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  • Author or Editor: Johannes Storz x
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SUMMARY

Eight strains of Chlamydia psittaci isolated from swine with pneumonia, pleuritis, pericarditis, and enteritis were characterized through analysis of the major outer membrane protein gene ompA by a two-step polymerase chain reaction, by their interactions with cells in culture, and the morphologic features and ultrastructure of intra-cellular inclusions. Amplified chlamydial ompA dna fragments were differentiated by restriction endonuclease digestion. Chlamydial isolates were separated into 2 types on the basis of ompA restriction fragment length polymorphism. Strains of type L71 had finely granular inclusions, whereas those of type 1710S contained pleomorphic reticulate bodies (rb) in the inclusions, which are characteristic of aberrant chlamydial developmental forms. Chlamydial types L71 and 1710S required centrifuge-assisted inoculation for efficient infection of cell cultures. Cultivation in cell culture medium containing cycloheximide increased the numbers of chlamydial inclusions about 1.5-fold. These strains formed few elementary bodies in yolk sac cells of chicken embryos. Ultrastructurally, unique doublet rb were observed, particularly in strains of the ompA type L71. These doublets consisted of 2 rb, bounded by a cytoplasmic membrane, contained within a common cell wall and an extended periplasmic space. Ultrastructural examination of strains of the ompA type 1710S confirmed the aberrant chlamydial developmental forms, but evidence of viral infection of the rb as a cause of these aberrant forms was not found. The strain S45 isolated from intestinal sites of swine was a trachoma restriction fragment length polymorphism type. With the mouse biotype, it represented the second isolate from animals of Chlamydia trachomatis.

Free access
in American Journal of Veterinary Research

Abstract

Objective

To analyze the antigenic cross reactivity of various proteins of strains of Chlamydia pecorum, C psittaci, and C trachomatis.

Samples and Procedures

Strains FC-Stra and LW-613 of C pecorum; strains B577, Fitz-9, and 6BC of C psittaci, and strain LGV-2 of C trachomatis were studied. Strains of C pecorum were propagated in Georgia bovine kidney cells, and other chlamydial strains were propagated in L cells or Georgia bovine kidney cells. Partially purified chlamydial elementary bodies propagated in RAG cells, a BALB/c cell line cloned from a renal adenocarcinoma of BALB/c mice, were used to immunize BALB/c mice for production of monoclonal antibodies. Rabbits were inoculated with yolk sack-propagated, purified elementary bodies to produce polyclonal antisera. The reaction of monoclonal antibodies and polyclonal antisera with chlamydial proteins was analyzed by use of immunoblot techniques.

Results

Two monoclonal antibodies reacted with a 90-kd protein of C psittaci and a 94-kd protein of C pecorum strains. One monoclonal antibody reacted strongly with a 67-kd protein of C pecorum and strain B577 of C psittaci, but weakly with proteins of strains 6BC and LGV-2. Another monoclonal antibody reacted with a 46-kd protein of 2 C pecorum strains and of strain B577 of C psittaci, but not with those of strains 6BC and LGV-2. Two monoclonal antibodies reacted with a 20-kd protein of C pecorum and a 22-kd protein of C psittaci and LGV-2 strains. Polyclonal antisera reacted similarly with the proteins identified by monoclonal antibodies in the various chlamydial strains.

Conclusions

Reactions of several monoclonal antibodies with chlamydial antigens indicated that 67- and 46-kd proteins contain genus- and species-specific epitopes, respectively; a 94-kd protein of C pecorum is homologous to a 90-kd protein of C psittaci and C trachomatis strains; and a 20-kd protein of C pecorum corresponds to a 22-kd protein of C psittaci and C trachomatis. (Am J Vet Res 1996;57:1720–1725)

Free access
in American Journal of Veterinary Research

Abstract

Objective

To investigate the susceptibility of polarized epithelioid human rectal tumor (HRT-18G) cells to bovine coronaviruses (BCV) isolated from enteric (EBCV) and respiratory (RBCV) tract infections.

Procedure

Cells of the G clone of HRT-18 were grown to confluent monolayers on permeable supports, and were directionally infected at the apical and basolateral domains with 3 wild-type BCV strains, RBCV-LSU-94LSS-051-2, RBCV-OK-0514-3, and EBCV-LY138-2, and 1 cell culture-adapted strain, EBCV-L9-80. Sequential cytopathic changes were microscopically monitored. Medium samples for titration of hemagglutinins and viral infectivity were collected directionally from both domains of the infected cell cultures at various intervals.

