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Experimental studies of bacterial cholecystitis
EXPERIMENTAL
STUDIES OF BACTERIAL HANS G.
From the Department
ARONSOHN, of Surgery,
CHOLECYSTITIS *
M.D.
University
of Chicago
CHICAGO
B
OTH Andrews’ and Magner,2 studying the contents of human gaII bIadders removed at operation, have found 65 per cent to contain sterile biIe. 0sIer3 pIaces this figure at 52 per cent. An important roIe in the production of cholecystitis seems to be pIayed by factors other than infection, mainIy those of a chemicaI nature. Recent investigations conducted by us4 have shown that bile acids, which Iong have been known to be of high toxicity for tissues outside of the gaI1 bIadder, deveIop a definite toxicity for the gaIl-bladder waI1 itseIf if present in a suffrcientIy high concentration. Further studies are now in progress on the exact role pIayed by the biIe acids. AIthough the importance of infection in this connection is possibIy not primary, its existence as a factor cannot be compIetely disregarded for severa reasons. First, one is tempted to draw an anaIogy as regards the gaI1 bIadder from the fact that infection plays an important part in the pathology of most other organs. Secondly, one must take into account the fact that many reIiabIe workers, afterextensive studies, have concIuded that infection is the main factor in the production of cholecystitis. Very IittIe is known about the actual route which the infection foI10~s. The five possibihties, as given in Graham’s textbook,5 are as foIIows: (I) descending infection from the Iiver; (2) ascending from the duodenum via the common duct; (3) hematogenous; (4) Iymphatic; (5) directIy through the galIbIadder waI1 from an inflamed adjacent organ. To determine the route by which the infection spreads is not the essentia1 point of this study. Our purpose is to ascertain by anima1 experimentation whether or not *Work
an increase either in number or viruIence of the bacteria in bile Ieads to choIecystitis, and whether different bacteria show different behaviors in this regard. A considerabIe number of bacteria may be present in norma biIe, but since these gaI1 bIadders do not manifest any changes either chnicaIIy or anatomicaIIy, we are not justified in caIIing such biIe “infected.” BiIe may be considered as “infected” if it contains either an abnormaIIy high number of bacteria or bacteria of especiaIIy great viruIence. The organisms might be those which occur normaIIy or might accrue in the biIe from any pathoIogica1 sources in the body. Any bacterioIogica1 study of the biIiary system presupposes knowIedge of the norma bacterioIogica1 ffora of the Iiver and gaI1 bIadder as we11 as the kind and origin of pathoIogica1 organisms and the route by which these might reach the gaI1 bladder. Since our experimenta studies are based on certain fundamental factors which are considered to be proved, a brief review of the Iiterature is in place. The occurrence of cholecystitis in cases of typhoid fever Ied earIy investigators to consider the infectious nature of gaII-bIadder disease. As earIy as 1763, John Hunter6 associated inff ammation of the gaI1 bIadder with typhoid fever, an idea which was proved bacterioIogicaIly by GiIbert and Girode’ in 1890 and confirmed cIinicalIy by Chiari.8 There foIIowed a series of experimenta.1 studies which attempted to produce “carriers ” or even reactions of the gaI1 bIadder, by injection of typhoid baciIIi. BIacksteing found the biIe to contain bacteria for many days or weeks foIIowing the intravenous injection of BaciIIus coli and Bacillus typhosus in rabbits. Other
done in part under a grant from the DougIas Smith Foundation. 18
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Aronsohn-ChoIecystitis
bacteria were subsequentIy used by further observers, Sherringtonl” injecting BaciIIus anthracis, BaciIIus pyocyaneus, and staphyIococci and obtaining I 8 “infected ” biles from 49 cases. Whether or not the gaIIbIadder waI1 showed any pathology in these experiments couId not be ascertained from the data given in the Iiterature, since the chief interest of the author was directed towards the production of “carriers.” GiIbert,l’ in I 894, considered BaciIIus coIi to be the most important organism in the production of cho1ecystiti.s and favored the idea of an ascending infection from the duodenum. This view was opposed by Cushing and Livingood,12 who found the duodenum to be very Iow in bacteria1 content and even steriIe when empty. Further contributions were made whiIe attempting to produce stones experimentaIIy. Cushing,13 after direct inocuIation of typhoid baciIIi into the gaI1 bIadders of rabbits, observed inflammation of the gaI1 bladders with the formation of concrements. SimiIar experiments were performed on dogs and rabbits by ItaIia,14 who used attenuated cuItures of BaciIIus coIi and BaciIIus typhosus. Th is author mentions an inflamed gaII-bIadder mucosa, incidental to the formation of concrements. RoIIeston15 emphasizes the need for an additiona factor other than the presence of bacteria, such as trauma, stones, or stasis, for the production of choIecystitis. This author points out that though the frequency of typhoid fever is decreasing (BaciIIus typhosus is found in the biIe in aImost a11 severe cases of typhoid fever), there is an increase in the frequency of choIecystitis. SimiIarIy, OsIerlfi states that choIecystitis in typhoid fever occurred in onIy 19 of his series of 1500 cases. Later workers again considered the bIood stream to be the route by which bacteria reach the gaI1 bIadder and produce choIecystitis. There was great disagreement, however, as to whether the organisms reach the gaI1 bIadder directIy through the arteria1 bIood stream or indirectly by way of the porta vein, Iiver, and duct system. Rosenow, l7 who demonstrated the
American
Journal
of Surgery
19
specific affinity of streptococci and partIy aIso of BaciIIus coIi to the gaI1 bIadder, considers the direct arteria1 route to be the
Diagram: Showing carmula and gaII bIadder after the common duct had been opened and the catheter introduced into the gal1 bladder.
usua1 mode of infection, since he obtained positive reactions by intravenous bacteria1 injection and negative results by direct inocuIation into the gaI1 bIadder. SchiibI,‘* on the other hand, who tried to produce “cholera carriers” experimentally, obtained the opposite resuIts, direct inocuIation into the gaI1 bIadder being more effective than intravenous injection. There is no mention in this work whether or not a change occurred in the gaII-bIadder waI1. Doerrlg observed that excretion of typhoid baciIIi into the biIe, which occurred reguIarIy after injection of the former into the systemic veins, could be prevented by Iigation of the cystic duct. On the basis of this experimenta work he concIuded that bacteria indirectIy reach the gal1 bIadder by excretion through the Iiver. Koch20 and ChiaroIanza,21 however, arrived at the opposite concIusion. Typhoid baciIIi were found in the gaI1 bIadder after intravenous injection even after a preceding Iigation of the cystic duct. This observation, together with the fact that microscopic study of a large number of inflamed gaI1 bIadders reveaIed “emboIi” in the
20
American Journd of Surgery
Aronsohn-ChoIecystitis
foIds of the mucous membrane, Ied these authors to con&de that “a transverse infection of the waI1” is the usua1 route. NichoIs22 concurred with the hematohepatogenous route of Doerr on the basis of his work on rabbits. This author found that injection of typhoid baciIIi into the mesenteric veins yieIded “gaI1-bIadder Iesions ” in 63 per cent of the cases, whiIe injection into the ear vein resuIted in onIy 4 I per cent of positive cases. Emmerich and Wagner,23 after direct injection of typhoid baciIIi into the gaI1 bIadders of rabbits, obtained a chronic infiItration and thickening of the gaIl-bIadder waI1. However, the fact that these animaIs had been immunized with typhoid serum previous to the injections might have sIackened and diminished the inffammatory reaction. In contrast to these observations, Venema,24 in simiIar experiments, obtained a norma gaI1 bIadder. In rgzr Meyer25 and his co-workers confirmed the vaIidity of the hematohepatogenous route, which they beIieved to be the onIy one proved definiteIy in the Iiterature, by observing positive reactions in the gaI1 bIadders of rabbits both by intravenous injection of typhoid baciIIi and direct inocuIation of the same organism into the gaI1 bladder. Rosenow’s theory of a hematogenous infection was further enhanced by Brown,26 who found mainly streptococci in the emuIsihed gaI1 bIadder tissue from human cases of ChoIecystitis. This author demonstrated seroIogfcaIIy the direct etioIogica1 importance of these streptococci and demonstrated their specific affinity for the gaI1 bIadder by intravenous injection in rabbits. The very compIete survey by Magner and Hutcheson reveaIed that the most common organism in the cavity of the gaI1 bIadder is BaciIIus coIi, whiIe in the waI1 it is streptococcus. These workers injected staphyIococci into both the waI1 and cavity of rabbits’ gaI1 bIadders and in every instance of subserous injection obtained positive reactions in the gaIIbIadder waIIs, but in only a limited number of cases foIIowing injection into the Iumen. examining gaI1 bIadders HunIemiiIIer,27
APRIL, 1936
