Lung lesions in SCID-bo and SCID-bg mice after intratracheal inoculation with wild-type or leucotoxin-deficient mutant strains of Mannheimia haemolytica serotype 1.

The purpose of this study was to investigate SCID-bg mice engrafted with bovine haematolymphoid tissues (SCID-bo) as a model for studying bovine Mannheimia haemolytica serotype 1- induced pneumonia, in which leucotoxin (LKT) plays a major role. In experiment A, SCID-bo and SCID-bg mice were inoculated intratracheally with either (1) phosphate-buffered saline (PBS), (2) M. haemolytica wild-type strain 89010807N ("LKT(+)WT"), (3) a M. haemolytica leucotoxin-deficient mutant of strain 89010807N ("LKT(-)mutant"), or (4) the M. haemolytica wild-type Oklahoma strain. Mice were killed for examination at intervals between 20 and 44h after inoculation. Lung lesions consisted of thickened alveolar septa and neutrophil and macrophage infiltrates in the bronchioles and alveoli. Lung lesion scores in the SCID-bo mice inoculated with LKT(+)WT or LKT(-) mutant were significantly (P<0.05) greater than those of the PBS control group, but the two bacterial strains produced results that did not differ significantly. M. haemolytica was isolated from lung, liver and spleen after inoculation but less frequently as time progressed. In experiment B, SCID-bg mice were inoculated intratracheally with live LKT(+)WT or formalin-killed LKT(+)WT and killed 24, 48 or 96 h later. Lung lesions were histologically similar to those observed in experiment A; however, there were no significant differences in the lung lesion scores between groups. It was concluded that the lesions seen in this study were probably not due to LKT, and that the SCID-bo mouse does not provide a good rodent model for bovine pneumonia.

Analysis of tandem, multiple genes encoding 30-kDa membrane proteins in Pasteurella haemolytica A1.

A number of outer membrane proteins (OMPs), including a 30-kDa protein, may be important in eliciting immunity to Pasteurella haemolytica A1, the causative agent of bovine pneumonic pasteurellosis. To better understand the nature of the 30-kDa antigen, several genes encoding this protein were sequenced. Sequence analysis revealed that three separate genes encoding similar, yet distinct, versions of the 30-kDa protein are tandemly arranged on the P. haemolytica A1 chromosome. The genes appear to be transcribed from a single promoter. The deduced amino acid sequences of the proteins encoded by these genes are similar to a 28-kDa inner membrane lipoprotein of Escherichia coli and a 28-kDa membrane protein which may contribute to the virulence of Haemophilus influenzae type b strains.

Construction of isogenic mutants of Pasteurella haemolytica by allelic replacement.

We describe methods for the mutagenesis of cloned Pasteurella haemolytica (Gram-) genes and for the construction of P. haemolytica mutants by allelic exchange. We used these methods to construct isogenic mutants of P. haemolytica which no longer synthesize three membrane lipoproteins (Lpp). A single genetic locus, consisting of three tandemly arranged genes encoding 28-30-kDa membrane Lpp, was replaced with a mutated locus which carries the beta-lactamase-encoding ApR gene from a 4.2-kb P. haemolytica plasmid. The inactivated locus was introduced into P. haemolytica by electroporation of a plasmid which carries the mutated locus, but is incapable of replicating in P. haemolytica. Southern and Western blot analyses indicate that the wild-type locus was replaced by the mutated locus through a double-crossover recombination event and that the membrane Lpp were no longer produced by the mutant strain. These methods should be useful in constructing mutant loci which can be used to analyze the roles for various P. haemolytica proteins in the pathogenesis of bovine pneumonic pasteurellosis.

Analysis of two genes encoding heat-labile toxins and located on a single Ent plasmid from Escherichia coli.