Results

Polarized epithelioid HRT-18G cells from apical domains had maximal susceptibility to infection with the EBCV and RBCV strains, and those from basolateral surfaces had minimal susceptibility. Titers of hemagglutinins and infective progeny BCV reached 1,280 hemagglutinin units and 4.2 × 108 plaque-forming units/ml for apical samples, but were minimal for basolateral samples. Asymmetric virus release occurred through the apical surfaces of the HRT-18G cells by 12 hours after infection when cell fusion as a sign of cytopathic changes began. When cells were infected basolaterally, progeny virions released from apical surfaces reinfected the target cells from the apical domains and induced cytopathic changes were delayed about 12 hours, compared with changes detectable in apically exposed cultures.

Conclusions

EBCV and RBCV, isolated from cattle, had marked tropism for polarized epithelioid HRT-18G cells. Entry of BCV into the polarized HRT-18G cells was effected maximally through the apical domains and minimally through the basolateral domains. Release of progeny BCV occurred preferentially from the apical domains. (Am J Vet Res 1997;58:1120–1124)

Free access
in American Journal of Veterinary Research

Abstract

Objective

To investigate the antigenic diversity of lipo-oligosaccharides of Haemophilus parasuis.

Procedures

Immunoblot assays were done with monoclonal and polyclonal antibodies on whole-cell lysates. Individual colonies of H parasuis strains H 54, H 53, and H 128 were tested for reactivity with lipo-oligosaccharide-specific monoclonal antibodies after a single passage on chocolate agar, and colonies of strain H 54 were analyzed after 10 passages. Colony blot tests were used to screen H parasuis strains for spontaneously occurring antigenic variation in their lipo-oligosaccharides.

Results

Eight H parasuis strains were separated into 4 lipo-oligosaccharide serovars on the basis of immunoblot reactions with 3 polyclonal rabbit antisera. Nine monoclonal antibodies against lipo-oligosaccharides of a lipo-oligosaccharide-serovar I strain reacted with all tested serovar I strains but failed to react with other H parasuis strains.

Conclusions

Variations in the antigenic reactivity after 1 or 10 passages on chocolate agar were not observed. The serovar I lipo-oligosaccharide strains included virulent as well as avirulent H parasuis strains, indicating that these epitopes do not correlate directly with virulence properties of H parasuis. (Am J Vet Res 1996;57:63-67)

Free access
in American Journal of Veterinary Research

Abstract

Objective—To identify cytocidal viruses and Pasteurella spp that could be isolated from cattle involved in 2 natural outbreaks of shipping fever.

Design—105 and 120 castrated male 4- to 8-monthold feedlot cattle involved in 1997 and 1998 outbreaks, respectively.

Animals—Nasal swab specimens and blood samples were collected, and cattle were vaccinated on arrival at an order-buyer barn from 4 local auction houses. Four days later, they were transported to a feedlot, and additional nasal swab specimens and blood samples were collected. Nasal swab specimens were submitted for virus isolation and bacterial culture; blood samples were submitted for measurement of respiratory bovine coronavirus (RBCV) hemagglutinin inhibition titers.

Results—93 of 105 cattle and 106 of 120 cattle developed signs of respiratory tract disease during 1997 and 1998, respectively, and RBCV was isolated from 81 and 89 sick cattle, respectively, while at the orderbuyer's barn or the day after arrival at the feedlot. During the 1997 outbreak, bovine herpesvirus 1 was isolated from 2 cattle at the order-buyer's barn and from 5 cattle 7 and 14 days after arrival at the feedlot, and parainfluenza virus 3 was isolated from 4 cattle 14 days after arrival at the feedlot. During the 1998 outbreak, bovine herpesvirus 1 was isolated from 2 cattle at the order-buyer's barn and on arrival at the feedlot and from 5 cattle 7 and 14 days after arrival at the feedlot, and parainfluenza virus 3 was isolated from 1 animal the day of, and from 18 cattle 7 and 14 days after, arrival at the feedlot. Pasteurella spp was cultured from 4 and 6 cattle at the order-buyer's barn and from 92 and 72 cattle on arrival at the feedlot during the 1997 and 1998 outbreaks, respectively.

Conclusion and Clinical Relevance—Results suggest that RBCV may play a causative role in outbreaks of shipping fever in cattle. More than 80% of the sick cattle shed RBCV at the beginning of 2 outbreaks when the Pasteurella spp infection rate was low. (J Am Vet Med Assoc 2000;216:1599–1604)

Full access
in Journal of the American Veterinary Medical Association