Biomedical subjects

A M Friedlander

Publications and source records attributed to A M Friedlander.

At least 37 records · Page 2Linked to original sources

PubMed1999

Protective efficacy of recombinant Yersinia outer proteins against bubonic plague caused by encapsulated and nonencapsulated Yersinia pestis.

To evaluate the role of Yersinia outer proteins (Yops) in conferring protective immunity against plague, six yop loci from Yersinia pestis were individually amplified by PCR, cloned, and expressed in Escherichia coli. The recombinant proteins were purified and injected into mice. Most Yop-vaccinated animals succumbed to infection with either wild-type encapsulated Y. pestis or a virulent, nonencapsulated isogenic variant. Vaccination with YpkA significantly prolonged mean survival time but did not increase overall survival of mice infected with the nonencapsulated strain. The only significant protection against death was observed in YopD-vaccinated mice challenged with the nonencapsulated strain.

Animals↗

PubMed1999

Immune response to Yersinia outer proteins and other Yersinia pestis antigens after experimental plague infection in mice.

There is limited information concerning the nature and extent of the immune response to the virulence determinants of Yersinia pestis during the course of plague infection. In this study, we evaluated the humoral immune response of mice that survived lethal Y. pestis aerosol challenge after antibiotic treatment. Such a model may replicate the clinical situation in humans and indicate which virulence determinants are expressed in vivo. Immunoglobulin G enzyme-linked immunosorbent assay and immunoblotting were performed by using purified, recombinant antigens including F1, V antigen, YpkA, YopH, YopM, YopB, YopD, YopN, YopE, YopK, plasminogen activator protease (Pla), and pH 6 antigen as well as purified lipopolysaccharide. The major antigens recognized by murine convalescent sera were F1, V antigen, YopH, YopM, YopD, and Pla. Early treatment with antibiotics tended to reduce the immune response and differences between antibiotic treatment regimens were noted. These results may indicate that only some virulence factors are expressed and/or immunogenic during infection. This information may prove useful for selecting potential vaccine candidates and for developing improved serologic diagnostic assays.

Animals↗

PubMed1998

Anthrax as a potential biological warfare agent.

Anthrax is a zoonotic illness recognized since antiquity. Today, human anthrax has been all but eradicated from the industrialized world, with the vast majority of practitioners in the United States unlikely to have seen a case. Unfortunately, the disease remains endemic in many areas of the world, and anthrax poses a threat as a mass casualty-producing weapon if used in a biological warfare capacity.

Animals↗

PubMed1998

Protection against experimental bubonic and pneumonic plague by a recombinant capsular F1-V antigen fusion protein vaccine.

The current human whole-cell vaccine is ineffective against pneumonic plague caused by typical F1 capsule positive (F1+) strains of Yersinia pestis. The authors found this vaccine to also be ineffective against F1-negative (F1-) Y. pestis strains, which have been isolated from a human case and from rodents. For these reasons, the authors developed a recombinant vaccine composed of a fusion protein of F1 with a second protective immunogen, V antigen. This vaccine protected experimental mice against pneumonic as well as bubonic plague produced by either an F1+ or F1- strain of Y. pestis, gave better protection than F1 or V alone against the F1+ strain, and may provide the basis for an improved human plague vaccine.

Aerosols↗

PubMed1998

Comparative efficacy of experimental anthrax vaccine candidates against inhalation anthrax in rhesus macaques.

