Lesson 25: Anaerobic Respiration PDF

Lesson 25 Lesson 25 Ecological groups of anaerobic bacteria: the main reasons of anaerobic...

Lesson 25 Lesson 25 Ecological groups of anaerobic bacteria: the main reasons of anaerobic respiration The production of energy requires The electron transport chain, where the majority of ATP is oxygen. formed, requires a large input of oxygen. However, many organisms have developed strategies to carry out metabolism without oxygen, or can switch from aerobic to anaerobic cell respiration when oxygen is scarce. ANAEROBIC RESPIRATION SBYJU'S During cellular respiration, some species of bacteria use an organic molecule as the final electron acceptor. Pro Ethanol cesses that that use an organic molecule to regenerate NAD+ s from NADH are collectively referred to as fermentation. Lactate < Co In contrast, some some species of bacteria an inor use ganic molecule as a final electron acceptor. Both meth ods are called anaerobic cellular respiration, where or 2x NAD 2x NADH ganisms convert energy for their use in the absence of 2x (ATP 4x (ADP) Oxygen. Alcohol and lactic acid fermentation are the two forms of anaerobic respiration. It is a less effective cellular Pyruvate 2x (ADP) 4x(ATP) process, but anaerobic respiration converts energy more Glucose quickly than aerobic respiration. Aerobic And Anaerobic Respiration Anaerobic respiration is favorable for those organisms which live in such places where there is little oxvoen (ps://can1.byJUS.com/swp-content'uploads/2019/04/Aerobic-And-Anaerobic Respiration-2,png) like soil and digestive tract. It should be noted that all forms of fermentation, except lactic acid fermentation, produce gas. The production of particular types of gas is used as an indicator of the fermentation of specific carbohydrates, which plays a role in the laboratory identification of the bacteria. Louis Pasteur during the 1850s and 1860s showed fermentation is initiated by living organisms in a series of investigations. He defined fermentation (incorrectly) as "Life without air", yet he correctly showed how specific types of microorganismscau se specific types of fermentations and specific end products. He has proved that life can exist in atmos Types of anaerobes phere free of oxygen and that such bacteria as clostridia can live in anaerobic conditions. Obligate anaerobic bacteria- Are those bacteria that can grow in the absence of free oxygen, but fails to multiply in the presence of oxygen Ecological groups of anaerobic bacteria: bacteria which on the surface of nutritionaly adequate solid media incubated in room are highly sensitive to the presence oxygen and of or in a CO2 incubator (containing 5-10% CO:). eg.- Bacteroides aerotolerant bacteria fragilis, Clostridium perfringens, C. novyi, Porphyromonas, There are two main group of anaerobic bacteria: aerotol Fusobacterium. erant anaerobes and obligate or strict anaerobes. Aerotolerant anaerobes- Anaerobic bacteria that will show limited or Aerotolerant organisms can survive in the presence of scanty growth on agar in room air or in a 5-10% CO: incubator, but oxygen (tolerate the presence of it), but they are anaero show good growth under anaerobic conditions. eg.- C. carnis, C. bic because they do not use it as a terminal electron ac histolyticum, C. tertium etc. ceptor. Aerotolerant bacteria die only after prolonged contact with oxygen. (htps:/alchetron.com/cdn/anaerobic-organisn-21 4ccdbe-c845-4b8b-bd29. Obligate (strict) anaerobes are immediately killed by ea373lc2985-resize-750,jpeg) Oxygen. Classification of angerobes Anaerobic conditions exist naturally in the intestinal tract of animals. Obli BACILLI COCCI gate anaerobes, mainly Bacteroidetes, represent a large fraction of the mi crobes in the human gut. GRAM SPORING PeprococcUs POSITIVE PeptostreptococcUs Clostridium spp. Another type of obligate anaerobe encountered in the human body is Clos NON SPORING tridium spp. Their ability to form endospores allows them to survive in the Eubacterium Propionibacterium presence of oxygen. GRAM Bacteroides Veillonella Classification of anaerobic bacteria NEGATIVE Prevotella The group of Gram-positive ecological anaerobic bacteria includes: cocci Porphyromonas Fusobacterium belonging to genera Peptococcus and Peptostreptococcus; the