Lorne A. Babiuk

Viral Bacterial Synergistic Interaction in Respiratory-Disease

Publisher Summary Humans and animals are constantly inoculated with various microorganisms resident in the upper respiratory tract and by inhaled aerosols, yet pneumonia is a relatively rare event. This implies the existence of very efficient defense mechanisms that are capable of eliminating the vast majority of microorganisms before they colonize and multiply to sufficient levels, resulting in clinical diseases. In order to overcome this continuous barrage of microorganisms, there is a complex array of defense mechanisms present in the upper and lower respiratory tract capable of eradicating these organisms. However, in individuals suffering from a variety of diseases, including virus infections, colonization occurs rapidly with subsequent development of pneumonia. Thus, it is estimated that 90% of bacterial pneumonias develop after a viral infection. Furthermore, individuals suffering from a viral pneumonia have a 40% chance of developing bacterial pneumonia. The reasons for the increased colonization of the lung by bacteria following virus infections are related to the surface properties of epithelial cells lining the respiratory tract, the physiological environment of the respiratory tract, and the alteration of the specific and nonspecific defense mechanisms of the lung that occurs as a result of virus infection.

Analysis of bovine herpes virus-type 1 isolates by restriction endonuclease fingerprinting.

SummaryIn an effort to determine whether distinct types of bovine herpesvirus 1 are responsible for causing specific syndromes, the polypeptides and DNA of 93 BHV1 isolates from the province of Alberta as well as vaccine strains and numerous other Canadian, U.S. and European isolates were analyzed by PAGE and restriction endonuclease fingerprinting, respectively. The polypeptide patterns showed only slight variations: only six isolates contained polypeptides that varied from the norm in their molecular weight, or were absent. Although on the basis of endonuclease patterns the isolates could be categorized into three “strain”s and nine “sub-strains”, we were unable to associate any of the strains with specific clinical signs. This suggests that the type of disease caused may be determined more by the route of infection and animal management practices than by the inherent properties of certain types of BHV1. Most of the Albertan isolates were of one BHV1 type and sub-type.

Effect of Chronic Administration of Recombinant Bovine Tumor Necrosis Factor to Cattle

Cachectin/tumor necrosis factor-alpha (TNF), a protein produced by macrophages upon stimulation, has been implicated as an important mediator of inflammatory processes and of clinical manifestations in chronic infectious diseases. In order to study further the potential role of TNF in infectious diseases, a homologous system was employed in which recombinant Escherichia coli (E. coli) derived bovine TNF (rBoTNF) was injected in cattle, either as a single bolus or in a repetitive treatment-regime. No clinical signs were observed, although changes occurred in hematologic and immunologic parameters when less than 0.5 mg of TNF/100 kg body weight was administered twice daily for 18 days. Prolonged treatment with 0.05–0.5 mg/100 kg induced histologic but no gross changes in the kidneys and liver. When doses were increased above 0.5 mg/100 kg, depression, anorexia, cachexia, and diarrhea appeared rapidly. Pathologic changes were apparent in various tissues including liver, kidneys, and lymphoid organs; body fat depots were depleted. Most of these changes appeared to be reversible; return to normal tissue-morphology occurred within 3 weeks of withdrawal of rBoTNF. The clinical and pathologic changes induced by prolonged rBoTNF administration resembled those observed in some chronic parasitic and viral infections of cattle in which macrophage-activation characteristically occur. Our finding may be relevant to the elucidation of the pathogenesis of these and other chronic infections.

In vitro generation of hydrogen peroxide and of superoxide anion by bovine polymorphonuclear neutrophilic granulocytes, blood monocytes, and alveolar macrophages.