A clinical isolate of Escherichia coli harbored two copies of the heat-labile toxin (LT)-encoding gene (elt) on a 157-kb plasmid. The arrangement of the gene copies is different from the cholera toxin-encoding gene duplication described for some strains of Vibrio cholerae. The nucleotide sequences of the elt alleles are not identical (differing by 2 bp) and the duplicated region flanking the alleles extends 314 bp on one side and 1122 bp on the other side of each copy. Different partial copies of IS600 were identified 280 bp 3' to the stop codon of each allele. Partial and complete copies of other IS were also identified near the elt alleles. The data suggest that the regions surrounding the genes are hot spots for IS which would account for the observed heterogeneity in DNA flanking elt genes.

On metaphoric representation.

The article discusses claims that conceptual structure is in some part metaphorical, as identified by verbal metaphors like LOVE IS A JOURNEY. Two main interpretations of this view are discussed. In the first, a target domain is not explicitly represented but is instead understood through reference to a different domain. For example, rather than a detailed concept of love per se, one could make reference to the concept of a journey. In the second interpretation, there is a separate representation of love, but the content of that representation is influenced by the metaphor such that the love concept takes on the same structure as the journey concept. It is argued that the first interpretation is not fully coherent. The second interpretation is a possible theory of mental representation, but the article raises a number of empirical and theoretical problems for it. It is concluded that many of the data cited as evidence for metaphoric representations can be accounted for by structural similarity between domains.

Thematic relations in adults' concepts.

Concepts can be organized by their members' similarities, forming a kind (e.g., animal), or by their external relations within scenes or events (e.g., cake and candles). This latter type of relation, known as the thematic relation, is frequently found to be the basis of children's but not adults' classification. However, 10 experiments found that when thematic relations are meaningful and salient, they have significant influence on adults' category construction (sorting), inductive reasoning, and verification of category membership. The authors conclude that concepts function closely with knowledge of scenes and events and that this knowledge has a role in adults' conceptual representations.

Development of a non-radioactive DNA probe and in situ hybridization for detection of Anaplasma marginale in ticks and cattle.

A non-radioactive DNA probe was developed for detection of Anaplasma marginale in ticks and cattle. The probe was labeled with digoxigenin 11-dUTP by polymerase chain reaction. The probe was tested on bovine blood and was found to be a sensitive and specific detection method for A. marginale in cattle. The DNA probe was then adapted for in situ hybridization (ISH) of A. marginale in Dermacentor andersoni and D. variabilis ticks infected either as nymphs or adults. One-half of each tick was studied with ISH while the other half was examined with light and electron microscopy. In male ticks infected as adults, tick gut cells first became infected with A. marginale while ticks fed on an infected calf, and they remained infected as they transmission fed on a second, susceptible calf. At the onset of transmission feeding, salivary glands became infected with A. marginale. During transmission feeding infection was also observed in interstitial, reproductive, skeletal muscle, fat body and Malpighian tubule tissue, resulting in a generalized A. marginale infection. When adult ticks that acquired infection as nymphs were examined with ISH and microscopy, gut tissues of both D. andersoni and D. variabilis became infected with A. marginale. However, salivary gland infection was seen only in D. variabilis, even though both species of ticks transmitted A. marginale to susceptible calves. A. marginale was not seen with ISH or microscopy in hemocytes collected from both species of ticks and, thus, hemocytes do not appear to play a role in the development of A. marginale in ticks.

Genetic and immunological analyses of a 38 kDa surface-exposed lipoprotein of Pasteurella haemolytica A1.

Pasteurella haemolytica serotype A1 is the bacterial pathogen most frequently isolated from the lungs of cattle with bovine respiratory disease. As part of a study to characterize P. haemolytica antigens which are important in eliciting resistance to pneumonic pasteurellosis, we have cloned and sequenced the gene encoding a 38 kDa lipoprotein, Lpp38. The deduced amino acid sequence of Lpp38 is similar to those of the Escherichia coli polyamine transport proteins PotD (70%) and PotF (33%). P. haemolytica Lpp38 is present in both inner membrane and outer membrane fractions of the cell envelope. Susceptibility of Lpp38 to cleavage by extracellular proteases indicates that portions of the protein are surface-exposed. A protein of similar molecular mass in P. haemolytica strains from all 12 serotypes of biotype A and in an untypeable strain was detected by an anti-Lpp38 monoclonal antibody. Lpp38 is recognized by sera from calves resistant to infection after natural exposure to P. haemolytica and by sera from calves protected against infection by vaccination with P. haemolytica A1 outer membranes or with live bacteria. These data suggest a role for this protein in the development of immunity to P. haemolytica infection.