The authors examined the efficacy of Bacillus anthracis protective antigen (PA) combined with adjuvants as vaccines against an aerosol challenge of virulent anthrax spores in rhesus macaques. Adjuvants tested included i) aluminum hydroxide (Alhydrogel), ii) saponin QS-21 and iii) monophosphoryl lipid A (MPL) in squalene/lecithin/Tween 80 emulsion (SLT). Animals were immunized once with either 50 micrograms of recombinant PA plus adjuvant, or with Anthrax Vaccine Adsorbed (AVA), the licensed human anthrax vaccine. The serological response to PA was measured by enzyme linked immunosorbent assay. Lymphocyte proliferation and serum neutralization of in vitro lethal toxin cytotoxicity were also assayed. In all vaccine groups, anti-PA IgM and IgG titers peaked at 2 weeks and 4-5 weeks postimmunization, respectively. Five weeks postimmunization, animals in all vaccine groups demonstrated PA-specific lymphocyte proliferation and sera that neutralized in vitro cytotoxicity. Six weeks after immunization, the animals were challenged by aerosol with approximately 93 LD50 of virulent anthrax spores. Animals were bled daily for 1 week to monitor bacteremia, and deaths were recorded. Anti-PA ELISA titers in all groups of immunized animals were substantially increased 2 weeks after challenge. One dose of each vaccine provided significant protection (> 90%) against inhalation anthrax in the rhesus macaques.

Administration, Inhalation↗

PubMed1998

Antibiotic treatment of experimental pneumonic plague in mice.

A mouse model was developed to evaluate the efficacy of antibiotic treatment of pneumonic plague; streptomycin was compared to antibiotics with which there is little or no clinical experience. Infection was induced by inhalation of aerosolized Yersinia pestis organisms. Antibiotics were administered by intraperitoneal injection every 6 hours for 5 days, at doses that produced levels of drug in serum comparable to those observed in humans treated for other serious infections. These studies compared in vitro to in vivo activity and evaluated the efficacy of antibiotics started at different times after exposure. Early treatment (started 24 h after challenge, when 0 of 10 mice tested had positive blood cultures) with netilmicin, ciprofloxacin, ofloxacin, ceftriaxone, ceftazidime, aztreonam, ampicillin, and rifampin (but not cefazolin, cefotetan, or ceftizoxime) demonstrated efficacy comparable to streptomycin. Late treatment (started 42 h after exposure, when five of five mice tested had positive blood cultures) with netilmicin, ciprofloxacin, ofloxacin, and a high dose (20 mg/kg of body weight every 6 h) of gentamicin produced survival rates comparable to that with streptomycin, while all of the beta-lactam antibiotics (cefazolin, cefotetan, ceftriaxone, ceftazidime, aztreonam, and ampicillin) and rifampin were significantly inferior to streptomycin. In fact, all groups of mice treated late with beta-lactam antibiotics experienced accelerated mortality rates compared to normal-saline-treated control mice. These studies indicate that netilmicin, gentamicin, ciprofloxacin, and ofloxacin may be alternatives for the treatment of pneumonic plague in humans. However, the beta-lactam antibiotics are not recommended, based upon poor efficacy in this mouse model of pneumonic plague, particularly when pneumonic plague may be associated with bacteremia.

Animals↗

PubMed1998

Analysis of the Yersinia pestis V protein for the presence of linear antibody epitopes.

The V protein expressed by pathogenic Yersinia pestis is an important virulence factor and protective immunogen. The presence of linear B-cell epitopes in the V protein was investigated by using a series of 17 overlapping linear peptides. Groups of 10 mice were immunized intraperitoneally with 30 microg of each peptide on days 0, 30, and 60. Although the V protein-specific antibody response to the peptides varied, most of the peptides elicited high antibody titers. The immunized mice were challenged subcutaneously with 60 50% lethal doses (LD50) (1 LD50 = 1.9 CFU) of a virulent Y. pestis strain, CO92. None of the peptide-immunized mice survived challenge. The animals immunized with the V protein were completely protected against challenge. The immunogenicity of some of the V peptides was increased by conjugating them to keyhole limpet hemocyanin. Only one peptide (encompassing amino acids 1 to 30) conjugate demonstrated some protection; the others were not protective. In additional experiments, V peptides that reacted well with sera from mice surviving Y. pestis infection were combined and used to immunize mice. Although the combined peptides appeared to be very immunogenic, they were not protective. Therefore, the protective B-lymphocyte epitope(s) in the V protein is most likely to be conformational.

Amino Acid Sequence↗

PubMed1998

Short- and long-term efficacy of single-dose subunit vaccines against Yersinia pestis in mice.

