MYCOBACTERIOSIS 



" A COMPARISON OF HISTOPATHOLOGIC TECHNIQUES FOR DETECTING ACID-FAST BACILLI. Mote, R. F., 

 Gigstad, D. C, Himes, E. M. and Muhm, R. L. (Project Report) 



Summary 



Two hundred and fifty suspected tuberculous bovine specimens were examined for acid-fast bacilli using three 

 different histopathologic techniques. These included an auramine o-stained smear, an auramine-o stained tissue 

 section and a new fuchsin-stained tissue section procedure. The combination of an auramine o-stained smear and an 

 auramine o-stained tissue section had a statistically signilicant advantage for detecting mycobacteria. 



Introduction 



In support of the bovine tuberculosis eradication program suspected granulomas received from slaughter 

 establishments are routinely examined at Veterinary Services Diagnostic Laboratory, Ames, Iowa. Upon collection, 

 tissues are split and one portion is put in 10 percent buffered formalin to be examined by the Ilistopathology 

 Section and the other portion is put in saturated sodium borate solution for examination by the Mycobacteriology 

 Section. This study presents histopathology results from 250 suspected tuberculous submissions and compares three 

 different staining procedures. Also included are mycobacteriology results if available. 



Materials and Methods 



Histopathologic preparations for microscopic examination were made on each of 250 submission suspected of 

 being tuberculous. These were '"run of the mill" specimens, biased only by selection for the presence of a gross 

 lesion large enough to furnish sufficient material for examination. There was some variation in the content of the 

 individual submissions. Some contained only lymph node, others lymph nodes and lung, or liver, or spleen or various 

 other combinations of tissues. 



Tissues were removed from formalin, examined for lesions, cut in for paraffin impregnation and a portion of 

 lesion was collected for smear preparation. Smears were stained with auramine-o (AO) (2). 1 1'araffin-impregnated 

 tissues were blocked, sectioned at six microns and mounted on slides. Replicate tissue sections were prepared, one 

 stained with a modified new fuchsin (NF) (1) procedure, another stained with a AO preparation (3) and a third 

 stained with hematoxylin and eosin. A Leitz Orthoplan microscope equipped with a Xenon lamp and BG-12 blue 

 light, BG-38 heat and K-530 barrier filters was used for fluorescent examinations. A Leitz Orthoplan equipped for 

 light microscopy was used for examination of H&E and NF-stained sections. 



Mycobacterial isolation and typing results were obtained for 220 of the 250 submissions. (Some had not been 

 submitted for culture and others not examined because they had been previously diagnosed as Coccidioidomycosis.) 



A comparison of the techniques for detecting acid-fast bacilli was made to determine if there were statistically 

 signilicant dillerences in efficiency using one or a combination of staining procedures. 



Results 



The results of using the different techniques to detect acid-fast bacilli have been presented in table 1. The 

 AO-stained smear and AO-stained section combination had a significant advantage when compared statistically with 

 a combination of the AO-stained smear and the NF-stained section (p. >.05). More positives were detected in 



Numbers in parentheses refer to References at the end of this report. 



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