spore forming rods belonging to the genus Clostridium, and also the rods which (https://image.slidesharecdn.com/ be ravisanaerobicinfection4-17090407141 1/95/anaerobic are not spore-formers belonging to the genera Propionibacterium and Eu bacterium. Group of Gram-negative anaerobic bacteria includes cocci be infection-pp-7-638 jpg?cb=1504509288) longing to the genus Veilonella and the rods which don't form spores and belong to the genera Bacteroides, Porphyromonas, Prevotella and Fusobacteriun. Clostridia: main characteristics Clostridium, genus of rod-shaped, gram-positive, spore-forming bacteria, members of which are found in soil, water, and the intestinal tracts of humans and other animals. Most species grow only in the complete absence of oxygen (are obligate anaer obes); however, some species are aerotolerant. Clostridium spores, which are inactive (dormant)forms of the bacteria, are highly resistant to heat, desiccation, and toxic chemi Lesson 25 cals and detergents and enable bacteria to survive when environmental conditions are difficult. When conditions are favorable, cach spore germinates into an active bacterium. If endospores inside Clostridium cells are present, they are wider than cells, conferring bacterial cells a swollen end or spindle shaped appearance, which gives them their name, from the Greek "kAootp" or spindle. Pathogenic Clostridium species produce tissue-destructive protein toxins that contribute to disease manifestations. Clostridia: classification Clostridium contains around 250 species that include common free-living bacteria, as well as important pathogens. The genus belongs to the family Clostridiaceae and the order Clostridiales. In medicine clostridia are classified into two groups: clostridia which cause wound infections and Scientific classitication clostridia which cause enteric infections. The first group contains the pathogens causing anaerobic Domain: Bacteria wound infections, such as gas gangrene - perfringens, C. novyi (itwas called earlier C. oedemati C. Phylum: Firmicutes ens; these species names used as synonyms now), C. histolyticum, C. septicum and others; tetanus - Class: Clostridia the pathogen is C. tetani. The second group is composed of the types of C. perfringens which cause Order: Clostridiales food-poisoning toxic infections, clostridia causing botulism, C. botulinum and C. difficile the patho gen causing Clostridium difficile infection, Family: Clostridiaceae In 2016 the last species was transferred from the genus Clostridium to Clostridioides, thus giving it Genus: Clostridium the binonmial Clostridioides difficile. This new name reflects the taxonomic differences between this Prazmowski 1880 species and members of the genus Clostridium, while maintaining names Clostridium difficile and C. diff as synonyms. (https:/len. wikipedia.org wiki/Clostridium) Clostridium difficile infection (CDIor C-dif), also known as Clostridioides difficile infection, is a symptomatic infection making up about 20% of cases of antibiotic-associated diarhea. Complica tions may include pseudomembranous colitis, toxic megacolon, perforation of the colon, and sepsis. Clostridia: natural habitats Clostridium species inhabit soils, aquatic sediments and the intestinal tract of' animals, including humans. Clostridium is a nor mal inhabitant of the healthy lower reproductive tract of females. Their fermentation of organic compounds, large amounts of CO, and like sugars, H; as produces well as volatile organic com pounds like acetic and butyric acid, acetone and Metabolism of substrates like amino acids butanol. and fatty acids results in foul -smelling degradation products. Clostridia also have an extended range of extracellular enzymes that degrade large biological molecules in the environment into fermentable compounds. Clostridia: the ability to produce spores Endospores are the most important single feature of the genus Clostridium. Thus, whilst the pathogenesis of its pathogens is as cribed to the devastating toxins produced (neurotoxins, endotoxins and cytotoxins), it is their capacity to produce spores that lies at the heart of the diseases they cause. This is because spores play the pivotal role in the spread of infection (e.g., C. dificile) and in foodstuff contamination and food