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BACTERIAL WILT OF CUCURBITS. 15 



On February 17 and again on March 8 the soil was all respaded and each compart- 

 ment set to plants grown in sterile soil and the soil in the bed reinoculated with 

 Bacillus tracheiphilus as before. In no case did a single plant in any of the compart- 

 ments contract wilt, although the suspensions used in inoculating the soil were 

 virulent, as shown by needle-punctured control inoculations. 



This set of experiments was discontinued about March 25, 1917, and the whole 

 cage wa3 taken up and removed to one of our experimental fields near Tuxedo, Md., 

 where with the slight variations noted below the experiment was repeated. 



Compartment 1. Control. Soil not inoculated, but cucumber seeds sown. 



Compartments 2 and 3. Soil inoculated with a well-water bacterial suspension from 

 4-day-old beef-agar cultures and sown to cucumber seeds while the soil was still moist. 



Compartment 4. A large number of cucumber vines freshly wilted from pure- 

 culture inoculation were buried in- the soil just before sowing cucumber seeds. 



Compartments 5 and 6. Controls. Not inoculated, but cucumber seeds sown and 

 wilt-free striped and spotted cucumber beetles introduced. 



Compartments 7 and 8. Soil inoculated and planted to cucumber seed as in Nos. 2 

 and 3, but in these cases the soil had been drenched with formalin (1-320) two 

 weeks previously to kill any insects and insect eggs and pupae present in the soil. 



In due season the seeds germinated, and the plants in all the compartments con- 

 tinued healthy and vigorous. On June 15 the soil was again inoculated with Bacillus 

 tracheiphilus as at first, and wilt-free striped and spotted cucumber beetles were 

 introduced into all the compartments except the control, No. 1. 



Up to June 27 no wilt had occurred anywhere in the compartments and all the 

 plants were pulled ; the soil was reinoculated as before, immediately replanted with 

 cucumber seed, and two days later more wilt-free cucumber beetles were added. 



Again on July 23 the soil was very heavily reinoculated. The cucumber plants 

 were now in full bloom. No wilt had yet appeared. 



On August 22 the soil was again inoculated. Beetles were present in all the com- 

 partments and the vines had been fed upon freely. The cage was under constant 

 observation up to September 15, but at no time did any wilt develop. 



In this same field similar tests were carried out at varying intervals throughout 

 the season, using three of the large cloth-covered cages (PI. II, fig. 1). In two of 

 these cages the soil was inoculated by bacterial suspension and in the third by bury- 

 ing 24 wilted vines in the soil about 1 inch below the surface. In the latter case 

 cucumber seeds were planted directly among these wilted vines. Wilt-free beetles 

 were added to each cage and the soil was inoculated several times during the season. 

 No wilt developed in these cages at any time. 



In all these experiments the virulence of the bacterial suspensions used was proved 

 by needle-puncture inoculations into cucumber seedlings. 



In these numerous greenhouse and field tests where the soil was 

 heavily inoculated with virulent bacteria or when freshly wilted 

 vines were buried in the soil and where wilt-free cucumber beetles 

 and other insects, nematodes, etc., were introduced with the cucum- 

 ber seeds sown in these soils no single case of bacterial wilt developed. 

 These results fully corroborate the findings from careful field obser- 

 vations, namely, that bacterial wilt of cucurbits develops under 

 field conditions only when introduced into the plant by human 

 agencies or by cucumber beetles which have previously fed upon 

 plants infected with the wilt organism. 