obtained at human operations, revealed positive bacteria1 cuItures in IOO per cent of cases when he incIuded in his statistics both the gall-bIadder waI1 and the Iiver. During the Iast ten to fifteen years the beIief in the infectious origin of choIecystitis has become considerabIy weakened. Wherever the infectious theory is stiI1 maintained, the route is considered to be especiaIIy by way of the Iymphatics from the Iiver, as suggested by Graham. In 1922, Drennan,28 who emphasized the antibacteria1 activity of the biIe, found onIy rg of IOO biIes removed at operation, to be infected. The scarcity of cIinica1 signs of infection, the Iower fever and sIight Ieucocytosis, as we11 as the rare occurrence of Ieucocytes in biIe even in cases of so-caIIed “empyema,” based upon a study of a Iarge amount of surgica1 and necropsy materia1, convinced Denton2g that infection does not pIay a primary roIe in the production of ChoIecystitis but that certain other factors, such as mechanica or chemica1 ones, are the basic causes. FeinbIatt,30 in a study of 400 cIinica1 cases, obtained resuIts similar to Drennan and Denton, observing very few inffammatory changes in the gaIIbIadder waI1 and very few Ieucocytes in the biIe or ffuid content. From this brief review of the Iiterature, one can see the present confused state of the question of the relationship between the bacteria1 content of the gaI1 bIadder and the pathoIogy of that organ. The information yieIded by different authors varies considerabIy and often is contradictory. WhiIe bacteria1 studies of the gaI1 bIadder obtained at human operations and postmortem examinations are reIiabIe and vaIuabIe, experimenta studies of the same probIem are much Iess satisfactory, due to the differences in the techniques empIoyed by various investigators, some of which may be actuaIIy misIeading. The commonest method, introduction of the bacteria by means of a needIe puncture through the gaII-bIadder waI1, often resuIts in a IocaIized injury to the wal1 at the point of puncture, thus making the resuIts uncon-
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Aronsohn-Cholecystitis
trollabIe. This disadvantage was recognized by many earIy workers, Rosenow stating that “localized cholecystitis occurred at the point of puncture when the injection was made directIy into the gaII-bIadder.” SimiIarIy Venema mentions the fact that the infection produced by the usua1 technique, which consisted of needIe puncture with subsequent tying off of that part of the wall, is especiaIIy marked and is accompanied by a severe injury to the gaI1 bladder waI1. According to WiIkie,3’ the changes at the neck of the gaI1 bIadder, where the injection had been made, contrasted sharpIy with the fundus of the organ, which stiI1 appeared normal. The work of Phemister32 and Graham,33 however, indicates that it is possibIe to obtain normal gaI1 bIadders even with this usuai technique, if the proper degree of care is administered. In experiments with dogs, Phemister injected Streptococcus viridans intramuraIIy and performed in addition a cystic duct Iigation, yet the gaI1 bladders remained norma in a11 but one case, where a subsequent mixed infection accounted for the marked inflammatory changes. SimiIarIy, Graham and Peterman obtained negative resuIts folIowing the injection of BaciIIi coIi into the waI1 of the gaI1 bIadder in dogs. In this instance, however, additiona Iigation of the bIood vesseIs or cystic duct resulted in definite changes. Some improvement of the needIe puncture technique was made by Venema, who cauterized the waI1 at the point of the injection. It is perhaps to this improved technique that we can attribute the fact that this author obtained normal gaI1 bIadders after direct inocuIation of typhoid baciIIi, while simiIar experiments by Emmerich and Wagner yieIded positive reactions. Hospers34 varied the procedure by making the injection through the Iiver surface of the gaI1 bIadder, a method which Ieaves the free surface of the gaI1 bIadder practicaIIy unharmed, as we have seen, but may cause occasiona bIeeding in the liver tissue and thus inffuence the adjacent gaI1 bladder. The method of intravenous
American Journal of Surgery
21
injection practiced by Rosenow and many others is much more satisfactory. A positive reaction of the gaI1 bIadder foIIowing an intravenous injection of bacteria may be attributed with certainty to a specific affinity of these organisms to the biIiary system. Negative resuIts are vaIueIess in this connection. AIthough the organisms may not reach the gal1 bIadder via the blood stream, they may stiI1 possess a definite affnity for the biliary system if brought there by any other way. Another disadvantage of intravenous injection is the production of a bacteremia, which ordinariIy is not present in cases of human ChoIecystitis, and might faciIitate the production of choIecystitis by causing an abnorma1 reaction of the organism as an entity. It is now evident that there is great doubt as to which route bacteria actuaIIy foIIow, under either norma or pathoIogica1 conditions, which are found eventuaIIy in the gaI1 bIadder. ConsequentIy we wiI1 Iimit ourseIves to the effects of the actual presence of organisms in the gaI1 bIadder either of especiaIIy great viruIence or in abnormaIIy high numbers. This condition can be created experimentaIIy by means of technique.35 We our “nontraumatizing” empIoyed this procedure in most of our experiments, using the method of injection by a fine needIe puncture in onIy a few instances, which served to compare the two techniques. Method. Only dogs were used in our experimenta work, the operations being performed under ether anaesthesia and strictIy aseptic conditions. The abdomen was opened by a midline incision and the common duct incised close to the duodenum. A soft French rubber catheter was introduced into the gaI1 bIadder and IO to 13 C.C. of biIe were aspirated by means of a syringe. Injection of the bacteria1 suspension was accompIished at this point by means of another syringe, and this was foIlowed by re-injection of the aspirated biIe into the gaI1 bIadder, thus washing the common and cystic ducts as free as possibIe
22
American Journal of Surgery
Aronsohn-ChoIecystitis
from bacteria. It might incidentahy be mentioned at this point that this washing was suffrcientIy effective to prevent any marked infection of the mucous membrane of the ducts. The catheter was then withdrawn and a gIass cannuIa inserted into the opening which had been made in the common duct. By means of this technique, the gaI1 bIadder in most instances was not visuaIized during the entire procedure, practicaIIy assuring freedom from traumatization. In those instances in which we failed to introduce the catheter into the gaI1 bIadder, we injected the bacteria1 suspension by means of a direct needle puncture and rn addition performed a Iigation of the cystic or common duct. For this injection, we preferred aIways to use the Iiver surface of the gal1 bIadder rather than the free surface, thus minimizing the amount of trauma to the gaII-bIadder waI1. In severa cases a needIe puncture was used for the injection without any duct Iigation, to determine whether or not stasis of the biIe favors infection of the gaIl-bIadder waI1. The simpIe catheter-cannuIa technique is the onIy method that has yieIded norma controIs. It has aIways been a handicap in gaI1 bIadder studies that if injection was made through the gaII-bIadder waI1 a considerable degree of choIecystitis might arise and furthermore if the duct was Iigated or drained externaIIy, the contro1 experiments very frequentIy were not normaI. Therefore we have never been abIe previousIy to either remove or introduce materia1 into the gaI1 bIadder without causing some degree of damage, which vitiates the experiment. The diagram shows the cannuIa in pIace after the common duct had been opened and the catheter introduced into the gaI1 bIadder. The bacteria used for injection were obtained from human sources, as acute adenitis, sinusitis, acute and chronic mastoiditis, etc. The cuItures were of recent isoIation. The organisms were suspended in saIine soIution, in a concentration of approximateIy 500 miIIion bacteria per c.c.,
APRIL, ,936
the criterion used being the cIoudiness of the suspension as compared with one of known bacteria1 content. One C.C. of this suspension was generaIIy the amount injected. Wherever a marked reaction occurred, the experiment was repeated with 0.5 C.C. or Iess, whiIe 2 C.C. were used in those cases in which there was no reaction. The dogs were eIectrocuted twentyfour hours after the operation and postmortem examination performed as soon as possibIe. A microscopic examination of the gaII-bIadder waI1 was made in every case except those which died SpontaneousIy and where postmortem changes had occurred. Those animaIs which died from any other cause, as Ieakage from the gal1 bIadder or ducts, pneumonia, etc. were not incIuded in this study. Protocols. In aI1, 36 dogs were operated. In Charts I, II and III are shown the detaiIed resuIts of the experiments. A gaI1 bIadder was considered to be norma if it reveaIed no changes either on gross or microscopic examination. “ Cholecystitis I ” indicates that the gall-bIadder waI1 was of normal thickness, but there was some edema and congestion in the serosa. A fibrinous membrane was either compIeteIy absent or seen 0nIy in pIaces. “ChoIecystitis 2 ” incIuded those cases in which the gaII-bIadder waI1 was thickened up to 4 mm. and the inAammatory signs were more definite, the fibrinous exudate covering the serosa as a thick membrane. “Cholecystitis 3 ” indicates that the waI1 is thicker than 4 mm., the gaI1 bIadder usuaIIy Iies imbedded deepIy in the Iiver, and the adjacent organs, the duodenum, omentum, etc. form a Iarge inffammatory tumor around it. The terms “about normaI” or “sIight reaction” were used in those cases in which the changes were non-inflammatory in character, i.e., distention of the gaI1 bIadder, congestion of the serosa without any edema or fibrinous exudate, mechanica injury to the waI1, etc. Whenever the introduction of the catheter or insertion of the cannuIa was attended with diffrcuIty, the term “trauwas incIuded in the column matization”
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I
Wherever this IabeIIed “comphcations.” happened the value of the experimenta resuIts are Iimited, as they are in those instances in which the animals died spontaneousIy and postmortem changes occurred in the gaII bladder. SimiIarIy, experiments in which the common or cystic duct was tied are of vaIue onIy when the
American Journal of Surgery
23
within fifteen hours after the injection, the second was moribund in twenty-four hours, and the third was in poor condition when kiIIed. OnIy three drops had been injected into the anima1 which died. These resuIts are in direct contrast to the experiments on nine dogs in which the other three strains of Streptococcus hemoIyticus were injected.