poisoning (e.g., C. botulinum and C. perfringens). The processes of spore formation (sporulation) and germination (return of the dormant spore to toxin-producing, vegetative cells), therefore, represent key inter vention points. Sporulation can be placed occurs as in the external in the bacterial environment, cell centrally, terminally and in (i.e. the large intestine. Tne spores are of at one of the edge of bacterial o spherical Seca oval va (between n the center of the rod and one of its edge). Clostridia: their resistance to the unfavourable conditions of environment The resistance of usual vegetative cells of clostridia to Clostridia of gas gangrene the unfavourable conditions of environ ment is similar to the resistance of ma jority of bacteria but the resistance of their spores is very high. Clostridia: the pathogenicity factors Main panogenicity factors of clostridia C perfringens Gram stain smear. in C.perfringens C.novyi are the n toxins produced by these Capsules can be bacteria. The toxins frequently possess seen. the enzymatic activity and can act not (https:/ only as bacterial toxins, but also as viru slideplayer.com/ lence enzymes. slide 12262107/72/ images/20/ of the clostridia Ctperfringenst- Main characteristics Characteristicsjpg) producing gas gangrene C. perfringens is the only one species of clostridia that is C. septicum C.histolyticyShared able to produce capsule. Although they lack flagella, C. perfringens bacteria are able to because their bodies are lined with glide across filaments surfaces from end-to (https:/h3.googieter0ZAieMCGvCJOASF.com/proxy/dUcA69eHaZobc2aXSTxiSjs -end. The specific name perfringens is derived from the INrQkGHehNeBEUH2šQPFEKNkinE5Jfrg NbykWdFVYDA) Lesson 25 Latin per (meaning "through") and frango ("burst"), referring to the disruption of tissue that occurs during gas gangrene. The action of C. perfringens on dead bodies is called tissue gas. It causes extremely accelerated decomposition, and cannot be stopped by normal embalming measures. C. perfringens is the most commonbacterial agent for gas gangrene. C. novyi (oedematiens) are the clostridia which have the largest size, and straight or slightly curved shape. This species comes in three types, labelled A, B, and a non-pathogenic typeC by the range of toxins they produce.Growth in culture distinguished proceeds through 3 stages: initial growth wherein no toxin is produced;vigorous growth wherein toxin is produced; and spore formation wherein endosporesare formed and toxin production decreases. Isolating and identifying C. novyi is difficult due to its extreme anaerobic nature. It is also fastidious and to culture, requiring the presence of thiols. C. septicum polymorphic rods arranged in the smears in chains. This species can cause gas gangrene, but unlike other Clos are tridium no trauma is necessary at the site of the infection. It is thought that the infection is established by hematogenous species spread from the gastrointestinal tract. Gas gangrene caused by Clostridium septicum is associated with colorectal cancer and other defects of the bowel. C. histolyticum can be plated on Zeissler plate agar, and appear as small, dewlike colonies of either rough or smooth morpholo gy, by a zone of weak hemolysis. These bacteria tend to surrounded in pairs or short chains and in wound smears closely resembles C. perfringens, but without the capsule. Cells are richly flagellate and very motile. Clostridium histolyticum produces large endosporesand are asaccharolytic and proteolytic. This species is difficult to culture becausegrowth is inhibited by sugars, and spores are not very heat resistant. The pathogenicity factors of clostridia producing gas gangrene Clostridium perfringens produces a C. perfringens Virulen ce Factors variety of virulence factors. The mech anism of action of these factors usually Virulence Factors Biologic Activity falls into one of three groups. Some of these virulence factors, such as the a toxin alp which is phospholipase toxin, C, Lethal toxin; phospholipase C (lecithinase); increases and the kappa toxin, which is a colla vascular permeability; hemolysin; produces necro genase, are enzymes that hydrolyze tizing activity substances essential to the integrity of B toxin Lethal toxin; necrotizing