Studies were conducted to compare the capacity of bovine blood monocytes, polymorphonuclear granulocytes (PMNs), and alveolar macrophages (AMs) to generate hydrogen peroxide and Superoxide anion. Following stimulation with opsonized zymosan, bovine PMNs respond with an immediate and vigorous liberation of both oxygen species, generating 4.7 ± 0.3 nmol H2O2/106 cell and 12.3 ± 1.8 nmol O2−/106 cell during the initial 15 min. This is more than twice the amount generated by AMs (1.2 nmol H2O2/106 cell; 2.5 nmol O2/106 cell) and blood monocytes (0.5 nmol H2O2/106 cell; 2.1 nmolO2−/106 cell) during the same period. However, AMs continue generating H2O2 and O2 at a steady rate for a longer period and consequently produce amounts equal to those of PMNs when measured over a longer time span. Also, AMs can be stimulated with nonopsonized zymosan in contrast to PMNs. However, the AM population appears to comprise at least two subpopulations, which can be clearly distinguished by their capacity for generation of reactive oxygen species, and which correlate with their tendency for adherence to a plastic surface. In contrast to what has been found in other species, the bovine phagocytes were found to lack receptors for tuftsin and formylated oligopeptides, and thus remained unresponsive to these compounds. The in vitro activity of the three cell types was found to be very dependent on culture conditions, such as cell density and an adherent versus suspended state. In addition, a comparison with macrophages and PMNs elicited into the mammary gland suggest that in vivo factors can significantly influence the in vitro activities. The mammary gland cells have lower activity than blood and alveolar cells, even though they have been “primed” by chemotactic factors), and this is probably caused by milk components, i.e., the microenvironment. Our observations are discussed with respect to the results obtained from different laboratories, different species, and different cell types; emphasis is placed on the problem of drawing conclusions about in vivo functions of cells from parameters assayed in vitro.

Role of interferon-γ in inducing cytotoxicity of peripheral blood mononuclear leukocytes to bovine herpesvirus type 1 (BHV-1)-infected cells☆

Abstract In the present study, the possible role of Interferon (IFN)-γ on the induction of cytotoxic activity of peripheral blood mononuclear leukocytes (PBML) from BHV-1-immune cattle was investigated. Supernatants obtained from BHV-1-immune PBML, stimulated under conditions similar to those required to demonstrate cytotoxicity, contained an antiviral substance capable of inducing 2′-5′-oligoadenylate synthetase activity in MDBK cells and MHC class II antigen expression on epithelial cells. These supernatants also contained IFN-α, but were devoid of tumor necrosis factor and interleukin-2 biological activities. Further studies during primary infection and hyperimmunization with BHV-1 showed that IFN-γ production and non-MHC-restricted cytotoxicity against BHV-1-infected targets always occurred concomitantly, suggesting that they represent an important part of the detectable CMI responses mounted against this virus. Furthermore, it was also demonstrated that cytotoxicity of PBML against BHV-1-infected cells was reduced with the addition of antibodies to bovine IFN-γ to the cytotoxic assay. Bovine recombinant IFN-γ was able to enhance the in vitro cytotoxic activity of PBML from immune cattle, but not from their nonimmune counterparts. This suggests that other factors, in addition to IFN-γ, may be essential in the development of non-MHC-restricted cytotoxic responses during BHV-1 infection.

Antigenic and immunogenic characteristics of bovine herpesvirus type-1 glycoproteins GVP 3/9 and GVP 6/11a/16, purified by immunoadsorbent chromatography.