CHART I STREPTOCOCCUS
Strain
Exp’t. No.
Strain
Injected
Amount
HEMOLYTICUS
from: (I) Acute mastoiditis 1-3 dogs (2) Acute sinusitis--6 dogs (3) Acute mastoiditis 11-2 dogs (4) Chronic mastoiditis--r dog Method of Introductior
1
Duct
Complications
ResuIts
-.
9 IO II I2
Acute mastoiditis
I
0.5 cc.
Catheter
CannuIa
Acute mastoiditis
I
3 drops
Catheter
CannuIa
Acute mastoiditis Acute sinusitis Acute sinusitis
I
3 drops
Catheter Catheter Catheter
CannuIa CannuIa CannuIa
2 cc. I cc.
Acute sinusitis Acute sinusitis Acute sinusitis
2 cc.
Acute sinusitis
I cc.
Acute mastoiditis Acute mastoiditis Chronic mastoiditis
I cc. I cc.
II
I cc.
II
I cc. I cc.
Catheter CannuIa Catheter CannuIa Common NeedIe punt :Iigation ture Catheter CannuIa Catheter Catheter Catheter
gaI1 bladder remained norma in spite of the bacteria1 injection and duct Iigation. Results. Since streptococci and BaciIIus coIi are the organisms most commonly accused of being the causative agents in the production of gaII-bIadder infection, we pIaced specia1 emphasis in our experimenta work on the actua1 effect which these bacteria have. Very striking resuIts were obtained with Streptococcus hemoIyticus, the detaiIs of which appear on Chart I. Four different strains of the organism were used for the injections. AIthough three of the four strains used were from acute human infectious processes, onIy one produced changes in the gaI1 bIadder. The changes were very marked and occurred in the 3 cases in which this particuIar strain was injected. One of these animaIs died
CannuIa Cannula CannuIa
Dying when electrocuted Died Very sick
ChoIecystitis 3, biIe peritonitis ChoIecystitis 3, postmortem changes ChoIecystitis 2-3
I 1: 1: 1: 1: 1: 1: t$ZrnIZ%E duct
!
( ““““” i :::::::::::: ~
n~E.t~
GaII bIadder norma ChoIecystitis I Thickening of galIbIadder wall Gal1 bIadder about norma Gal1 bIadder norma GaII bladder norma Gal1 bIadder norma
None of these animaIs died spontaneously nor did any of them exhibit any marked signs of iIIness during the twenty-four hours between the injection and eIectrocution. In 5 of these g cases, postmortem examination reveaIed perfectIy norma gaI1 bIadders, whiIe in 3 other cases the changes were very sIight and not inflammatory at aI1. In only one instance, that of dog No. 7, did a shght but definite cho1ecystiti.s deveIop foIIowing injection of the Streptococcus hemoIyticus from an acute sinusitis, and the technique was not accountabIe for this change. The injection of BaciIIus coIi yieIded definiteIy negative resuIts, which are summarized on Chart II. In 3 cases in which the catheter method had been empIoyed the gaI1 bIadder remained perfectly normaI.
American
24
Journal
of Surgery
Aronsohn-ChoIecystitis
In 4 animaIs in this group (4, 6, 7 and 8), the shght changes manifested couId definitely be attributed either to traumatization at the time of operation or to the Iigation of the cystic or common duct. In none of these were the changes so marked that they couId be attributed to the bacteria1 injection. OnIy one gaII bIadder (dog No. 3) reveaIed an intense reaction, apparentIy due to the ligation of the common duct. From Chart III it can be seen that the results Iikewise are negative. None of the organisms caused any inffammatory changes on the gaII-bIadder wall and any sIight change which occurred couId easiIy CHART BACILLUS
Exp’t. NO. I
Bacteria Injected
Amount
3 4 5
B. B. B. B. B.
CoIi Coli Coli Coli Coli
2 C.C.
6
B. CoIi
I C.C.
7
B. CoIi
I C.C.
8
B. Coli
1.5 C.C.
2
I C.C. I C.C.
I C.C. I C.C.
Method of Introduction
CannuIa Cannula CannuIa CannuIa Common ligation
Needle puntture NeedIe puntture Needle puntture
Common ligation Common ligation Cystic Iigation
be attributed to traumatization or duct Iigation. The charts show that when the same type of bacteria is injected through the catheter the gal1 bIadder remains normaI, but if injected by needle puncture, or if the cystic or common duct Iigation is added, then they cause cholecystitis or death. Microscopic examination of the gaI1 bladder in the experiments where streptococcus hemoIyticus isoIated from acute mastoiditis cases had been injected, reveaIs that the enormous thickening of the waI1 is caused chiefly by a swehing of the serosa1 Iayer to about five times its normaI thickness. The Iatter is edematous and hyper-
1936
emit, contains a Ieucocytic infiltration, and is covered with a puruIent fibrinous exudate. Although there is a scattered Ieucocytic invasion of the muscuIar layer, this as well as the mucosa1 surface is mostIy unchanged. Those gal1 bIadders, however, into which had been injected another strain of Streptococcus hemoIyticus as we11 as Staphylococcus, BaciIIus WeIchii and BaciIIus coli, proved to be practicaIIy norma both grossIy and microscopically. In summarizing these experimenta resuits, we may conclude that there is IittIe tendency for an acute bacteria1 infection of the gaIl-bladder waI1 to resuIt from the contents of that organ. OnIy Streptococcus II COLI
Duct
Catheter Catheter Catheter Catheter Needle puntture
APRIL.
Complications
Results
. ..,
duct
Some traumatization Some traumatization died
duct
Very sick
duct duct
GaII bladder norma Gall bIadder norma GaII bIadder normal ChoIecystitis I Cholecystitis 3, postmortem change, bile peritonitis DiIated gal1 bIadder with fibrinous exudate Gal1 bIadder about norma1 Slight reaction on gal1 bIadder
hemoIyticus gave a positive reaction, and this onIy in the case of one of the four strains used. Since traumatization or extension from an adjacent pathologica process can be definitely ruIed out as a cause for this change, we may deduce that the organism or its toxins present in the biIe reached the serosa1 Iayer of the waI1 by traversing the mucosa1 and muscular Iayers without causing any changes in these. None of the other organisms injected, aIthough present in the biIe in overwheIming numbers, caused any change in the gaII-bIadder wall. The experimenta resuIts we obtained partIy confirm and partly contradict those reported in the literature. We found bacteria1 infection of the gaII-bIadder waI1
Nrw
%nlts
VOL. XxX11,
Aronsohn-Cholecystitis
No. I
occur Iess frequently than recorded by other authors. AI1 observers have described the streptococcus as pIaying an important part in producing inflammation of the gallto
bladder waII. The diverse results we obtained with different strains of this organism concur with the observations of Rosenow and Magner and Hutcheson, who stressed the varying activity within this group on the basis of differences in viru-
Bacteria Injected
,\lethod of Introduction
Amount
_~
.I
Staph. Staph. Staph.
albus albus albus
16
I
cc.