A single, subcutaneous, 30-microg dose of either a combination of the Yersinia pestis proteins F1+V or a F1-V fusion protein adsorbed to the adjuvant aluminum hydroxide, protected Hsd:ND4 mice for one year against pneumonic plague. The recombinant F1+V vaccine provided significant protection as early as day 14 postimmunization. The current Plague Vaccine USP in a single 0.2-ml dose did not provide significant protection in this mouse model. Antibody titers to F1 and V peaked at approximately 5-12 weeks postimmunization and were still detectable one year later. These F1 and V subunit vaccines may offer effective long-term immunity with a reduced dosage schedule when compared with the presently licensed, formalin-killed, whole-cell vaccine.

Animals↗

PubMed1998

The pathology of experimental anthrax in rabbits exposed by inhalation and subcutaneous inoculation.

OBJECTIVE: Although rhesus monkeys are considered to be an appropriate model for inhalational anthrax in humans, an alternative for vaccine and therapeutic efficacy studies is desirable. This study characterized the pathology of lethal anthrax in rabbits challenged by subcutaneous inoculation and aerosol exposure. MATERIALS AND METHODS: New Zealand white rabbits were exposed by subcutaneous inoculation or aerosol to lethal doses of Bacillus anthracis spores. RESULTS: The pathology of anthrax in rabbits exposed by either route was similar, with principal findings occurring in the spleen, lymph nodes, lungs, gastrointestinal tract, and adrenal glands. The cardinal changes were hemorrhage, edema, and necrosis, with bacilli and limited leukocytic infiltration. Features that depended on the route of exposure included mediastinitis in aerosol-exposed rabbits, a primary dermal lesion after subcutaneous inoculation, and differences in the pattern of lymph node involvement. Lesions observed in rabbits were comparable to those of inhalational anthrax in humans and rhesus monkeys. Noteworthy differences included the lack of leukocytic infiltration in brain and meningeal lesions, the relatively mild mediastinal lesions, and a lower incidence of anthrax-related pneumonia in rabbits compared with humans. These differences may be attributed to the greater susceptibility of rabbits to anthrax. Increased susceptibility is associated with both reduced leukocytic response to the bacilli and a more rapid progression to death, which further limits development of leukocytic infiltrates in response to the basic lesions of hemorrhage and necrosis. Primary pneumonic foci of inhalational anthrax, which may be influenced by preexisting pulmonary lesions in humans, were not observed in our rabbits, which were free of preexisting pulmonary disease. CONCLUSION: Anthrax in rabbits may provide a useful model for evaluating prophylaxis and therapy against inhalational anthrax in humans.

Administration, Inhalation↗

PubMed1997

Clinical recognition and management of patients exposed to biological warfare agents.

Concern regarding the use of biological agents--bacteria, viruses, or toxins--as tools of warfare or terrorism has led to measures to deter their use or, failing that, to deal with the consequences. Unlike chemical agents, which typically lead to violent disease syndromes within minutes at the site of exposure, diseases resulting from biological agents have incubation periods of days. Therefore, rather than a paramedic, it will likely be a physician who is first faced with evidence of the results of a biological attack. We provide here a primer on 10 classic biological warfare agents to increase the likelihood of their being considered in a differential diagnosis. Although the resultant diseases are rarely seen in many countries today, accepted diagnostic and epidemiologic principles apply; if the cause is identified quickly, appropriate therapy can be initiated and the impact of a terrorist attack greatly reduced.

Anthrax↗

PubMed1997

Studies on the role of plasminogen activator in systemic infection by virulent Yersinia pestis strain C092.