Glycoproteins GVP 3/9 and GVP 6/11a/16, two of the major glycosylated proteins specified by bovine herpesvirus type-1 (BHV-1), were purified on immunoadsorbents consisting of the appropriate monoclonal antibodies linked to Affigel-10. Each glycoprotein, whether purified from virus-infected cells or from virus, retained antigenic activity and induced high titers of monospecific antibodies in rabbits. These antibodies could neutralize virus and mediate complement-dependent lysis of virus-infected cells. Denatured glycoproteins GVP 3, GVP 6, GVP 11a, and GVP 16, which were purified by immunoadsorbent chromatography, followed by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis, also retained antigenicity and immunogenicity, though to a lesser extent than the native glycoproteins. Antibodies induced by GVP 9, GVP 6, and GVP 11a could also neutralize and mediate immune lysis. Even though GVP 16 induced high levels of antibody, these antibodies could not neutralize virus or participate in antibody and complement-mediated cytolysis. These results may suggest that the orientation of GVP 6/11a/16 in the membrane is such that GVP 11a is better exposed on the virion envelope and the cell surface than GVP 16. Cross-reactivity between monospecific antibodies against GVP 3 and GVP 9, as well as GVP 6, GVP 11a, and GVP 16 supported the previously proposed hypothesis that GVP 3 (180K) is a dimer of GVP 9 (91K) and that GVP 6 exists in two forms: one being a 130K polypeptide and the other composed of GVP 11a (74K) and GVP 16 (55K) linked by disulfide bonds. These data suggest that, thus far, either GVP 6/11a/16 or GVP 3/9 may be a potential candidate for a subunit vaccine against BHV-1 infection.

Characterization of surface receptors on bovine leukocytes.

Three bovine leukocyte populations--peripheral blood lymphocytes (PBL), mammary gland polymorphonuclear neutrophils (PMN) and macrophages (Mo)--were characterized with respect to five surface markers: surface immunoglobulin (SIg), sheep erythrocyte receptor, complement (C) receptor and Fc receptors for both IgG and IgM. The majority of PMN and Mo possessed C and Fc receptors for IgG, but lacked SIg and the erythrocyte receptor. The PMN, but not Mo, also expressed a Fc receptor for IgM. The PBL were heterogeneous with respect to their surface characteristics and evidence was presented for the following subtypes: (a) cells with the E receptor alone; (b) cells with E receptor plus the Fc(IgG) receptor; (c) cells with SIg plus the C receptor but minus the Fc(IgG) receptor; (d) lymphocytes with SIg plus the C receptor and the Fc(IgG) receptor, and (e) cells lacking E receptors and SIg but bearing Fc(IgG). It was assumed, but not proven, that some of these latter cells must also bear the C receptor. The significance of the various cell types in antiviral defense is briefly discussed.

Bovine Interferon - its Biology and Application in Veterinary-Medicine

Abstract Investigations of the production and potential use of bovine interferons against viral infections have occurred since the first descriptions of interferons in other systems. The recent advent of recombinant DNA-technology has facilitated such studies and furthered our knowledge about the bovine interferon system in general. This review gives an overview of the biology, antiviral and immunomodulatory activities of bovine interferons. Areas in which the interferons are now applied or have potential application in viral diseases in cattle are described. Finally, the value of studies of the bovine interferon system with respect to comparative interferon research is discussed.

Bovid herpesvirus type-1 (Infectious bovine rhinotracheitis virus) induced thymidine kinase

Abstract This communication reports the presence of a new thymidine kinase (dTK) in bovine kidney cells infected with Bovid herpesvirus 1(BHV-1). The virus-induced enzyme activity differs from the host cell enzyme with respect to substrate specificity, thermostability, and the ability to use phosphate donors other than ATP. Thus, the BHV-1-induced dTK could be selectively assayed when CTP was used in place of ATP as the phosphate donor in the enzyme reaction. The virus-induced activity can phosphorylate thymidine, bromodeoxyuridine, bromovinyldeoxyuridine, and methylmethoxydeoxyuridine but not deoxycytidine, bromodeoxycytidine, deoxyguanosine, or acycloguanosine.

The role of antibody dependent cytotoxicity in recovery from herpesvirus infections

Abstract In order to implicate antibody dependent cytotoxicity (ADC) 1 as playing a possible role in recovery from herpesvirus infections, target cells were infected with virus and the time of susceptibility to ADC was determined. Additional virus-host cell events were measured in parallel including the time of extracellular and intracellular transmission of infectious virus. Cells were susceptible to ADC at about the same time as intracellular transmission of virus occurred. The presence of effector cells and antibody in infected cultures markedly diminished viral transmission, indicating that ADC may play an important role in curtailing infections.