activity membranes or body structures. Major other € toxin Lethal toxin; permease Other virulence factors, such as the L toxin Lethal binary toxin; necrotizing activity;, adenosine beta, episolon, and iota toxins, act pri diphosphate (ADP)ribosylating marily on the vascular endothelium, causing increased capillary permeabil S toxin Hemolysin 0 toxin ity. Still others, such as the delta and Heat- and oxygen-labile hemolysin;cytolytic theta toxins, are essentially hemoly K toxin Collagenase; gelatinase; necrotizing activity sins. Theta toxin is similar in action Minor A toxin Protease and serologically related to streptoly u toxin Hyaluronidase sin O. v toxin Deoxyribonuclease; hemolysin;necrotizing activity Three of C. novyi produce six types Enterotoxin Alters membrane permeability (cytotoxic, entero toxins. he alpha-toxin is characterized toxic) as lethal and necrotizing. This toxin is Neuraminidase Alters cell surface oedematising. It acts by causing mor ganglioside receptors; promotes phological changes to all cell types capillarv thrombosis Cnovyi type Toxins (https://image.slidesharecdn.com/'clostridialtoxins-c-18033 1 094237/95/c S/clostridial-toxins clostridium-perfringens-and-clostridium-difficile-12-638,jpg?cb= 1522489408) alpha, gamma, delta, epsilon B alpha, beta, zeta especially endothelial cells by inhibition of signal transduction pathways, resulting in the breakdown of cytoskeletal structures. The cells of the microvascular system become C gamma spherical and the attachments to neighboring cells are reduced to thin strings. This re sults in leakage from the cap lea g o ocdema. The beta-toxin is characterized e MS HOrmaly produced by Ous types C. novyi the vari as hemolytic, necrotizing gamma-toxin is characterized as (https:/len. wikipedia. orgwiki/ lecithinase. The delta-toxin is characterized as oxygen labile hemolysin. The epsilon Clostridium novyi) toxin is characterized as lecithino-vitelline and thought to be responsible for the pearly layer found in cultures. The zeta-toxin is characterized as hemolysin. The major virulence factor produced by C. septicum is a-toxin, a pore-forming cytolysin. Pores formed by alpha toxin are ap proximately 1.5 nm in diameter, allowing the release of potassium ions, thus disrupting the ionic equilibrium within the cell. Clostridium histolyticum The alpha-toxin is the major toxigenic factor of C. histolyticum. Alpha-toxin produces five toxins. is a necrotizing, This toxin is secreted, as it is isolated from filtrates of C. histolyticum cultures. Additional but not hemolytic, toxin. ly, alpha-toxin is is readily inactivated by proteolytic enzymes. Only about 29% of C. histolyticum strains isolated from soil actual ly produce this alpha-toxin. The beta-toxin of C. histolyticum is a group of seven collagenases. Collagenases are zinc metallopro teases that cleave collagen and gelatin into small fragments. Beta-toxin plays a major role in the pathogenicity of C. histohyticum, due to its ability to destroy collagen fibers in the body and cause necrosis. Gamma-toxin is a cysteine-activated proteinase that digests hide powder, gelatin, and casein. It is not active against collagen. Delta- toxin is an elastase. Epsilon-toxin is an oxygen hemolysinsimilar to the 0-toxin of C. perfringens and the ö-toxins of C. septicum, and C. novyi. Gas gangrene: pathogenesis of the disease Gas gangrene is a highly lethal infection of soft tissue, caused by Clostridium species, with Clostridium perfringens being the Lesson 25 187 most common. This is synonymous with myonecrosis and is characterized by rapidly progressive gangrene of the injured tissue along with the production of foul PATHOGENESIS smelling gas. SPORES ENTER DEVITALISED TISSUE The Clostridia producing gas gangrene penetrate into the wound, multiply and secrete protein toxins which cause necrosis and decay of the tissue. The pathological reac tions are not accompanied by inflammation, but active GERMINATE processes of gas production deeply in the affected tissues BACTERIAL MULTIPLICATION A EXOTOXINS SYSTEMIC:.Disease gas gangrene 1.Entry 2. (ponetratig wound) doad Muliplication (in LOCALA From: Toins anaerobio isouo, MUSCLE CARDIAC Gaso0ckets condons NECROSIS+ GAs DEPRESSION. 