Identification of immunogenic, surface-exposed outer membrane proteins of Pasteurella haemolytica serotype 1.

Pasteurella haemolytica serotype 1 (S1) is the bacterium most frequently recovered from the lungs of cattle that have succumbed to shipping fever pneumonia. P. haemolytica outer membrane proteins (OMPs) are important immunogens in the development of resistance to pneumonic pasteurellosis. The purpose of this study was to identify the repertoire of immunogenic, surface-exposed P. haemolytica (S1) OMPs, that could be important in the development of protective immunity. We determined surface exposure of OMPs by (1) their susceptibility to protease treatment and (2) their ability to adsorb out antibodies from bovine immune sera. For a comprehensive identification of immunogenic, surface-exposed OMPs, we used bovine antisera from calves that were resistant to experimental P. haemolytica challenge after (1) natural exposure to P. haemolytica, (2) vaccination with live P. haemolytica, or (3) vaccination with P. haemolytica OMPs. We identified 21 immunogenic, surface-exposed P. haemolytica OMPs. Most were recognized by all three immune sera. However, some were recognized by one or two of the three antisera. Our analyses identified surface-exposed, immunogenic proteins that were not identified in previous studies.

Outer membrane proteins of bovine Pasteurella multocida serogroup A isolates.

The outer membrane proteins (OMPs) of P. multocida serotypes A3 (7 isolates), A4 (2 isolates), A3,4 and A2 (one isolate each) obtained from pneumonic cattle (10 isolates) and from one pig isolate were investigated to identify potential immunogens. SDS-PAGE of P. multocida OM isolated by SDG centrifugation of spheroplasts revealed eight major OMPs. Outer membranes isolated by sarcosyl extraction or SDG had similar protein composition on Coomassie blue-stained SDS-PA gel and on immunoblots. Two major OMPs (M(r)s of 35 and 46 kDa at 100 degrees C) demonstrated heat modifiability with apparent M(r)s of 30 and 34 kDa at 37 degrees C, respectively. The N-terminal aa sequences of these heat modifiable proteins revealed homology with E. coli OmpA and Hib P1 proteins, respectively. Protease treatment of whole cells followed by western immunoblots using bovine convalescent sera identified several immunogenic, surface-exposed and conserved OMPs among the eleven P. multocida isolates examined. The whole organism SDS-PAGE profiles of the eleven P. multocida isolates differed such that six patterns were seen. These patterns could potentially be used as a typing system for P. multocida bovine isolates based on the molecular weights of whole cell proteins. The above observations have potentially important implications relative to the immunity to infection.

Category learning with minimal prior knowledge.

In 6 experiments, the authors examined the use of prior knowledge in category learning. Previous studies of the effects of knowledge on category learning have used categories in which knowledge was related to all of the category's features. However, people's knowledge of real-world categories often consists of many "rote" features that are not related to their prior knowledge. Five experiments found that even minimal prior knowledge (1 knowledge-relevant feature and 5 rote features per exemplar) can facilitate category learning. Posttests revealed that although the knowledge aided learning, subjects also acquired the rote features that were not related to knowledge, contradicting predictions of an attentional explanation of the knowledge effect. The results of Experiment 6 suggested that subjects attempt to link even rote features to their knowledge.

Category-based predictions: influence of uncertainty and feature associations.

Four experiments examined how people make inductive inferences using categories. Subjects read stories in which 2 categories were mentioned as possible identities of an object. The less likely category was varied to determine if people were using it, as well as the most likely category, in making predictions about the object. Experiment 1 showed that even when categorization uncertainty was emphasized, subjects used only 1 category as the basis for their prediction. Experiments 2-4 examined whether people would use multiple categories for making predictions when the feature to be predicted was associated to the less likely category. Multiple categories were used in this case, but only in limited circumstances; furthermore, using multiple categories in 1 prediction did not cause subjects to use them for subsequent predictions. The results increase the understanding of how categories are used in inductive inference.

