use of PCR for discrimination of the subspecies using various forms of long arbitrary primers or primers specific for repetitive extragenic palindromic sequences (REP) or enterobacterial repetitive intragenic consensus (ERIC) sequences. Patterns generated by use of REP, ERIC, or long arbitrary primers allowed differentiation at the species level and of the four subspecies of F. tularensis. With each of these three methods, similar or identical clustering of strains was found, and groups of strains of different geographical origins or differing in virulence showed distinct patterns. The discriminatory indices of the methods varied from 0.57 to 0.65; thus, the patterns were not sufficiently discriminatory to distinguish individual strains. The sequence of a fragment generated by amplification with an arbitrary primer was determined, and a region showing interstrain heterogeneity was identified. Specific primers were designed, and a PCR was developed that distinguished strains of F. tularensis subsp. holarctica from strains of other F. tularensis subspecies, including strains of the highly virulent F. tularensis subsp. tularensis. Notably, one European isolate showed the genetic pattern typical of the highly virulent F. tularensis subsp. tularensis, generally believed to exist only in North America. It is proposed that a combination of the specific PCR together with one method generating subspecies-specific patterns is suitable as a rapid and relatively simple strategy for discrimination of Francisella species and subspecies.

DNA Primers↗

Conformational differences in human apolipoprotein B-100 among subspecies of low density lipoproteins (LDL). Association of altered proteolytic accessibility with decreased receptor binding of LDL subspecies from hypertriglyceridemic subjects.

We asked at what point in the metabolic cascade of very low density lipoproteins (VLDL) to low density lipoproteins (LDL) the accessibility of proteolytic cleavage sites in B-100 changes, and we evaluated the effect of hypertriglyceridemia on the proteolytic accessibility, secondary structure, and receptor-binding affinity of B-100 in LDL subspecies of varying density. Limited proteolysis with Staphylococcus aureus V8 protease and cathepsin D identified the density (about 1.033 g/ml) between two LDL subspecies, designated LDL-1 and -2, as the transition point during VLDL metabolism of both normolipidemic (N-) and hypertriglyceridemic (HTG-) subjects at which accessibility to protease attack changed in three peptide regions of B-100. Hypertriglyceridemia greatly altered proteolytic accessibility of B-100 in the denser LDL subspecies. Specifically, B-100 in HTG-LDL exposed more cleavage sites than in N-LDL, including two novel sites, approximately 120 and approximately 130 kDa from the NH2 terminus in the small and dense subspecies (designated LDL-4, -4.5 or -5, d = 1.048-1.062 g/ml). Analysis of circular dichroic spectra indicated no difference in helical content between B-100 in N- and HTG-LDL but showed a greater content of beta-structure in HTG-LDL. Binding affinity for the LDL receptor of human fibroblasts decreased markedly with increasing density among HTG-LDL subspecies (by approximately 50% for LDL-4.5 or -5). We conclude that the changes in proteolytic accessibility observed between LDL-1 and -2 and in LDL-4, -4.5, or -5 indicate significant differences in local conformation of B-100 at specific peptide regions. The association of exposure of more cleavage sites, especially novel sites in the NH2-terminal regions, with greatly decreased receptor-binding affinity in LDL-4.5 or -5 suggests that altered local conformation in B-100 apart from the putative receptor-binding domain might affect interaction with the receptor.

Apolipoprotein B-100↗

Virulence characterization of a strain of Salmonella enterica subspecies houten (subspecies IV) with chromosomal integrated Salmonella plasmid virulence (spv) genes.

The Salmonella plasmid virulence genes (spv) are commonly found on plasmids contained in a small number of serotypes of Salmonella belonging to subspecies I, where they are important for survival within macrophages and the establishment of successful systemic infection. However, in this study, spv genes were detected by the polymerase chain reaction in the chromosome of a plasmid-free strain of S. IV 16:z4, z32:- (Salmonella subspecies IV). The full range of spv genes (spvR, spvA, spvB, spvC and spvD) was demonstrated, but a 216-bp deletion, accompanied by an insertion of 59-bp cryptic DNA, was present in spvA. S. IV 16:z4, z32:- was avirulent in mice and did not become virulent with the introduction of a fully functionally serotype-associated virulence plasmid (SAP) from S. typhimurium. By use of an spvRAB'-chloramphenicol acetyl transferase fusion gene, it was demonstrated that S. IV 16:z4, z32:- did not express the spv genes. Salmonella subspecies IV is monophasic, and in phylogenetic analyses it clusters distantly to Salmonella subspecies I, where all the serotypes that normally carry SAPs are found. The mechanisms by which spv genes have been transferred to this serotype remain unknown.