Catheter Needle puncture NeedIe puncture
I
cc. cc.
Catheter Catheter
Cystic ligation CannuIa Cannula Cannula Cannula
2 cc.
Catheter Catheter
Staph. aureus
2 cc.
Needk
Staph. aureus
I cc.
Catheter
Strep. viridans
I cc.
Catheter
Common ligation Cannula
Strep. viridans
I cc.
Catheter
Cannula
Catheter Catheter Catheter Catheter
Cannula CannuIa Cannula Common ligation
1 B. Welchii
0.3
cc.
I cc.
0.5
Traumatization
., Traumatization
puncture
NeedIe puncture
cc.
duct
Common ligation
-
Ience. Magner and Hutcheson divided the streptococci into two chief groups, which they classified as typical and atypical: TypicaI streptococci were characterized by the formation of smooth, non-hemolytic, colorless or green coIonies on blood agar, and by the fact that they were kilIed by exposure to the action of bile. AtypicaI streptococci were characterized by the formation of rough, nonhemolytic, white colonies on bIood agar, and by
duct
Died
I ( , /:‘.:::...:::I duct
_.___
Gall bIadder norma Gall bladder norma Slight reaction on gaI1 bladder Gal1 bladder norma Gall bladder about normal Gal1 bladder normal BiIe peritonitis, some reaction on gall bIadder SIight reaction on gal1 bladder Cholecystitis 3, postmortem changes GaII bladd er about normal normal, Gall bladder some bile peritonitis GaII bladder normal Gail bladder normal GaII bladder norma Postmortem change on gaII bladder
I
Staph. aureus Staph. aureus
I cc.
Results -___-.
---
Cannula
1
2 cc.
I_-
I
Staph. aureus Staph. aureus
Strep. viridans B. Welchii B. Welchii B. WeIchii
Complications _~.
cc. cc.
2 cc.
dogs dogs 3 dogs
Duct I ____~.
1
I
23
The negative resuIts we obtained using Bacilli coli stand in direct disagreement both with the clinical data reported by GiIbert and the experimental work done with this organism by Italia. However, the fact that
Staphylococcus aureus--6 (3) Streptrococcus viridans(4) Bacilli Welchii-4 dogs
1
of Surgery
gall bladder.
(2)
-
Journal
the fact that they were not killed by the action of bile. Chronic cholecystitis was produced in 100 per cent of cases by injecting typica streptococci under the serous coat of a rabbit’s
CHAKT III (I ) Staphylococcus albus-
Esp’t. No.
American
: Traumatizxtion died
duct
-I
Gail bladder mal
about
nor-
StaphyIococcus aureus and albus, Streptococcus viridans and BaciIIus Welchii, a11 failed to produce cholecystitis, agrees in the greater part with the resuhs reported in the literature. SUMMARY I. Animal experimentation conducted on 36 dogs indicates that an increase either in the number or the virulence of bacteria
26
American Journal of Surgery
Aronsohn-ChoIecystitis
in gaII-bIadder biIe using a non-traumatizing technique, seIdomIy produces a reaction in the gaII-bIadder waI1. A severe infection of the waI1 was obtained in 3 dogs, which had been injected with one particuIar strain of Streptococcus hemoIyticus. The gaII-bIadder waI1 remained practicaIIy unchanged, however, upon the injection of three other strains of Streptococcus hemoIyticus, as we11 as BaciIIus coli, Streptococcus viridans, Staphylococcus aureus and aIbus, and BaciIIus WeIchii. 2. In many cases the same organisms injected directIy through the gaII-bIadder waI1, or with an accompanying Iigation of the common or cystic duct, produce severe or even fata choIecystitis. CONCLUSION
The resuIts seem to confirm the earIier statement of Graham that choIecystitis of biIiary origin is diffrcuIt to produce without the additiona factor of trauma or stasis. REFERENCES I.
2.
3. 48
5. 6. 7.
ANDREWS, E. and HENRY, L. D. The bacteriology of norma and diseased eaII bladders. Waitine publication in the Arch. Iit. Med. MAGNER, W. and HUTCHESON,G. M. ChoIecystitis. Can. Med. Assn. Jour. 27: 469, 1932. OSLER. Modern Medicine. PhiIadeIphia, Lea and Febiger, 1907. ARONSOHN, H. G. and ANDREWS, E. Bile saIt ChoIecystitis: PreIiminary report. Proc. Sot. Exp. Biol. and Med. In press. GRAHAM. Diseases of the GaII BIadder and BiIe Ducts. PhiIadeIphia, Lea and Febiger, 1928, I 14. HUNTER, JOHN. cited in RoIIeston, Diseases of Liver, GaIIbIadder and BiIe Ducts. GILBERT and GIRODE. Compt.-rend. Sot.’ de biol., Paris, p. 756, 1890; ibid. p. 956, 1893.
APRIL,1936
22, 1893. 8. CHIARI. Prag. med. Wcbnscbr., No. Ztscbr. f. Heilk., Bd. xv, p. 199. 9. BUCKSTEIN. Bull. Johns Hopkins Hosp., pp. g6 and 121, 1891. i, 10. SHERRINGTON, T. PathoIogy and BacterioIogy, P. 259, 1892-93. II. GILBERT ET DOMINICI. Compt.-rend. Sot. de biol., p. 38, (Jan. 20) 1893. 12. CUSHING and LIVINGOOD. Jobns Hopkins Hosp. Rep. 9: 543, 1900. ‘3. CUSHING, H. Bull. Johns Hopkins Hosp., IO: 166, 1899. 14. ITALIA, F. E. Policlinico, 1901; cited Zentralbl. f. Cbir. 28: 693, 1901. ‘5. ROLLESTON. Diseases of the Liver and Bile Ducts. rgo5 and 1929. 16. OSLER, WILLIAM. Practice of Medicine, 1905, p. 83. 17. ROSENOW. E. C. J. A. M. A. 61: 1835, 1914; and -_- . _Jour. of Inj. Dis. 19: 527, rgr6. Experimental choIera carriers. 18. SCHBBL, Orro. Jour. of Znj. Dis. 18: 307, 19x6. 19. DOERR: Zentralbl. j. Bakt. 39: 624, 1905. 20. KOCH, T. Ztscbr. .f. Hyg., I: I, 1908. f. Hyg.: I : I I, 1908. 21. CHIAROLANZA, R: Z&r. 22. NICHOLS, H. T. Jour. Exp. Med. 24: 497, 19x6. 23. EMMERICHand WAGNER: Zentralbl. j. allg. Path. u. patb. Anat. 27: 433, 1916. 24. VENEMA, T. A. Berl. klin. Wocben. 815, 1917. 25. MEYER, K. F., Nelson, N. M. and FENSIER, M. L. Jour. Inj. Dis. 28: 457, 1921. 26. BROWN, R. 0. Arch. Int. Med. 23: 185, 1919. 27. HUNLEMOLLER.Klin. Wocben. iii: 349, 1924. 28. DRENNAN, T. G. Ann. Surg. 76: 482, 1922. 29. DENTON, T. Arch. of Surg. 14: i, 1927. 30. FEINBLATT, H. M. New EngIand Med. 199: 1073, 1928. 31. WILKIE, A. L. Brit. Jour. of Surg. 15, 1927-28. 32. PHEMISTER, D. B. Calcium carbonate gaItstones and their experimenta production. Ann. Surg. P. 595, (Oct.) ‘932. 33. GRAHAM, E. A. and PETERMAN, M. G. Arch. Surg. 4: 23, 1922. 34. HOSPERS, C. A. Arch. of Patb. 14, 66-78, (JuIy)
1932. 35. ARONSOHN,H. G. and ANDREWS, E. A non-traumatizing technique for study of chemica1 choIecystitis. Proc. Sot. Exp. Biol. and Med. 32, 16291631, 1935.
STUDIES OF BACTERIAL HANS G.
