The Natural History of Pneumonic Tularemia in Female Fischer 344 Rats after Inhalational Exposure to Aerosolized Francisella tularensis Subspecies tularensis Strain SCHU S4.

The inbred Fischer 344 rat is being evaluated for testing novel vaccines and therapeutics against pneumonic tularemia. Although primary pneumonic tularemia in humans typically occurs by inhalation of aerosolized bacteria, the rat model has relied on intratracheal inoculation of organisms because of safety and equipment issues. We now report the natural history of pneumonic tularemia in female Fischer 344 rats after nose-only inhalational exposure to lethal doses of aerosolized Francisella tularensis subspecies tularensis, strain SCHU S4. Our results are consistent with initial uptake of aerosolized SCHU S4 from the nasal cavity, lungs, and possibly the gastrointestinal tract. Bacteremia with hematogenous dissemination was first detected 2 days after exposure. Shortly thereafter, the infected rats exhibited fever, tachypnea, and hypertension that persisted for 24 to 36 hours and then rapidly decreased as animals succumbed to infection between days 5 and 8 after exposure. Tachycardia was observed briefly, but only after the core body temperature and blood pressure began to decrease as the animals were near death. Initial neutrophilic and histiocytic inflammation in affected tissues became progressively more fibrinous and necrotizing over time. At death, as many as 1010 colony-forming units were found in the lungs, spleen, and liver. Death was attributed to sepsis and disseminated intravascular coagulation. Overall, the pathogenesis of pneumonic tularemia in the female F344 rat model appears to replicate the disease in humans.

Autophagy protects against active tuberculosis by suppressing bacterial burden and inflammation.

Autophagy is a cell biological pathway affecting immune responses. In vitro, autophagy acts as a cell-autonomous defense against Mycobacterium tuberculosis, but its role in vivo is unknown. Here we show that autophagy plays a dual role against tuberculosis: antibacterial and anti-inflammatory. M. tuberculosis infection of Atg5(fl/fl) LysM-Cre(+) mice relative to autophagy-proficient littermates resulted in increased bacillary burden and excessive pulmonary inflammation characterized by neutrophil infiltration and IL-17 response with increased IL-1α levels. Macrophages from uninfected Atg5(fl/fl) LysM-Cre(+) mice displayed a cell-autonomous IL-1α hypersecretion phenotype, whereas T cells showed propensity toward IL-17 polarization during nonspecific activation or upon restimulation with mycobacterial antigens. Thus, autophagy acts in vivo by suppressing both M. tuberculosis growth and damaging inflammation.

Cowpox virus inhibits human dendritic cell immune function by nonlethal, nonproductive infection.

Orthopoxviruses encode multiple proteins that modulate host immune responses. We determined whether cowpox virus (CPXV), a representative orthopoxvirus, modulated innate and acquired immune functions of human primary myeloid DCs and plasmacytoid DCs and monocyte-derived DCs (MDDCs). A CPXV infection of DCs at a multiplicity of infection of 10 was nonproductive, altered cellular morphology, and failed to reduce cell viability. A CPXV infection of DCs did not stimulate cytokine or chemokine secretion directly, but suppressed toll-like receptor (TLR) agonist-induced cytokine secretion and a DC-stimulated mixed leukocyte reaction (MLR). LPS-stimulated NF-κB nuclear translocation and host cytokine gene transcription were suppressed in CPXV-infected MDDCs. Early viral immunomodulatory genes were upregulated in MDDCs, consistent with early DC immunosuppression via synthesis of intracellular viral proteins. We conclude that a nonproductive CPXV infection suppressed DC immune function by synthesizing early intracellular viral proteins that suppressed DC signaling pathways.

Stress-induced Hsp70 gene expression and inactivation of Cryptosporidium parvum oocysts by chlorine-based oxidants.

Our research on the mechanisms of action of chlorine-based oxidants on Cryptosporidium parvum oocysts in water revealed a dual-phase effect: (i) response to oxidative stress, which was demonstrated by induced expression of the Hsp70 heat shock gene, and (ii) oocyst inactivation as a result of long-term exposure to oxidants. The relative biocidal effects of sodium hypochlorite (bleach) and electrolytically generated mixed oxidant solution (MOS) on C. parvum oocysts were compared at identical free chlorine concentrations. Oocyst inactivation was determined by quantitative reverse transcription-PCR (qRT-PCR) amplification of the heat-induced Hsp70 mRNA and compared with tissue culture infectivity. According to both assays, within the range between 25 and 250 mg/liter free chlorine and with 4 h contact time, MOS exhibits a higher efficacy in oocyst inactivation than hypochlorite. Other RNA-based viability assays, aimed at monitoring the levels of beta-tubulin mRNA and 18S rRNA, showed relatively slow decay rates of these molecules following disinfection by chlorine-based oxidants, rendering these molecular diagnostic viability markers inappropriate for disinfection efficacy assessment.

Genetic similarity of Puumala viruses found in Finland and western Siberia and of the mitochondrial DNA of their rodent hosts suggests a common evolutionary origin.

A total of 678 small mammals representing eight species were trapped in western Siberia in 1999-2000 and assayed for the presence of hantaviruses. Eighteen animals, all Clethrionomys species, were antigen positive by enzyme-linked immunosorbent assay (ELISA). Small and medium genome segments were recovered by RT-PCR from six samples from Clethrionomys glareolus and three from Clethrionomys rufocanus. Sequence comparison and phylogenetic analysis revealed that these hantaviruses were Puumala virus and were similar to hantavirus strains from Finland. To confirm these data, partial nucleotide sequences of the rodent hosts' cytochrome b genes were obtained, as well as several sequences from genes from rodents trapped at different localities of European Russia and western Siberia. The cytochrome b sequences of Siberian bank voles were similar to sequences of C. glareolus, trapped in Finland. These data suggest that the Puumala hantaviruses, as well as their rodent hosts, share a common evolutionary history. We propose that these rodents and viruses may be descendents of a population of bank voles that expanded northward from southern refugia during one of the interglacial periods.