Investigation Of Experimental Pathology And Bacteriology Of Vibrio Vulnificus Infections

OBJECT: Vibrio vulnificus is an invasive gram-negative bacillus that may cause necrotizing cellulitis, bacteremia, and/or sepsis. Although V vulnificus infection is uncommon, it is frequently fatal and is usually attributed to ingestion of raw shellfish or traumatic exposure to a marine environment. In this study,we tried to find the reason of infections through the alimentary canal with Vibrio vulnificus(Vv) seldom inducing obvious symptoms digestive system.And we also wanted to find the pathological and bacteriological characters of Vv infection to provide theoretic foundation for clinical therapy of Vv infection.METHOD:1.Biochemical and morphological characters of Vv(1.1758 strain)and virulence experiment:After anabiosis-culture of Vv(1.1758 strain),we observed its morphological characters under light microscope,scanning microscope and transmission microscope and morphological characters of its colony on different culture mediums; 20 KM mice(SPF) were divided randomly into 2 groups,study group(n=24)and control group(n=15). Vv bacterial fluid was inoculated into the abdominal cavity in study group and control group was injected into the peritoneal cavity. The tissue taken from blood,heart,liver,sleen,lung and kidney were observed under light microscope.2.Pathogenicity of Vibrio vulnificus infection in mice through the enteron:The animal model of Vibrio vulnificus infection through the alimentary canal was established by feeding Vv fluid not only under normal gastric acid condition but also under anti-acid condition. And positive control group was fed with sterile culture medium and vacant control group was fed with Vibrio cholera fluid. The tissue taken from oesophagus,stomach and intestines were observed under light microscope. The tissue taken from stomach were also observed under scanning microscope.And we also observed the morphologic characteristics of Vv in acid(close to the gastric juice) under 37°C.We regarded Vv existing in stomach and necrosis of gastric mucous membrane as standard to judge whether Vv in certain concentration has induced the injury of stomach to find critical concentration of Vv infection in stamoch. 3.The study of bacteriology and pathology on the infected tissue with Vv:The animal model of Vibrio vulnificus infection was established by making wounded skin exposed to a marine environment with Vv. We also established other 3 kinds of animal models,like traumatic tissue infected by Vv in non-marine environment, wounded skin exposed to a marine environment with no Vv and only skin wounded. We tried to find all kinds of bacteria in infected tissue , make quantitative bacteriology study on Vv infected wound and analysed the datas with statistics.RESULT:1. Vv(1.1758 strain) grew into mucoid colony cutured on solid medium and it could not ferment lactose.lt didn't have capsule on its surface.2. The injury of lung and kidney were most severe among the main viscera of mice when primary septice caused by Vv occur.The pathological changes of lung injury were characterized by a lot of abnormal red blood cells existing in the bronchus cavity and stroma, the cells of bronchiolar epithelium falling off into thecavity and extravasate appearing in the alveolus;The pathological changes of kiney were characterized by degeneration and necrosis of the epidermic cells of renal tubule and hemorrhage.3. We didn't observed typical symptoms of digestive system after Vv fluid was fed under normal gastric condition.The gastric mucosa was damnified with degeneration and necrosis of the epidermic cells, and the anamorphic and swoln Vv were found in the stomachs under microscopes in the study group. The culture of Vv from the tissues was negative in both of the study and the control group. And Vv in the acid were anamorphic and swoln,too. The concentration range of Vv inducing damages of gastric mucosa was 108cfu/L~ 109cfu/L.4.We didn't observed typical symptoms of digestive system after Vv fluid was fed under condition of anti-acid. But the injury of gastric mucosa was more obvious and severe.Besides this,we also observed the injury of intestines characterized by soakage of inflammatory cells and the degeneration of flesh fibre.5. Bacteria in the wound exposed to a marine evironment with Vv was more various than a terrestrial evironment without Vv. Both of them were mixed bacillus infection.6. Bacterial quantification of 2 Vv infection groups tended to increase with the time. The total count of bacterium and the count of W in infected tissue exposed to a marine evironment with Vv were larger and the speed of bacteria proliferation in it was fast,too.7. Four bacterial growth dynamics models were set up,i.e.:y(1)= e (6-9893+0-5672X)......................................................... (1)Y(2)= e <7-2212+?>......................................................... (2)lgY=3. 0154+0. 2506x ......................................................... (3)lgY=3. 3160+0. 1638x ......................................................... (4)FomularC 1 )and( 3 Represent bacterial growth dynamics models of Vv infection in the seawater group,and fomular (2) and (4) represent bacterial growth dynamics models of Vv infection in the physiological saline solution group.X represents time and Y represents the bacterial count in per gram tissue.8. Using the dynamics model, we found the bacterial count of Vv infection in the seawater group would reach the critical point for clinical infection in 7.98h and Vv infection in the physiological saline solution group would reach the same bacterial count 3.4h later.9. The results of Vv culture from blood of 7 mice in Vv infection in the seawater group were positive.And the different degree injury of main viscera of these mice were observed.10. The degree of tissue injury of Vv infection in the seawater groups tended to increase with the time and its injury was the most severe among four groups.The injury was characterized by fargoing necrosis, abscess and slow repair .And the pathological changes of 2 Vv infection groups were more severe than 2 groups without Vv infection.CONCLUSIONS:1. The test of lactose zymolysis couldn't be used as a definitive definitive to identification of Vv.2. Vv 1.1758 stain had powerful virulence but had a mild pathogenicity in gastric mucosa by enteron infection.3. The primary septicemia and/or multi- organ's dysfouction and failure caused by severe Vv infection were the critical reasons of body quick death.4. Vv couldn't survive in the acidic gastric juice naturally and it had a mild pathogenicity in gastric mucosa by enteron infection. And just this two key points were the reason that infections through the alimentary canal with Vibriovulnificus(Vv) seldom induce obvious symptoms digestive system which was different to other vibrio.5. The concentration range of Vv inducing damages of gastric mucosa was 108cfu/L~109cfu/L6. There were more severe tissue damage,pollution and ealier infection in skin wound exposed a marine environment with Vv. And the tissue damage was characterized by purulent inflammation and lytic necrosis. Early use of antimicrobial based on drug sensitivity test was efficacious and cleaning out the seawater in the wound early is also emphasized.