From the Department
ARONSOHN, of Surgery,
CHOLECYSTITIS *
M.D.
University
of Chicago
CHICAGO
B
OTH Andrews’ and Magner,2 studying the contents of human gaII bIadders removed at operation, have found 65 per cent to contain sterile biIe. 0sIer3 pIaces this figure at 52 per cent. An important roIe in the production of cholecystitis seems to be pIayed by factors other than infection, mainIy those of a chemicaI nature. Recent investigations conducted by us4 have shown that bile acids, which Iong have been known to be of high toxicity for tissues outside of the gaI1 bIadder, deveIop a definite toxicity for the gaIl-bladder waI1 itseIf if present in a suffrcientIy high concentration. Further studies are now in progress on the exact role pIayed by the biIe acids. AIthough the importance of infection in this connection is possibIy not primary, its existence as a factor cannot be compIetely disregarded for severa reasons. First, one is tempted to draw an anaIogy as regards the gaI1 bIadder from the fact that infection plays an important part in the pathology of most other organs. Secondly, one must take into account the fact that many reIiabIe workers, afterextensive studies, have concIuded that infection is the main factor in the production of cholecystitis. Very IittIe is known about the actual route which the infection foI10~s. The five possibihties, as given in Graham’s textbook,5 are as foIIows: (I) descending infection from the Iiver; (2) ascending from the duodenum via the common duct; (3) hematogenous; (4) Iymphatic; (5) directIy through the galIbIadder waI1 from an inflamed adjacent organ. To determine the route by which the infection spreads is not the essentia1 point of this study. Our purpose is to ascertain by anima1 experimentation whether or not *Work
an increase either in number or viruIence of the bacteria in bile Ieads to choIecystitis, and whether different bacteria show different behaviors in this regard. A considerabIe number of bacteria may be present in norma biIe, but since these gaI1 bIadders do not manifest any changes either chnicaIIy or anatomicaIIy, we are not justified in caIIing such biIe “infected.” BiIe may be considered as “infected” if it contains either an abnormaIIy high number of bacteria or bacteria of especiaIIy great viruIence. The organisms might be those which occur normaIIy or might accrue in the biIe from any pathoIogica1 sources in the body. Any bacterioIogica1 study of the biIiary system presupposes knowIedge of the norma bacterioIogica1 ffora of the Iiver and gaI1 bIadder as we11 as the kind and origin of pathoIogica1 organisms and the route by which these might reach the gaI1 bladder. Since our experimenta studies are based on certain fundamental factors which are considered to be proved, a brief review of the Iiterature is in place. The occurrence of cholecystitis in cases of typhoid fever Ied earIy investigators to consider the infectious nature of gaII-bIadder disease. As earIy as 1763, John Hunter6 associated inff ammation of the gaI1 bIadder with typhoid fever, an idea which was proved bacterioIogicaIly by GiIbert and Girode’ in 1890 and confirmed cIinicalIy by Chiari.8 There foIIowed a series of experimenta.1 studies which attempted to produce “carriers ” or even reactions of the gaI1 bIadder, by injection of typhoid baciIIi. BIacksteing found the biIe to contain bacteria for many days or weeks foIIowing the intravenous injection of BaciIIus coli and Bacillus typhosus in rabbits. Other
done in part under a grant from the DougIas Smith Foundation. 18
NEW SEHES VOL. XxX11,
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I
Aronsohn-ChoIecystitis
bacteria were subsequentIy used by further observers, Sherringtonl” injecting BaciIIus anthracis, BaciIIus pyocyaneus, and staphyIococci and obtaining I 8 “infected ” biles from 49 cases. Whether or not the gaIIbIadder waI1 showed any pathology in these experiments couId not be ascertained from the data given in the Iiterature, since the chief interest of the author was directed towards the production of “carriers.” GiIbert,l’ in I 894, considered BaciIIus coIi to be the most important organism in the production of cho1ecystiti.s and favored the idea of an ascending infection from the duodenum. This view was opposed by Cushing and Livingood,12 who found the duodenum to be very Iow in bacteria1 content and even steriIe when empty. Further contributions were made whiIe attempting to produce stones experimentaIIy. Cushing,13 after direct inocuIation of typhoid baciIIi into the gaI1 bIadders of rabbits, observed inflammation of the gaI1 bladders with the formation of concrements. SimiIar experiments were performed on dogs and rabbits by ItaIia,14 who used attenuated cuItures of BaciIIus coIi and BaciIIus typhosus. Th is author mentions an inflamed gaII-bIadder mucosa, incidental to the formation of concrements. RoIIeston15 emphasizes the need for an additiona factor other than the presence of bacteria, such as trauma, stones, or stasis, for the production of choIecystitis. This author points out that though the frequency of typhoid fever is decreasing (BaciIIus typhosus is found in the biIe in aImost a11 severe cases of typhoid fever), there is an increase in the frequency of choIecystitis. SimiIarIy, OsIerlfi states that choIecystitis in typhoid fever occurred in onIy 19 of his series of 1500 cases. Later workers again considered the bIood stream to be the route by which bacteria reach the gaI1 bIadder and produce choIecystitis. There was great disagreement, however, as to whether the organisms reach the gaI1 bIadder directIy through the arteria1 bIood stream or indirectly by way of the porta vein, Iiver, and duct system. Rosenow, l7 who demonstrated the
American
Journal
of Surgery
19
specific affinity of streptococci and partIy aIso of BaciIIus coIi to the gaI1 bIadder, considers the direct arteria1 route to be the
Diagram: Showing carmula and gaII bIadder after the common duct had been opened and the catheter introduced into the gal1 bladder.
usua1 mode of infection, since he obtained positive reactions by intravenous bacteria1 injection and negative results by direct inocuIation into the gaI1 bIadder. SchiibI,‘* on the other hand, who tried to produce “cholera carriers” experimentally, obtained the opposite resuIts, direct inocuIation into the gaI1 bIadder being more effective than intravenous injection. There is no mention in this work whether or not a change occurred in the gaII-bIadder waI1. Doerrlg observed that excretion of typhoid baciIIi into the biIe, which occurred reguIarIy after injection of the former into the systemic veins, could be prevented by Iigation of the cystic duct. On the basis of this experimenta work he concIuded that bacteria indirectIy reach the gal1 bIadder by excretion through the Iiver. Koch20 and ChiaroIanza,21 however, arrived at the opposite concIusion. Typhoid baciIIi were found in the gaI1 bIadder after intravenous injection even after a preceding Iigation of the cystic duct. This observation, together with the fact that microscopic study of a large number of inflamed gaI1 bIadders reveaIed “emboIi” in the
20
American Journd of Surgery
Aronsohn-ChoIecystitis
foIds of the mucous membrane, Ied these authors to con&de that “a transverse infection of the waI1” is the usua1 route. NichoIs22 concurred with the hematohepatogenous route of Doerr on the basis of his work on rabbits. This author found that injection of typhoid baciIIi into the mesenteric veins yieIded “gaI1-bIadder Iesions ” in 63 per cent of the cases, whiIe injection into the ear vein resuIted in onIy 4 I per cent of positive cases. Emmerich and Wagner,23 after direct injection of typhoid baciIIi into the gaI1 bIadders of rabbits, obtained a chronic infiItration and thickening of the gaIl-bIadder waI1. However, the fact that these animaIs had been immunized with typhoid serum previous to the injections might have sIackened and diminished the inffammatory reaction. In contrast to these observations, Venema,24 in simiIar experiments, obtained a norma gaI1 bIadder. In rgzr Meyer25 and his co-workers confirmed the vaIidity of the hematohepatogenous route, which they beIieved to be the onIy one proved definiteIy in the Iiterature, by observing positive reactions in the gaI1 bIadders of rabbits both by intravenous injection of typhoid baciIIi and direct inocuIation of the same organism into the gaI1 bladder. Rosenow’s theory of a hematogenous infection was further enhanced by Brown,26 who found mainly streptococci in the emuIsihed gaI1 bIadder tissue from human cases of ChoIecystitis. This author demonstrated seroIogfcaIIy the direct etioIogica1 importance of these streptococci and demonstrated their specific affinity for the gaI1 bIadder by intravenous injection in rabbits. The very compIete survey by Magner and Hutcheson reveaIed that the most common organism in the cavity of the gaI1 bIadder is BaciIIus coIi, whiIe in the waI1 it is streptococcus. These workers injected staphyIococci into both the waI1 and cavity of rabbits’ gaI1 bIadders and in every instance of subserous injection obtained positive reactions in the gaIIbIadder waIIs, but in only a limited number of cases foIIowing injection into the Iumen. examining gaI1 bIadders HunIemiiIIer,27
APRIL, 1936