3 Bloodvessel FORMATION MOF compression- Lipases Necrosis The principle scheme of the Gas gangrene pathogenesis MOF= Stage 2 multiorgan failure B Muscle necrosis and oxooxin further production and organismspread (https:/image.slidesharecdn.com'gasgangrene-160928204018/95/gas (Cpertringens and otherspecios) gangrene-8-638jpg?cb-1475095326) Edema Gas pockets Stage 3 and also development of intoxication are characteristic for the patho gangrene genesis of the infectious process caused by C. perfringens. If C. novyi is additionally involved into the infectious process will cause severe Bioodvessetconpresslon it edema and also intoxication. Stage 1 Organismsaccompany C. perfringens cause 80% to 90%, of gas gangrene cases, but other foregn penotrating otbjoct skin ormuscle species can cause infection. In order of prevalence, they are Clostridi um (40%), C. septicum (20%), C. histolyticum (10%), Clostridi novyi um bifermentans (10%), Clostridium fallax (5%), and C. sordelli. Dermis These organisms are in the soil and organic waste especially if con taminated with fecal material. Muscle Health care workers should suspect igas gangrene if anaerobic gram positive bacilli are present in a wound with necrosis of soft tissue and Bloodvessel muscle. The organisms produce a gas identifiable on x-ray or CT scans. Only about 5% of the wounds colonized with clostridial organ Pathogenesis Myonecrosis of (https://medchrome. comwp-content/uploads/2010/11/gas isms will develop an infection. Therefore, host factors and anatomic gangrene-pathogenesis,jpg) location of inoculation of the organisms help determine whether the bacteria will develop into a clostridial myonecrosis infection. For example, a deep penetrating wound into muscle tissue in which the host is immunocompromised is more likely to develop infection, compared to a host with a healthy immune systemn and good nutritional status. More open superficial wounds are less likely to become infected espedsPs cleaned and dressed compared to deeper penetrat if ing or with crush iniury and tissue ischemia. The clostridial and theta toxins that cause ex organisms produce alpha tensive tissue damage. The infection can spread quickly, and within a matter of several hours, the patient may develop overwhelming shock, sepsis, and death. Tissue thatat is better oxygenated with 70mmHg oxy gen tension will inhibit the organism growth because clostridial species are anaerobes. If the oxygen tension of the tissue is less than 30 mm Hg, then the clostridial organ isms groW more will quickly. The infection can develop slowy over weeks or rapidly over hours depending on the Photograph before right leg amputation oxygen tension of the (hemipelvectomy) tissue and the amount of t a patient with gas gangrene. The right thigh organismn inoculated. is edematous (swollen) and discoloured with necrotic bullae (large blisters). Crepitation is detected The virulence of the or on deep palpation. At this juncture, the patient is in shock ganism depends on the (https://upload.wikimedia.org'w vikipedialcommons/ exotoxins produced; thumb/8/81/Gas gangrene.jpg300px-Gas gangrenejpg) Clostridium perfiringens is the most pathologic with 17 known toxins, with the most toxic being the alpha toxin, a lecithinase. Gas gangrene of a diabetic foot Alpha toxin iggering is a phospholipase platelet aggregation, (lecithinase) thrombosis. and that breaks bistamine relaeaes down cell present (https:/www.ncbi.nlm.nih. NBKS37030/bin'gas gangrene gov/books/ jpg) Lesson 25 hemagglutinins, and hemolysins. Theta toxins cause direct vascular injury and are collagenase, hyaluronidase, host inflammatory response to the infection. Collagenase breaks breakdown of leukocytes causing blunted rapid spread of the organism across tissue planes. This is one of the main down connective tissue allowing the tissue plains, spreading into the deeper muscle tissues. reasons that the infection can cross over connective until late in the illness, when dangerously low blood pressure (shock ) and coma Typically, people remain alert develop. Kidney failure and death rapidly follow. Without treatment, death occurs in 100% of infected people, usually within 48 hours. Even with treatment, about one in four people die. Growth on Fluid Thioglycolate Gas gangrene: laboratory diagnostics Clostridium infections mostly exhibit