METCALF 



Gross macroscopic examination of the lungs showed complete con- 

 solidation to have occurred in the mice kept at 4^ C while 50 per 

 cent consolidation was found in lungs removed from mice kept at 

 20° C. Enzyme production occurred in both groups, but inhibitor 

 levels declined only in the cold exposed mice. A cold environment, 

 therefore, led to a more rapid growth of virus which in turn led to a 

 greater yield of enzyme and a decline in inhibitor content (Fig, 8). 



The importance of neuraminidase activity to cell damage was 

 shown by the results obtained following inoculation of mice with Al 

 and A strains having no detectable enzyme action. Virus multipli- 

 cation was limited, no enzyme was produced and there was no loss 

 of inhibitor. The lungs removed at sacrifice were normal in appear- 

 ance and no cell damage could be demonstrated (Fig. 9). 



FLUOROCHROME STUDIES OF EXPERIMENTAL 

 INFLUENZA INFECTIONS 



Host response to virus infection at the cellular level was studied 

 with the help of the fluorochrome acridine orange. Monkey kidney 

 monolayers were infected with A2 strains of high and low enzyme 

 activity at environmental temperatures of 20° C and 37° C, and ex- 

 amined at intervals up to 5 days. "Flying coverslips" were fixed in 

 chilled absolute methyl alcohol, hydrated, and stained 5 to 9 minutes 

 with a 1 to 10,000 dilution of acridine orange at a pH of 3.8. The 

 coverslips were examined with a Fluorestar microscope used in 

 conjunction with an Osram HBO- 200 illuminator. The exciter filter 

 used was a Corning No. 5860, half- thickness filter. Photomicro- 

 graphs were made using Ektachrome, type B film. 



Non- infected monolayers showed the green fluorescing nuclei 

 characteristic of DNA material, with yellowish- white nucleoli. The 

 cytoplasm exhibited a rust color symptomatic of RNA. Six hours 

 after infection the cytoplasm around the nucleus began to show a 

 brighter rust fluorescence. By 18 hours the fluorescence became a 

 bright orange- red color and could easily be distinguished. At 30 



358 