obtained at human operations, revealed positive bacteria1 cuItures in IOO per cent of cases when he incIuded in his statistics both the gall-bIadder waI1 and the Iiver. During the Iast ten to fifteen years the beIief in the infectious origin of choIecystitis has become considerabIy weakened. Wherever the infectious theory is stiI1 maintained, the route is considered to be especiaIIy by way of the Iymphatics from the Iiver, as suggested by Graham. In 1922, Drennan,28 who emphasized the antibacteria1 activity of the biIe, found onIy rg of IOO biIes removed at operation, to be infected. The scarcity of cIinica1 signs of infection, the Iower fever and sIight Ieucocytosis, as we11 as the rare occurrence of Ieucocytes in biIe even in cases of so-caIIed “empyema,” based upon a study of a Iarge amount of surgica1 and necropsy materia1, convinced Denton2g that infection does not pIay a primary roIe in the production of ChoIecystitis but that certain other factors, such as mechanica or chemica1 ones, are the basic causes. FeinbIatt,30 in a study of 400 cIinica1 cases, obtained resuIts similar to Drennan and Denton, observing very few inffammatory changes in the gaIIbIadder waI1 and very few Ieucocytes in the biIe or ffuid content. From this brief review of the Iiterature, one can see the present confused state of the question of the relationship between the bacteria1 content of the gaI1 bIadder and the pathoIogy of that organ. The information yieIded by different authors varies considerabIy and often is contradictory. WhiIe bacteria1 studies of the gaI1 bIadder obtained at human operations and postmortem examinations are reIiabIe and vaIuabIe, experimenta studies of the same probIem are much Iess satisfactory, due to the differences in the techniques empIoyed by various investigators, some of which may be actuaIIy misIeading. The commonest method, introduction of the bacteria by means of a needIe puncture through the gaII-bIadder waI1, often resuIts in a IocaIized injury to the wal1 at the point of puncture, thus making the resuIts uncon-
NEW SERIES VOL. XxX11,
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I
Aronsohn-Cholecystitis
trollabIe. This disadvantage was recognized by many earIy workers, Rosenow stating that “localized cholecystitis occurred at the point of puncture when the injection was made directIy into the gaII-bIadder.” SimiIarIy Venema mentions the fact that the infection produced by the usua1 technique, which consisted of needIe puncture with subsequent tying off of that part of the wall, is especiaIIy marked and is accompanied by a severe injury to the gaI1 bladder waI1. According to WiIkie,3’ the changes at the neck of the gaI1 bIadder, where the injection had been made, contrasted sharpIy with the fundus of the organ, which stiI1 appeared normal. The work of Phemister32 and Graham,33 however, indicates that it is possibIe to obtain normal gaI1 bIadders even with this usuai technique, if the proper degree of care is administered. In experiments with dogs, Phemister injected Streptococcus viridans intramuraIIy and performed in addition a cystic duct Iigation, yet the gaI1 bladders remained norma in a11 but one case, where a subsequent mixed infection accounted for the marked inflammatory changes. SimiIarIy, Graham and Peterman obtained negative resuIts folIowing the injection of BaciIIi coIi into the waI1 of the gaI1 bIadder in dogs. In this instance, however, additiona Iigation of the bIood vesseIs or cystic duct resulted in definite changes. Some improvement of the needIe puncture technique was made by Venema, who cauterized the waI1 at the point of the injection. It is perhaps to this improved technique that we can attribute the fact that this author obtained normal gaI1 bIadders after direct inocuIation of typhoid baciIIi, while simiIar experiments by Emmerich and Wagner yieIded positive reactions. Hospers34 varied the procedure by making the injection through the Iiver surface of the gaI1 bIadder, a method which Ieaves the free surface of the gaI1 bIadder practicaIIy unharmed, as we have seen, but may cause occasiona bIeeding in the liver tissue and thus inffuence the adjacent gaI1 bladder. The method of intravenous
American Journal of Surgery
21
injection practiced by Rosenow and many others is much more satisfactory. A positive reaction of the gaI1 bIadder foIIowing an intravenous injection of bacteria may be attributed with certainty to a specific affinity of these organisms to the biIiary system. Negative resuIts are vaIueIess in this connection. AIthough the organisms may not reach the gal1 bIadder via the blood stream, they may stiI1 possess a definite affnity for the biliary system if brought there by any other way. Another disadvantage of intravenous injection is the production of a bacteremia, which ordinariIy is not present in cases of human ChoIecystitis, and might faciIitate the production of choIecystitis by causing an abnorma1 reaction of the organism as an entity. It is now evident that there is great doubt as to which route bacteria actuaIIy foIIow, under either norma or pathoIogica1 conditions, which are found eventuaIIy in the gaI1 bIadder. ConsequentIy we wiI1 Iimit ourseIves to the effects of the actual presence of organisms in the gaI1 bIadder either of especiaIIy great viruIence or in abnormaIIy high numbers. This condition can be created experimentaIIy by means of technique.35 We our “nontraumatizing” empIoyed this procedure in most of our experiments, using the method of injection by a fine needIe puncture in onIy a few instances, which served to compare the two techniques. Method. Only dogs were used in our experimenta work, the operations being performed under ether anaesthesia and strictIy aseptic conditions. The abdomen was opened by a midline incision and the common duct incised close to the duodenum. A soft French rubber catheter was introduced into the gaI1 bIadder and IO to 13 C.C. of biIe were aspirated by means of a syringe. Injection of the bacteria1 suspension was accompIished at this point by means of another syringe, and this was foIlowed by re-injection of the aspirated biIe into the gaI1 bIadder, thus washing the common and cystic ducts as free as possibIe
22
American Journal of Surgery
Aronsohn-ChoIecystitis
from bacteria. It might incidentahy be mentioned at this point that this washing was suffrcientIy effective to prevent any marked infection of the mucous membrane of the ducts. The catheter was then withdrawn and a gIass cannuIa inserted into the opening which had been made in the common duct. By means of this technique, the gaI1 bIadder in most instances was not visuaIized during the entire procedure, practicaIIy assuring freedom from traumatization. In those instances in which we failed to introduce the catheter into the gaI1 bIadder, we injected the bacteria1 suspension by means of a direct needle puncture and rn addition performed a Iigation of the cystic or common duct. For this injection, we preferred aIways to use the Iiver surface of the gal1 bIadder rather than the free surface, thus minimizing the amount of trauma to the gaII-bIadder waI1. In severa cases a needIe puncture was used for the injection without any duct Iigation, to determine whether or not stasis of the biIe favors infection of the gaIl-bIadder waI1. The simpIe catheter-cannuIa technique is the onIy method that has yieIded norma controIs. It has aIways been a handicap in gaI1 bIadder studies that if injection was made through the gaII-bIadder waI1 a considerable degree of choIecystitis might arise and furthermore if the duct was Iigated or drained externaIIy, the contro1 experiments very frequentIy were not normaI. Therefore we have never been abIe previousIy to either remove or introduce materia1 into the gaI1 bIadder without causing some degree of damage, which vitiates the experiment. The diagram shows the cannuIa in pIace after the common duct had been opened and the catheter introduced into the gaI1 bIadder. The bacteria used for injection were obtained from human sources, as acute adenitis, sinusitis, acute and chronic mastoiditis, etc. The cuItures were of recent isoIation. The organisms were suspended in saIine soIution, in a concentration of approximateIy 500 miIIion bacteria per c.c.,
APRIL, ,936
the criterion used being the cIoudiness of the suspension as compared with one of known bacteria1 content. One C.C. of this suspension was generaIIy the amount injected. Wherever a marked reaction occurred, the experiment was repeated with 0.5 C.C. or Iess, whiIe 2 C.C. were used in those cases in which there was no reaction. The dogs were eIectrocuted twentyfour hours after the operation and postmortem examination performed as soon as possibIe. A microscopic examination of the gaII-bIadder waI1 was made in every case except those which died SpontaneousIy and where postmortem changes had occurred. Those animaIs which died from any other cause, as Ieakage from the gal1 bIadder or ducts, pneumonia, etc. were not incIuded in this study. Protocols. In aI1, 36 dogs were operated. In Charts I, II and III are shown the detaiIed resuIts of the experiments. A gaI1 bIadder was considered to be norma if it reveaIed no changes either on gross or microscopic examination. “ Cholecystitis I ” indicates that the gall-bIadder waI1 was of normal thickness, but there was some edema and congestion in the serosa. A fibrinous membrane was either compIeteIy absent or seen 0nIy in pIaces. “ChoIecystitis 2 ” incIuded those cases in which the gaII-bIadder waI1 was thickened up to 4 mm. and the inAammatory signs were more definite, the fibrinous exudate covering the serosa as a thick membrane. “Cholecystitis 3 ” indicates that the waI1 is thicker than 4 mm., the gaI1 bIadder usuaIIy Iies imbedded deepIy in the Iiver, and the adjacent organs, the duodenum, omentum, etc. form a Iarge inffammatory tumor around it. The terms “about normaI” or “sIight reaction” were used in those cases in which the changes were non-inflammatory in character, i.e., distention of the gaI1 bIadder, congestion of the serosa without any edema or fibrinous exudate, mechanica injury to the waI1, etc. Whenever the introduction of the catheter or insertion of the cannuIa was attended with diffrcuIty, the term “trauwas incIuded in the column matization”
NEW SERIES VOL. XxX11,
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Aronsohn-ChoIecystitis
I
Wherever this IabeIIed “comphcations.” happened the value of the experimenta resuIts are Iimited, as they are in those instances in which the animals died spontaneousIy and postmortem changes occurred in the gaII bladder. SimiIarIy, experiments in which the common or cystic duct was tied are of vaIue onIy when the
American Journal of Surgery
23
within fifteen hours after the injection, the second was moribund in twenty-four hours, and the third was in poor condition when kiIIed. OnIy three drops had been injected into the anima1 which died. These resuIts are in direct contrast to the experiments on nine dogs in which the other three strains of Streptococcus hemoIyticus were injected.