The locus of knowledge effects in concept learning.

Three experiments investigated how knowledge influences concept formation and representation in a standard concept acquisition task. The primary comparison was among arbitrary concepts, which had meaningless features; meaningful concepts, which had meaningful features from different domains; and integrated concepts, which had meaningful features interconnected by common knowledge. Experiment 1 found that learning was superior for the integrated concepts but that there was little difference as a function of feature meaningfulness. Experiment 2 suggested that the integrated Ss were learning to form a knowledge-based schema as their concept representation because they did not distinguish the typicality of features that differed in frequency. Experiment 3 introduced a category whose features were from the same domain but were not otherwise related. This concept was as difficult to learn and use as the meaningful concepts were. These comparisons help specify the ways in which knowledge does and does not influence concept formation.

Categorizing objects in isolation and in scenes: what a superordinate is good for.

Many studies have shown that subjects are faster at categorizing objects into "basic" concepts than into more general superordinate concepts. However, all of these studies have used a categorization task in which single, isolated objects are identified. There is good reason to believe that superordinate concepts are typically used to refer to collections of objects rather than to individual objects. For example, people more often use the term furniture to refer to a number of pieces of furniture rather than to name a single piece. This suggest that superordinate concepts include information about multiple objects and their common relations, particularly the typical scenes in which such objects appear. Four experiments examined this possibility by investigating whether the basic concept advantage will decrease or reverse itself when subjects are asked to categorize an object as part of a scene. The results showed that the basic-superordinate difference did decrease when subjects categorized objects in scenes. Furthermore, when an object was placed in an inappropriate scene, there was more interference for superordinate identifications. The results suggest qualitative differences in the representations of superordinate and basic concepts.

Alignment and category learning.

Recent research shows that similarity comparisons involve an alignment process in which features are placed into correspondence. In 6 studies, the authors showed that alignment is involved in category learning as well. Within a category, aligned matches (feature matches occurring on the same dimension) facilitate learning more than nonaligned matches do (matches on different dimensions), although nonaligned matches still facilitate learning relative to nonmatches. Analogously, feature matches that cross category boundaries hurt learning more if they occur on the same versus a different dimension, and cross-category feature matches on different dimensions hurt learning relative to nonmatching features. Representational assumptions of category learning models must be modified to account for the differences between aligned and nonaligned feature matches.

Category differentiation in object recognition: typicality constraints on the basic category advantage.

When people are asked to decide whether an object is in a given category, they generally respond faster when the category is at the basic level (e.g., car) than when it is at the superordinate level (e.g., vehicle) or the subordinate level (e.g., sedan). Basic categories have shorter and more frequent names, are learned earlier, and are usually more highly differentiated than other categories (they are both specific and distinctive), but it is not clear which of these factors is responsible for the faster response to basic categories. In three experiments with natural language categories, we found evidence that objects can be identified fastest as members of differentiated categories, even when such categories have longer names and are learned later than less differentiated categories. Specifically, we argued that atypical subordinate categories (e.g., racing car) are highly differentiated and should therefore be responded to as fast as basic categories in object recognition. The results supported this view and also ruled out the hypothesis that objects are necessarily identified as members of basic categories before further identification. We discuss the implications of these findings for the use of category names as definite descriptions in discourse.

Predicting features for members of natural categories when categorization is uncertain.

An important function of concepts to allow prediction of unseen features. A Bayesian account of feature prediction suggests that people will consider all the categories an object could belong to when they judge the likelihood that the object has a feature. The judgment and decision literature suggests that they may instead use a simpler heuristic in which they consider only the most likely category. In 3 experiments, no evidence was found that participants took into account alternative categories as well as the most likely one when they judged feature probabilities for familiar objects in meaningful contexts. These results, in conjunction with those of Murphy and Ross (1994), suggest that although people may consider alternative categories in certain limited situations, they often do not. Reasons for why the use of alternative categories may be relatively rare are discussed, and conditions under which people may take alternative categories into account are outlined.

