Ann. Medit. Burns Club - vol. 6 - n. 2 - June 1993
HISTOLOGICAL ASSESSMENT OF THE LEVEL OF BURN WOUND INFECTION: DIAGNOSTIC AND THERAPEUTIC STRATEGIES Di Lonardo A., Ferrante M., Maggio G., Bucaria V., Del Zotti M., Brienza E. Istituto di Chirurgia Plastica Ricostruttiva ed Estetica, UniversitA clegli Studi, Bari, Italy SUMMARY. In the context of recent developments reported in the literature regarding methods for the histological assessment of burn wound infection, in addition to traditional microbiological techniques, the results are presented of a study of 40 patients admitted to the Bari Bums Centre in the period 1990-91. The histological analysis of the biopsies provided useful information which made it possible to have a better understanding of the parameters supplied by traditional microbiological tests and therefore to adopt a more personalized therapeutic approach in individual patients. The frequency and the gravity of septic complications in the seriously burned patient impose the need for an accurate diagnostic and therapeutic approach. The microbiological monitoring of surface swabs and biopsies from burn wounds provides precise information as to the type of infection present and the bacterial charge, but it gives no indication as to the depth of the infection. Microbiological investigation should therefore be complemented by histological examination in order to detect the possible presence of micro-organisms in the tissues beneath the eschar. This study combines the results obtained by microbiological monitoring and by histological analysis, and establishes the infection rate in burndamaged areas on a scale of values of increasing severity. Our experience has shown this method to be a very reliable criterion of assessment for the choice of therapeutic strategies, particularly as regards the timing and the site of surgical intervention. Material and methods The study included all burn patients admitted to
the Bari Burns Centre Intensive Care Unit between January 1990 and December 1991 within 24
hours of the burn lesion and belonging to classes III, IV and V of the Roi index. Biopsies
were taken from each patient, both from the burned areas and from apparently healthy areas
in the vicinity of the burn, on the basis of a predetermined map. Two samples were taken
from each biopsied area. One of these was fixed in 10% formalin and subjected to
histological examination; the other was kept dry in a sterile container and examined
microbiologically. Histological technique The biopsies fixed in 10% formalin were sent to the Bari University Pathological Histology Laboratory. After inclusion in paraffin, serial sections were prepared perpendicular to the skin surface, with a thickness of 5 ~t (6, 8). Twenty slides were prepared for each histological sample, with the following stains: The histological specimens were examined by optical microscope (magnification 400 and 1000 times). The depth reached by the bacterial invasion was graded according to the following scale (9): GRADE 0: no micro-organisms GRADE 1: some micro-organisms
GRADE 2: invasion of surface dermis GRADE 3: invasion of all dermis GRADE 4: invasion of hypodermis and/or underlying tissues. Microbiological technique The biopsies were weighed, homogenized, diluted as necessary in physiological solution and inoculated in doses of I ml into sterile Petri dishes to which about 15 ml of ordinary broth were added for the bacterial count and 15 ml of Sabouraud's agar for the cultivation of fungi. After 48-72 hours of incubation at 37 'C a count was made of any colonies that had developed. At the same time, the 1/100 dilution in physiological solution was used for seeding the following cultivation media: A portion of the solution (about 0.1/0.2 ml) was seeded in ordinary broth and in Sabouraud's agar from which subcultures were prepared after 24 hours respectively on the Murtz medium and on MSA from ordinary broth and on Sabouraud's agar from Sabouraud's broth. Case histories and results Using the above methods we studied 40 patients admitted to the Bari Burns Centre Intensive Care Unit between January 1990 and December 1991. The patients belonged to classes 3, 4 or 5 on the Roi prognostic index and were aged between 26 and 45 years. The burns ranged between 30% and 70% BSA. The results were obtained using the double-blind technique and relate to a total of 352 biopsies (176 histological and 176 microbiological). Histological results Both the level of infection in the tissues and the search for possible mycotic contamination were considered. Haematoxylin-eosin staining enabled us to distinguish the various tissue planes and to detect the presence of fungi, and also to hypothesize the possibility of bacterial infection. Grarn-modified and methylene blue staining made it possibile to detect the presence of bacteria in the tissues and to assess the depth reached. Table 1 shows the histological results related to the grading of infection observed in the 176 biopsies examined. Microbiological results The microbiological assessment of the various biopsies supplied a number of parameters. Here we give only those relative to the tissue bacterial charge (expressed as the number of bacteria per gm of tissue) and to the presence of fungal colonies. The results of the 176 biopsies are given in Table 1.
Discussion Two main considerations can be made on the
basis of these results. 1. From the point of view of reconstructive surgery, the concept of early escharectomy in the severely burned patient (which in our experience means operations performed before day 10) must be associated with that of emergency necrectomy when, in the presence of documented massive bacterial invasion of the viable tissues underlying the eschar, it is necessary to operate very early (day 4 or 5) in the districts most exposed to risk. The level to which escharectomy should
extend depends on the histological assessment of the depth of the infection. In areas
where bacterial colonization of the subcutaneous tissue is demonstrated (Grade 4)
necrectomy must be performed as far as the muscle fascia independently of the apparent
viability of the tissues as observed intraoperatively. Conclusions The histological investigation
conducted on burn lesion biopsies represents in our experience a further step towards a
better understanding and definition of the intricate problem of infection in seriously
burned patients. The method described here, followed according to standardized protocols
described in the literature (9), allowed a more rational analysis of the parameters
supplied by microbiological tests and thus enabled us to grade the tissue infection more
accurately. This method made it possible to adopt a personalized therapeutic approach on
the basis of precise laboratory indications, and not merely a standard therapy. RESUME. Dans le contexte des plus récents développements décrits dans la littérature pour ce qui concerne les méthodes pour l'évaluation histologique de l'infection des brûlures, outre les techniques microbiologiques traditionnelles, les auteurs présentent les résultats d'une étude effectuée sur 40 patients hospitalisés chez le Centre des Brûlés de Bari, pendant la période 1990-91. L'analyse histologique des biopsies a fourni des informations utiles qui ont permis aux auteurs d'avoir une meilleure compréhension des paramètres fournis par les analyses mierobiologiques traditionnelles et d'adopter en conséquence une approche thérapeutique plus personnalisée chez les patients individuels. BIBLIOGRAPHY
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