Chapter IV 



57 — 



Methods of Enumeration 



organisms on glass slides submerged in sea water was observed by ZoBell 

 and Allen (1933) who noted the appearance of dividing cells, chains of 

 bacteria, and micro-colonies. The microorganisms are so tenaciously at- 

 tached that it is necessary only to stain the slides without fixation for 

 microscopic examination. ZoBell and Allen (1935) made continuous 

 studies for several months on microorganisms found attached to standard 

 25 X 75 mm. glass slides submerged in the sea by means of a special car- 

 rier (Fig. 6). Besides observing spe- 

 cies of Leptothrix, Actinomyces, and 

 other microorganisms which were 

 never demonstrated by cultural pro- 

 cedures, they isolated bacteria from 

 submerged slides by plating them 

 with nutrient agar. 



According to Henrici (1933), 

 Karzinkin (1934) was the first to 

 use the submerged slide technic for 

 studying the occurrence of bacteria 

 in water. Karzinkin applied the 

 term bacterial periphyton to bacteria 

 which grow on submerged slides. 

 Henrici referred to them as periphy- 

 tic bacteria. The term seems hardly 

 an apt one to apply to bacteria be- 

 cause, as emphasized by Roll's 

 (1939) review of the literature on 

 periphytes and epiphytes, its conno- 

 tation is ambiguous from a view- 

 point of both etymology and current 

 usage. The terms sessile and seden- 

 tary have long been used to describe 

 organisms which grow attached to 

 solid surfaces. That most of the bac- 

 teria found attached to submerged 

 slides attach themselves there and 

 are not merely passively stuck is 

 proved by the work of ZoBell 

 (19436) on the relation of bacteria to 

 solid surfaces. ZoBell and Allen 

 (1935) used the term attachment bac- 

 teria to describe the bacteria found 

 on submerged slides, and Hotchkiss 

 and Waks]vla.n (1936) speak of the 

 attachment count as the number of 

 bacteria found on submerged slides. 

 According to Hotchkiss and Waksman (1936), the submerged slide 

 technic, in contrast to other direct microscopic methods, can be used in 

 the case of waters containing small numbers of bacteria. A direct correla- 

 tion was found between plate counts and attachment counts in sea water 

 samples at different temperatures, in sea water samples to which carbon 

 and nitrogen compounds were added, in sea water inoculated with pure 

 cultures, and in filtered sea water. These workers reported that "Not 

 only were high values for the plate counts associated with high values for 

 the attachment counts, but constants could be determined from experi- 



FiG. 6. — Carrier for submerging in 

 water glass slides to be examined by the 

 direct microscopic method. Several such 

 carriers may be submerged in tandem to 

 provide for the simultaneous collection of 

 attachment microorganisms from different 

 depth. A weight or anchor must be fas- 

 tened to the bottom cord. The carriers 

 may be constructed of wood, plastics, or 

 other materials. Glass slides are secured 

 by slipping them under the cord which 

 encircles the carrier and into the slot near 

 the bottom of the carrier. 