Isolation of Borrelia burgdorferi (Spirochaetales: Spirochaetaceae) from Ixodes scapularis and Dermacentor albipictus ticks (Acari: Ixodidae) in Oklahoma.

Borrelia burgdorferi was isolated from Ixodes scapularis Say and Dermacentor albipictus Packard that were removed as partially fed adults from white-tailed deer, Odocoileus virginianus Zimmermann, in Oklahoma. Isolation in media was accomplished only after homogenates of pooled field-collected ticks were inoculated into laboratory-reared Peromyscus leucopus and reisolated from the urinary bladder into BSK II media. Both isolates were confirmed by western blot analysis and reactivity with monoclonal antibody H5332. These are the first reported isolates of B. burgdorferi from Oklahoma from these two tick species and are the first isolates from ticks from the south-central United States that were infective for laboratory-reared P. leucopus.

Attempted transmission of Borrelia burgdorferi (Spirochaetales: Spirochaetaceae) (JDI strain) by Ixodes scapularis (Acari: Ixodidae), Dermacentor variabilis, and Amblyomma americanum.

Laboratory-reared Ixodes scapularis Say, Amblyomma americanum (L.), and Dermacentor variabilis (Say) were fed on New Zealand white rabbits experimentally infected with Borrelia burgdorferi (JDI strain). At repletion, spirochetes could be detected by dark-field microscopy only in I. scapularis. Acquisition rates were 18 and 21%. When previously exposed nymphs of each species were fed on susceptible rabbits, I. scapularis was the only tick of the three species that transmitted B. burgdorferi. When a single rabbit was experimentally infected with B. burgdorferi and infested at 7-d intervals with I. scapularis, A. americanum, D. variabilis, and a second time with I. scapularis, B. burgdorferi was detected again only in cultures from the two groups of I. scapularis. When molted nymphs from each tick species were allowed to feed on susceptible rabbits, spirochetes again were isolated only at necropsy from the rabbits on which the two groups of I. scapularis fed.

Developmental studies of Anaplasma marginale (Rickettsiales:Anaplasmataceae) in male Dermacentor andersoni (Acari:Ixodidae) infected as adults by using nonradioactive in situ hybridization and microscopy.

The development of Anaplasma marginale Theiler was studied in ticks using a nonradioactive in situ hybridization method developed in our laboratory. Male Rocky Mountain wood ticks, Dermacentor andersoni Stiles, were infected intrastadially by allowing them to feed for 7 d on an infected calf (acquisition feeding). The ticks were then removed and held in a humidity chamber for 5 d before being fed on a 2nd susceptible call for 10 d (transmission feeding). Two groups of 10 ticks were collected daily during the 22-d experiment. In one group one-half of each tick was processed and embedded in paraffin and in the other group one-half of each tick was embedded in LR White for in situ hybridization. The companion tick halves from each group were fixed and embedded in Dow Epoxy Resin resin for routine light and electron microscopy. As detected by in situ hybridization on LR White- and paraffin-embedded sections and by microscopy, initial infection of A. marginale in ticks occurred in gut tissues either on the 7th d of acquisition feeding or the 1st d of the held period and infection persisted throughout transmission feeding. The highest number of ticks with gut infection was observed on the 5th d of transmission feeding. Salivary glands became infected with A. marginale on the 1st day of transmission feeding and remained infected throughout the transmission feeding period. Peak infection was observed on day 4 of transmission feeding. After the beginning of transmission feeding, A. marginale infection was also observed in interstitial, reproductive, skeletal muscle, fat body, and Malpighian tubule tissues. Although A. marginale infection of ticks clearly originates in midgut epithelial cells, many tissues eventually become infected during transmission feeding, resulting in a generalized infection. The infection of multiple tissues may contribute to the ability of A. marginale infection to persist in intrastadially infected male ticks.