Plasminogen activator is an outer membrane protease of Yersinia pestis encoded by the pla gene on plasmid pPst. Pla of the KIM-10 strain of Y. pestis appears to be required for the virulence from a subcutaneous (sc) but not an intraperitoneal (ip) or intravenous (iv) route of infection in mice. However, other strains of Y. pestis are highly virulent by the sc route yet lack pPst and pla. In this study, the pPst- Pestoides F strain was lethal to mice inoculated sc, with an LD50 (3 cfu), equal to that of C092, a virulent pPst+ strain. To analyse further the role of Pla in invasive infection, isogenic derivatives of C092, including one harboring pla with a frameshift mutation and another cured of pPst, were made. Although the ip LD50 of pPst- C092 and of the pla mutant were nearly identical to that of the wild type, the subcutaneous LD50 of the cured and mutant strains were 4 to 6 logs greater than that of wild type. Thus, pPst appears to be required for development of a lethal infection by some strains after sc inoculation but not after direct ip inoculation. Pla-associated virulence did not appear to be mediated by interference with the phagocyte chemoattractant C5a, as shown by the lack of correlation of C5a production with susceptibility to Y. pestis in C5a+ and C5a- congenic mice. In a footpad model of the early host response to subcutaneous infection, pPst- C092 proliferated at the subcutaneous injection site to a similar extent as did the wild type parent strain, and elicited a similarly large, local inflammatory response. However, the wild type was present at higher concentrations at more distant sites such as the popliteal lymph node and spleen.

Animals↗

PubMed1997

Passive protection by polyclonal antibodies against Bacillus anthracis infection in guinea pigs.

The protective effects of polyclonal antisera produced by injecting guinea pigs with protective antigen (PA), the chemical anthrax vaccine AVA, or Sterne spore vaccine, as well as those of toxin-neutralizing monoclonal antibodies (MAbs) produced against PA, lethal factor, and edema factor, were examined in animals infected with Bacillus anthracis spores. Only the anti-PA polyclonal serum significantly protected the guinea pigs from death, with 67% of infected animals surviving. Although none of the MAbs was protective, one PA MAb caused a significant delay in time to death. Our findings demonstrate that antibodies produced against only PA can provide passive protection against anthrax infection in guinea pigs.

Animals↗

PubMed1997

Protection of mice from fatal bubonic and pneumonic plague by passive immunization with monoclonal antibodies against the F1 protein of Yersinia pestis.

Monoclonal antibodies (MAbs) to the fraction 1 (F1) protein of Yersinia pestis protected mice against fatal pneumonic as well as bubonic plague from wild-type F1+ organisms. The rare isolation of a virulent F1- isolate from surviving animals supports earlier studies suggesting that improved vaccines should consist of immunogens to protect against F1- variants. The high degree of protection with IgG MAb suggests that secretory IgA is not required for protection from pneumonic plague.

Animals↗

PubMed1997

Bacterial filamentation of Yersinia pestis by beta-lactam antibiotics in experimentally infected mice.

OBJECTIVE: To identify alternatives to streptomycin for treating pneumonic plague, we evaluated beta-lactam antibiotics to treat experimental pneumonic plague in mice. METHODS: Mice were exposed to a lethal inhaled dose of Yersinia pestis and treated with beta-lactam antibiotics administered every 6 hours, starting 42 hours postexposure. RESULTS: The mice died or were euthanized in extremis 3 days postexposure. We observed marked bacterial filamentation of Y pestis in the tissues of mice treated with ceftazidime (10/10 mice), aztreonam (9/10 mice), or ampicillin (1/10 mice), but not in the tissues of mice treated with cefotetan, cefazolin, ceftriaxone, or saline. There was no evidence of septation of the filamentous bacteria by light or electron microscopy. The filamentous bacteria were confirmed as Y pestis based on their reactivity with rabbit anti-Y pestis F1 serum. CONCLUSIONS: Marked bacterial filamentation of Y pestis can be produced in vivo by certain beta-lactam antibiotics. This antibiotic-induced morphologic change is important because filamentous bacteria in clinical samples could possibly be confused with filamentous actinomycotic organisms.

Animals↗

PubMed1996

Recombinant V antigen protects mice against pneumonic and bubonic plague caused by F1-capsule-positive and -negative strains of Yersinia pestis.

The purified recombinant V antigen from Yersinia pestis, expressed in Escherichia coli and adsorbed to aluminum hydroxide, an adjuvant approved for human use, was used to immunize outbred Hsd:ND4 mice subcutaneously. Immunization protected mice from lethal bubonic and pneumonic plague caused by CO92, a wild-type F1+ strain, or by the isogenic F1- strain C12. This work demonstrates that a subunit plague vaccine formulated for human use provides significant protection against bubonic plague caused by an F1- strain (C12) or against substantial aerosol challenges from either F1+ (CO92) or F1-(C12) Y. pestis.