Animals↗

Comparison of the fibronectin-binding protein FNE from Streptococcus equi subspecies equi with FNZ from S. equi subspecies zooepidemicus reveals a major and conserved difference.

The gene fnz from Streptococcus equi subspecies zooepidemicus encodes a cell surface protein that binds fibronectin (Fn). Fifty tested isolates of S. equi subspecies equi all contain DNA sequences with similarity to fnz. This work describes the cloning and sequencing of a gene, designated fne, with similarity to fnz from two S. equi subspecies equi isolates. The DNA sequences were found to be identical in the two strains, and sequence comparison of the fne and fnz genes revealed only minor differences. However, one base deletion was found in the middle of the fne gene and eight base pairs downstream of the altered reading frame there is a stop codon. An Fn-binding protein was purified from the growth medium of a subspecies equi culture. Determination of the NH(2)-terminal amino acid sequence and molecular mass, as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, revealed that the purified protein is the gene product of the 5'-terminal half of fne. Fn-binding activity has earlier only been found in the COOH-terminal half of FNZ. By the use of a purified recombinant protein containing the NH(2) half of FNZ, we provide here evidence that this half of the protein also harbors an Fn-binding domain.

Adhesins, Bacterial↗

Ultrastructure of Bacteroides species: Bacteroides asaccharolyticus, Bacteroides fragilis, Bacteroides melaninogenicus subspecies melaninogenicus, and B. melaninogenicus subspecies intermedius.

Representative strains of two subspecies of Bacteroides melaninogenicus (subspecies melaninogenicus and subspecies intermedius) and Bacteroides asaccharolyticus as well as B. asaccharolyticus strain 536B isolated from a human perirectal abscess and Bacteroides fragilis ATCC 25285 were examined by glutaraldehyde-osmium fixation, ruthenium red fixation and staining, and thorium hydroxyde staining as well as by the physical preparative techniques of critical point drying--transmission electron microscopy (CPD--TEM) and scanning electron microscopy (SEM). All strains, with the exception of B. fragilis 25285, possessed an electron-dense material external to their outer membranes. Ruthenium red staining further revealed a layer, external to the surface of the outer membrane, that was distinct for each species examined. Thorium hydroxide, as well as CPD--TEM and SEM, showed the cells to be interconnected by thin fibers that not only connected adjacent cells but also traversed several microns to connect cell aggregates.

Bacteroides↗

Differential down-regulation of protein kinase C subspecies in normal human melanocytes: possible involvement of the zeta subspecies in growth regulation.

Normal human melanocytes are often grown in vitro in the continuous presence of 12-O-tetradecanoylphorbol-13-acetate (TPA) for growth in vitro. The expression of protein kinase C (PKC) subspecies, which are the major cellular receptors for phorbol esters, was examined in melanocytes after long-term treatment with TPA to investigate the role of PKC subspecies in TPA-dependent cell growth. The PKC enzyme activity detected in quiescent melanocytes was almost completely depleted in cells after incubation with 85 nM TPA for 48 h. Immunoblot analysis indicated that, among the PKC subspecies alpha, beta, delta, epsilon, and zeta expressed in quiescent cells, alpha-, beta-, delta-, and epsilon-PKC were significantly down-regulated, whereas zeta-PKC remained at detectable levels in TPA-treated cells. TPA did not significantly affect the expression or subcellular distribution of zeta-PKC in melanocytes. Immunoprecipitation assay revealed that the enzyme activity of zeta-PKC was increased in both the cytosol and particulate cell fractions, but the increase was much greater in the latter. The activation of zeta-PKC lasted for 24 to 48 h after the addition of TPA; thereafter, zeta-PKC activity returned to basal levels. DNA synthesis was shown to change concomitantly with the activation of zeta-PKC in TPA-treated cells. These results indicate that TPA induces not only the down-regulation of alpha-, beta-, delta-, and epsilon-PKC, but also long-term activation of zeta-PKC in melanocytes, and that activation of zeta-PKC parallels the growth of normal human melanocytes.

Cell Division↗

Mycobacterium fortuitum subspecies acetamidolyticum, a new subspecies of Mycobacterium fortuitum.

Mycobacterium fortuitum subspecies acetamidolyticum is a new subspecies of M. fortuitum and has an intermediate growth rate. It is a nonphotochromogenic mycobacterium. It does not utilize glutamate but utilizes acetamide as a simultaneous nitrogen and carbon source. It is able to utilize acetate, malate, pyruvate, fumarate, glucose, fructose, and n-propanol as the sole sources of carbon in the presence of ammoniacal nitrogen, but does not utilize them in the presence of glutamate-nitrogen. It is easily differentiated from all rapidly growing mycobacteria by its inability to utilize glutamate as a simultaneous nitrogen and carbon source, and from all slowly growing mycobacteria by its capacity to utilize acetamide as a simultaneous nitrogen and carbon source. Its mycolic acid pattern is different from that of M. fortuitum. However, its deoxyribonucleic acid showed 94% relatedness with that of M. fortuitum. In view of the above findings, it has been designated as a new subspecies of M. fortuitum. The organism was isolated from sputum of a 56-year-old patient with lung disease and is considered to be a lung pathogen. The type strain is ATCC 35931 (NCH E11620).