the same As bacterial s sporogenes Medium 9ycolate symptoms, early diagnosis of gas gangrene rarely occurs. Microbiological methods include cultures of fluids from inflicted area, microsco py identification of strain of bacteria sampled from fluids of inflicted area (Gram stain). Robertson 's deep-wound A aerobic and anaerobic culture at Cooked Meat the time of the initial surgical debridement can Medium help determine the causative organism and di (https:// rect antibiotic are inoculated mi therapy. Spee media as hlood Reducing agents in into cooked meat crobeonline.com micrvarrphilie hligateEacultative obliuste the medium absortb agar media and the latter is incubated anaerobi ezoimgfnmt Oxygen and allow il.wp.com/ cally for 48-72 hours. In blood agar medium obligateanaerobes mi there is hemolysis around the colony. The bac to grow crobeonline. com/ terial culture is used for Nagler reaction and WP-content/ biochemical tests. (https://image2.slideserve.com/4787153/growth-on-fluid uploads2016/11/ thioglycolate-ljpg) Specimen are incoulated into cooked meat me Robertsons cooked-mneat dia as well as in a blood agar media and the latter is incubated anaerobically for 48-72 hours. In bloodagar medium.jpg? medium there is hemolysis around the colony. The bacterial culture is used for Nagler reaction (Lecithinase w-225&ss-1&e Test) and biochemical tests. zimgfmt- ng:webp Robertson's Cooked Meat (RCM) Ingcb2) medium is used for the cultivation of Antitoxin Nagler Plate Nagler's reaction: aerobic, microaerophilic, and anaerobic microorganisms, Use: ldentification of pertringens especially Clostridium also known as species. cooked meat broth (CMB)as it contains pieces of fat It is Cl. Clostridium perfringens. free minced cooked meat of ox heart and nutrient broth. Principle: CL spoogenes It : the growth of both spore-forming and non suPming obligate anaerobes es and also differentiates - Clostridiumperfringens spo between putrefactive and saccharolytic species. produces opalescence in human serum or Oxygen in culture media can be reduced by various agents sueh glucose, thioglycollate, cooked meat piec egg yolk media due to as and : eorbic acid. es, cysteine the production of lecithinase C Thioglycollate broth which contains nutrient broth and 1% thioglycollate is also used to cultivate anaerobes. (phospholipase). In litmus milk, C. produces “stormy clot perfringens reaction" due of milk agar and fermenta to proteolysis tion of lactose producing acid and vigorous gas. Lecithinase test or Nagler 's reaction is a biochemical test used to identify The species of the pathogen is identified by revealing of organisms ihat erringens. The alpha liberate phospholipases (lecithinases) e.g. C (a) toxin ofC, perfringens has phosnholin t activi- the specific protein toxin produced by the bacteria in ty and hence, helps in the differentiation ofC. perfringens from oth neutralization reactions performed in vivo with use of er Clostridium spp that also produce lecithinase (C.baratti, C.absonum, laboratory white mice or in vitro. The neutralization Cbifermantans, C.sordelli, and C.novyi) by neutralization of lecithin c activ- reaction is also applied to reveal the type of toxin direct ity by an antitoxin. ly in the specimens without going through the procedure (https:/slideplayer.com/slide'13406153/80 images/24/ of Diagnosist oft gastgangerene jpg) isolation of pure culture of the pathogen. Specific antigen (the component of bacterial cells) can be also detected directly in the specimens by apply ing IFR. Inoculated with Uninoculated tube Gas gangrene: immune prophylaxis and immune therapy C perfringens There are toxoids (perfringens and oedematiens) and antisera (antiperfringens, antioedematiens and antisepticum), but unfortunately anti -gangrenous serum and toxoid are ineffective both in the treatment and in the prevention of gas gangrene. Litmus milk test with 'stormy clot reaction (https:/microbeonline.com/ezoimgfmti2.wp.com/ The pathogen causing tetanus: main characteristics microbeonline.comhwp-contentuploads/2020/05/ Clostridium tetani is a common soil bacterium and the causative agent of Cl Hanerfringens-common-featuresjpg? tetanus. Vegetative cells of C. tetani are usually rod-shaped, but the bacte- resize o102536&slkeimgfmt-ngwebp/ngcb2)