Animals↗

PubMed1996

Fraction 1 capsular antigen (F1) purification from Yersinia pestis CO92 and from an Escherichia coli recombinant strain and efficacy against lethal plague challenge.

As a first step in formulating an improved plague vaccine, we developed a simple purification strategy that produced high yields of pure cell-associated and culture supernatant-derived fraction 1 capsular antigen (F1) from both avirulent Yersinia pestis C092 (Pgm- Lcr-) and an Escherichia coli F1-producing recombinant strain. Cell-associated F1 was partially purified by sequential ammonium sulfate precipitations of a sodium chloride extract of acetone-dried bacteria harvested from broth cultures. Cell-free F1 was precipitated directly from culture supernatants with a single application of 30% ammonium sulfate. By exploiting the aggregative property of F1, large quantities of purified high-molecular-weight F1 species from both cell extracts and supernatants were isolated in the void volume of a preparative gel filtration column. Highly purified, endotoxin-free F1, combined with two different adjuvants, induced very high F1 titers in mice and protected them against either subcutaneous (70 to 100% survival) or aerosol (65 to 84% survival) challenge with virulent organisms. This protection was independent of the source of the antigen and the adjuvant used. F1-induced protection against both subcutaneous and aerosol challenge was also significantly better than that conferred by immunization with the licensed killed whole-cell vaccine. Our results indicate that F1 antigen represents a major protective component of previously studied crude capsule preparations, and immunity to F1 antigen provides a primary means for the host to overcome plague infection by either the subcutaneous or respiratory route.

Animals↗

PubMed1996

Pathology of experimental pneumonic plague produced by fraction 1-positive and fraction 1-negative Yersinia pestis in African green monkeys (Cercopithecus aethiops).

OBJECTIVE: The protein capsule of Yersinia pestis, known as Fraction 1 or F1, is a protective immunogen and is an assumed, but not proven, virulence factor. Our objectives were to determine if inhaled F1-negative and/or F1-positive strains of Y pestis were virulent in the African green monkey and, if so, to differentiate F1-negative from F1-positive monkeys. Because F1-negative strains have been isolated from natural sources and have caused experimental fatal disease, we felt that this information was crucial to the development of future vaccines and diagnostic tests. MATERIALS AND METHODS: Adult African green monkeys were exposed by aerosol to F1-positive (CO92, n=15) or F1-negative (CO92-C12, n=6; Java-9, n=2) Y pestis strains. RESULTS: All monkeys died 4 to 10 days postexposure and had lesions consistent with primary pneumonic plague. Antibodies to F1 antigen and other Y pestis antigens allowed us to differentiate F1-positive from F1-negative Y pestis strains in fixed tissues. CONCLUSIONS: In this study, F1 antigen was not a required virulence factor. Therefore, there may be a need for vaccines and diagnostic assays that are not solely based on the F1 antigen.

Acute Disease↗

PubMed1995

Relationship between virulence and immunity as revealed in recent studies of the F1 capsule of Yersinia pestis.

Yersinia pestis, the causative agent of plague, possesses multiple virulence determinants encoded on its three plasmids and on its chromosome. We evaluated the role of the protein capsule F1 in virulence an immunity against plague. Strains lacking F1, either those that are naturally occurring or those with genetically defined nonpolar mutations in the structural gene, retained their virulence for mice and nonhuman primates. However, both active immunization with F1, from either a recombinant vector or Y. pestis, and passive immunization with F1 monoclonal antibody protected mice from experimental infection with wild-type F1-positive organisms. These results suggest that protective immunogens like F1 need not be essential for virulence. The rare isolation of virulent F1-negative organisms from F1-immunized animals infected with F1-positive strains supports this conclusion and also suggests that, in addition to F1, an optimal vaccine against plague should include essential virulence factors as immunogens.

Animals↗