Base Composition↗

The ability of Mycoplasma mycoides subspecies mycoides and closely related strains from goats and sheep to immunize mice against subspecies capri.

Small colony (SC) strains of Mycoplasma mycoides subsp. mycoides from contagious bovine pleuropneumonia (CBPP) and from goats were compared with large colony (LC) strains of so-called M. mycoides subsp. mycoides from goats and sheep by means of a cross-protection test in which mice were challenged with M. mycoides subsp. capri. Of 13 LC strains, all gave partial cross-protection, and 11 were shown to be more closely related than four SC strains to subspecies capri. In a further experiment, six SC strains--three from CBPP and three from goats--all gave weak partial cross-protection against subspecies capri.

Animals↗

Activation of macrophages and interference with CD4+ T-cell stimulation by Mycobacterium avium subspecies paratuberculosis and Mycobacterium avium subspecies avium.

Mycobacterium avium subspecies paratuberculosis (M. ptb) and M. avium subspecies avium (M. avium) are closely related but exhibit significant differences in their interaction with the host immune system. The macrophage line, J774, was infected with M. ptb and M. avium and analysed for cytokine production and stimulatory capacity towards antigen-specific CD4+ T cells. Under all conditions J774 cells were activated to produce proinflammatory cytokines. No influence on the expression of major histocompatibility complex (MHC) class II, intracellular adhesion molecule-1 (ICAM-1), B7.1, B7.2 or CD40 was found. However, the antigen-specific stimulatory capacity of J774 cells for a CD4+ T-cell line was significantly inhibited after infection with M. ptb, but not with M. avium. When a T-cell hybridoma expressing a T-cell receptor identical to that of the T-cell line was used, this inhibition was not observed, suggesting that costimulation which is essential for the CD4+ T-cell line is influenced by the pathogenic bacterium M. ptb.

Animals↗

Note: identification of Lactobacillus delbrueckii subspecies bulgaricus and subspecies lactis dairy isolates by amplified rDNA restriction analysis.

Amplification and restriction analysis of the 16S rRNA gene (PCR-ARDRA) was used to identify Lactobacillus delbrueckii dairy isolates and to assign them to its three subspecies (delbrueckii, lactis and bulgaricus). PCR-ARDRA allowed confirmation of predictions from the DNA database regarding the restriction mapping of the 16S sequence to be verified in practice. PCR-ARDRA was shown to be a reliable and rapid method for identifying Lact. delbrueckii isolates at the subspecies level and for differentiating this species from Lact. helveticus and Lact. acidophilus.

Blotting, Southern↗

Gene expression profiling of monocyte-derived macrophages following infection with Mycobacterium avium subspecies avium and Mycobacterium avium subspecies paratuberculosis.

Mycobacterium avium subspecies paratuberculosis (MAP) and Mycobacterium avium subspecies avium (MAA) represent two closely related intracellular bacteria with vastly different associated pathologies. MAA can cause severe respiratory infections in immune compromised humans but is nonpathogenic in ruminants and is more readily controlled by the bovine immune system than MAP. MAP causes a fatal wasting syndrome in ruminants, typified by granulomatous enteritis localized in the small intestine. MAP has also been cited as a potential cause of human Crohn's disease. We used a bovine immune-specific microarray (BOTL-5) to compare the response of mature bovine monocyte-derived macrophages (MDM cells) to MAP and MAA. Statistical analysis of microarray data revealed 21 genes not appreciably expressed in resting MDM cells that were activated following infection with either MAA or MAP. Further analysis revealed 144 genes differentially expressed in MDM cells following infection with MAA and 99 genes differentially expressed following infection with MAP. Of these genes, 37 were affected by both types of mycobacteria, with three being affected in opposite directions. Over 41% of the differentially expressed genes in MAA and MAP infected MDM cells were members of, regulated by, or regulators of the MAPK pathways. Expression of selected genes was validated by quantitative real-time reverse transcriptase PCR and in several key genes (i.e., IL-2 receptor, tissue inhibitor of matrix metalloproteinases-1, and Fas-ligand) MAA was found to be a stronger activating factor than MAP. These gene expression patterns were correlated with prolonged activation of p38 MAPK and ERK1/2 by MAA, relative to MAP.

Animals↗