CHART I STREPTOCOCCUS
Strain
Exp’t. No.
Strain
Injected
Amount
HEMOLYTICUS
from: (I) Acute mastoiditis 1-3 dogs (2) Acute sinusitis--6 dogs (3) Acute mastoiditis 11-2 dogs (4) Chronic mastoiditis--r dog Method of Introductior
1
Duct
Complications
ResuIts
-.
9 IO II I2
Acute mastoiditis
I
0.5 cc.
Catheter
CannuIa
Acute mastoiditis
I
3 drops
Catheter
CannuIa
Acute mastoiditis Acute sinusitis Acute sinusitis
I
3 drops
Catheter Catheter Catheter
CannuIa CannuIa CannuIa
2 cc. I cc.
Acute sinusitis Acute sinusitis Acute sinusitis
2 cc.
Acute sinusitis
I cc.
Acute mastoiditis Acute mastoiditis Chronic mastoiditis
I cc. I cc.
II
I cc.
II
I cc. I cc.
Catheter CannuIa Catheter CannuIa Common NeedIe punt :Iigation ture Catheter CannuIa Catheter Catheter Catheter
gaI1 bladder remained norma in spite of the bacteria1 injection and duct Iigation. Results. Since streptococci and BaciIIus coIi are the organisms most commonly accused of being the causative agents in the production of gaII-bIadder infection, we pIaced specia1 emphasis in our experimenta work on the actua1 effect which these bacteria have. Very striking resuIts were obtained with Streptococcus hemoIyticus, the detaiIs of which appear on Chart I. Four different strains of the organism were used for the injections. AIthough three of the four strains used were from acute human infectious processes, onIy one produced changes in the gaI1 bIadder. The changes were very marked and occurred in the 3 cases in which this particuIar strain was injected. One of these animaIs died
CannuIa Cannula CannuIa
Dying when electrocuted Died Very sick
ChoIecystitis 3, biIe peritonitis ChoIecystitis 3, postmortem changes ChoIecystitis 2-3
I 1: 1: 1: 1: 1: 1: t$ZrnIZ%E duct
!
( ““““” i :::::::::::: ~
n~E.t~
GaII bIadder norma ChoIecystitis I Thickening of galIbIadder wall Gal1 bIadder about norma Gal1 bIadder norma GaII bladder norma Gal1 bIadder norma
None of these animaIs died spontaneously nor did any of them exhibit any marked signs of iIIness during the twenty-four hours between the injection and eIectrocution. In 5 of these g cases, postmortem examination reveaIed perfectIy norma gaI1 bIadders, whiIe in 3 other cases the changes were very sIight and not inflammatory at aI1. In only one instance, that of dog No. 7, did a shght but definite cho1ecystiti.s deveIop foIIowing injection of the Streptococcus hemoIyticus from an acute sinusitis, and the technique was not accountabIe for this change. The injection of BaciIIus coIi yieIded definiteIy negative resuIts, which are summarized on Chart II. In 3 cases in which the catheter method had been empIoyed the gaI1 bIadder remained perfectly normaI.
American
24
Journal
of Surgery
Aronsohn-ChoIecystitis
In 4 animaIs in this group (4, 6, 7 and 8), the shght changes manifested couId definitely be attributed either to traumatization at the time of operation or to the Iigation of the cystic or common duct. In none of these were the changes so marked that they couId be attributed to the bacteria1 injection. OnIy one gaII bIadder (dog No. 3) reveaIed an intense reaction, apparentIy due to the ligation of the common duct. From Chart III it can be seen that the results Iikewise are negative. None of the organisms caused any inffammatory changes on the gaII-bIadder wall and any sIight change which occurred couId easiIy CHART BACILLUS
Exp’t. NO. I
Bacteria Injected
Amount
3 4 5
B. B. B. B. B.
CoIi Coli Coli Coli Coli
2 C.C.
6
B. CoIi
I C.C.
7
B. CoIi
I C.C.
8
B. Coli
1.5 C.C.
2
I C.C. I C.C.
I C.C. I C.C.
Method of Introduction
CannuIa Cannula CannuIa CannuIa Common ligation
Needle puntture NeedIe puntture Needle puntture
Common ligation Common ligation Cystic Iigation
be attributed to traumatization or duct Iigation. The charts show that when the same type of bacteria is injected through the catheter the gal1 bIadder remains normaI, but if injected by needle puncture, or if the cystic or common duct Iigation is added, then they cause cholecystitis or death. Microscopic examination of the gaI1 bladder in the experiments where streptococcus hemoIyticus isoIated from acute mastoiditis cases had been injected, reveaIs that the enormous thickening of the waI1 is caused chiefly by a swehing of the serosa1 Iayer to about five times its normaI thickness. The Iatter is edematous and hyper-
1936
emit, contains a Ieucocytic infiltration, and is covered with a puruIent fibrinous exudate. Although there is a scattered Ieucocytic invasion of the muscuIar layer, this as well as the mucosa1 surface is mostIy unchanged. Those gal1 bIadders, however, into which had been injected another strain of Streptococcus hemoIyticus as we11 as Staphylococcus, BaciIIus WeIchii and BaciIIus coli, proved to be practicaIIy norma both grossIy and microscopically. In summarizing these experimenta resuits, we may conclude that there is IittIe tendency for an acute bacteria1 infection of the gaIl-bladder waI1 to resuIt from the contents of that organ. OnIy Streptococcus II COLI
Duct
Catheter Catheter Catheter Catheter Needle puntture
APRIL.
Complications
Results
. ..,
duct
Some traumatization Some traumatization died
duct
Very sick
duct duct
GaII bladder norma Gall bIadder norma GaII bIadder normal ChoIecystitis I Cholecystitis 3, postmortem change, bile peritonitis DiIated gal1 bIadder with fibrinous exudate Gal1 bIadder about norma1 Slight reaction on gal1 bIadder
hemoIyticus gave a positive reaction, and this onIy in the case of one of the four strains used. Since traumatization or extension from an adjacent pathologica process can be definitely ruIed out as a cause for this change, we may deduce that the organism or its toxins present in the biIe reached the serosa1 Iayer of the waI1 by traversing the mucosa1 and muscular Iayers without causing any changes in these. None of the other organisms injected, aIthough present in the biIe in overwheIming numbers, caused any change in the gaII-bIadder wall. The experimenta resuIts we obtained partIy confirm and partly contradict those reported in the literature. We found bacteria1 infection of the gaII-bIadder waI1
Nrw
%nlts
VOL. XxX11,
Aronsohn-Cholecystitis
No. I
occur Iess frequently than recorded by other authors. AI1 observers have described the streptococcus as pIaying an important part in producing inflammation of the gallto
bladder waII. The diverse results we obtained with different strains of this organism concur with the observations of Rosenow and Magner and Hutcheson, who stressed the varying activity within this group on the basis of differences in viru-
Bacteria Injected
,\lethod of Introduction
Amount
_~
.I
Staph. Staph. Staph.
albus albus albus
16
I
cc.
Catheter Needle puncture NeedIe puncture
I
cc. cc.
Catheter Catheter
Cystic ligation CannuIa Cannula Cannula Cannula
2 cc.
Catheter Catheter
Staph. aureus
2 cc.
Needk
Staph. aureus
I cc.
Catheter
Strep. viridans
I cc.
Catheter
Common ligation Cannula
Strep. viridans
I cc.
Catheter
Cannula
Catheter Catheter Catheter Catheter
Cannula CannuIa Cannula Common ligation
1 B. Welchii
0.3
cc.
I cc.
0.5
Traumatization
., Traumatization
puncture
NeedIe puncture
cc.
duct
Common ligation
-
Ience. Magner and Hutcheson divided the streptococci into two chief groups, which they classified as typical and atypical: TypicaI streptococci were characterized by the formation of smooth, non-hemolytic, colorless or green coIonies on blood agar, and by the fact that they were kilIed by exposure to the action of bile. AtypicaI streptococci were characterized by the formation of rough, nonhemolytic, white colonies on bIood agar, and by
duct
Died
I ( , /:‘.:::...:::I duct
_.___
Gall bIadder norma Gall bladder norma Slight reaction on gaI1 bladder Gal1 bladder norma Gall bladder about normal Gal1 bladder normal BiIe peritonitis, some reaction on gall bIadder SIight reaction on gal1 bladder Cholecystitis 3, postmortem changes GaII bladd er about normal normal, Gall bladder some bile peritonitis GaII bladder normal Gail bladder normal GaII bladder norma Postmortem change on gaII bladder
I
Staph. aureus Staph. aureus
I cc.
Results -___-.
---
Cannula
1
2 cc.
I_-
I
Staph. aureus Staph. aureus
Strep. viridans B. Welchii B. Welchii B. WeIchii
Complications _~.
cc. cc.
2 cc.
dogs dogs 3 dogs
Duct I ____~.
1
I
23
The negative resuIts we obtained using Bacilli coli stand in direct disagreement both with the clinical data reported by GiIbert and the experimental work done with this organism by Italia. However, the fact that
Staphylococcus aureus--6 (3) Streptrococcus viridans(4) Bacilli Welchii-4 dogs
1
of Surgery
gall bladder.
(2)
-
Journal
the fact that they were not killed by the action of bile. Chronic cholecystitis was produced in 100 per cent of cases by injecting typica streptococci under the serous coat of a rabbit’s
CHAKT III (I ) Staphylococcus albus-
Esp’t. No.
American
: Traumatizxtion died
duct
-I
Gail bladder mal
about
nor-
StaphyIococcus aureus and albus, Streptococcus viridans and BaciIIus Welchii, a11 failed to produce cholecystitis, agrees in the greater part with the resuhs reported in the literature. SUMMARY I. Animal experimentation conducted on 36 dogs indicates that an increase either in the number or the virulence of bacteria
26
American Journal of Surgery
Aronsohn-ChoIecystitis
in gaII-bIadder biIe using a non-traumatizing technique, seIdomIy produces a reaction in the gaII-bIadder waI1. A severe infection of the waI1 was obtained in 3 dogs, which had been injected with one particuIar strain of Streptococcus hemoIyticus. The gaII-bIadder waI1 remained practicaIIy unchanged, however, upon the injection of three other strains of Streptococcus hemoIyticus, as we11 as BaciIIus coli, Streptococcus viridans, Staphylococcus aureus and aIbus, and BaciIIus WeIchii. 2. In many cases the same organisms injected directIy through the gaII-bIadder waI1, or with an accompanying Iigation of the common or cystic duct, produce severe or even fata choIecystitis. CONCLUSION
The resuIts seem to confirm the earIier statement of Graham that choIecystitis of biIiary origin is diffrcuIt to produce without the additiona factor of trauma or stasis. REFERENCES I.
2.
3. 48
5. 6. 7.
ANDREWS, E. and HENRY, L. D. The bacteriology of norma and diseased eaII bladders. Waitine publication in the Arch. Iit. Med. MAGNER, W. and HUTCHESON,G. M. ChoIecystitis. Can. Med. Assn. Jour. 27: 469, 1932. OSLER. Modern Medicine. PhiIadeIphia, Lea and Febiger, 1907. ARONSOHN, H. G. and ANDREWS, E. Bile saIt ChoIecystitis: PreIiminary report. Proc. Sot. Exp. Biol. and Med. In press. GRAHAM. Diseases of the GaII BIadder and BiIe Ducts. PhiIadeIphia, Lea and Febiger, 1928, I 14. HUNTER, JOHN. cited in RoIIeston, Diseases of Liver, GaIIbIadder and BiIe Ducts. GILBERT and GIRODE. Compt.-rend. Sot.’ de biol., Paris, p. 756, 1890; ibid. p. 956, 1893.
APRIL,1936
22, 1893. 8. CHIARI. Prag. med. Wcbnscbr., No. Ztscbr. f. Heilk., Bd. xv, p. 199. 9. BUCKSTEIN. Bull. Johns Hopkins Hosp., pp. g6 and 121, 1891. i, 10. SHERRINGTON, T. PathoIogy and BacterioIogy, P. 259, 1892-93. II. GILBERT ET DOMINICI. Compt.-rend. Sot. de biol., p. 38, (Jan. 20) 1893. 12. CUSHING and LIVINGOOD. Jobns Hopkins Hosp. Rep. 9: 543, 1900. ‘3. CUSHING, H. Bull. Johns Hopkins Hosp., IO: 166, 1899. 14. ITALIA, F. E. Policlinico, 1901; cited Zentralbl. f. Cbir. 28: 693, 1901. ‘5. ROLLESTON. Diseases of the Liver and Bile Ducts. rgo5 and 1929. 16. OSLER, WILLIAM. Practice of Medicine, 1905, p. 83. 17. ROSENOW. E. C. J. A. M. A. 61: 1835, 1914; and -_- . _Jour. of Inj. Dis. 19: 527, rgr6. Experimental choIera carriers. 18. SCHBBL, Orro. Jour. of Znj. Dis. 18: 307, 19x6. 19. DOERR: Zentralbl. j. Bakt. 39: 624, 1905. 20. KOCH, T. Ztscbr. .f. Hyg., I: I, 1908. f. Hyg.: I : I I, 1908. 21. CHIAROLANZA, R: Z&r. 22. NICHOLS, H. T. Jour. Exp. Med. 24: 497, 19x6. 23. EMMERICHand WAGNER: Zentralbl. j. allg. Path. u. patb. Anat. 27: 433, 1916. 24. VENEMA, T. A. Berl. klin. Wocben. 815, 1917. 25. MEYER, K. F., Nelson, N. M. and FENSIER, M. L. Jour. Inj. Dis. 28: 457, 1921. 26. BROWN, R. 0. Arch. Int. Med. 23: 185, 1919. 27. HUNLEMOLLER.Klin. Wocben. iii: 349, 1924. 28. DRENNAN, T. G. Ann. Surg. 76: 482, 1922. 29. DENTON, T. Arch. of Surg. 14: i, 1927. 30. FEINBLATT, H. M. New EngIand Med. 199: 1073, 1928. 31. WILKIE, A. L. Brit. Jour. of Surg. 15, 1927-28. 32. PHEMISTER, D. B. Calcium carbonate gaItstones and their experimenta production. Ann. Surg. P. 595, (Oct.) ‘932. 33. GRAHAM, E. A. and PETERMAN, M. G. Arch. Surg. 4: 23, 1922. 34. HOSPERS, C. A. Arch. of Patb. 14, 66-78, (JuIy)
1932. 35. ARONSOHN,H. G. and ANDREWS, E. A non-traumatizing technique for study of chemica1 choIecystitis. Proc. Sot. Exp. Biol. and Med. 32, 16291631, 1935.