Blood and Tissue Flagellates: Coccidians PDF

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Summary

COCCIDIANS largest group of apicomplexan protozoa falling under Class Conoidasida microscopic, spore-forming, single-celled, obligate intracellular protozoan members of Phylum Apicomplexa are provided w/ a cluster of secretory cells ↳ made up of rhoptries, micronemes and polar rings w/ microtubules ...

Description

COCCIDIANS - largest group of apicomplexan protozoa falling under Class Conoidasida - microscopic, spore-forming, single-celled, obligate intracellular protozoan - members of Phylum Apicomplexa are provided w/ a cluster of secretory cells ↳ made up of rhoptries, micronemes and polar rings w/ microtubules ↳ in some spp., a conoid may be found within polar rings ↳ secretion allows the parasite to enter the host cell

3.



gametogony: result in dev’t of male (micro) & female (macro) gametocytes (gamonts)

complexity is a challenge in terms of taxonomy

I. CRYPTOSPORIDIUM HOMINIS ✓ It was initially reported that the only species that infects mammals was C. parvum and was believed to be the species infecting humans.

Class



Conoidasida

Order



Eucoccidiorida

Suborder



Eimeriorina

PARASITE BIOLOGY

Disease



Coccidiosis ↳ one of the major problems in animal farming & in zoo management ↳ considered to be opportunistic in immunocompromised & immunodeficient humans

Oocyst s

✓ However, molecular tools (esp. DNA analysis) described the existence of C. hominis found mainly in humans.

   

4-5um in diameter round each contains 4 sporozoites ↳ present at the time of passage into the feces infectious

✓ All stages of dev’t are completed in the GI tract of the host. Habitat

Species with medical & veterinary significance

Life cycle



intestinal tract of most phyla of intervertebrates & all all classes of vertebrates (including humans)

1. 2. 3. 4. 5.

Cryptosporidium Cyclospora Cystoisospora Sarcocytis Toxoplasma



alternation of sexual & asexual multiplication characterized by 3 sequential stages 1. sexual cycle: sporogony producing oocysts 2. asexual cycle: schizogony (merogony) producing merozoites (meronts)



1. Oocyts when passed out are already infective.  oocytes produced by C. hominis are found in the feces of humans & other animals 2. Upon ingestion, the sporozoites attach to the surface of epithelial cells of the GI tract. 3. Sporozoites develop into small trophozoites & become intracellular but extracytoplasmic, and attach to the brush borders. 4. Trophozoites divide by schizogony producing merozoites that infect other cells. 5. Macro- and microgametocytes are eventually produced.

6. Macrogamete is fertilized by the microgamete to produce a zygote. 7. There are 2 types of oocyst resulting from the zygote: Thin-walled oocyst



Thick-walled oocysts

  

infect other enterocytes, resulting in autoinfection ↳ responsible for the chronicity of the infection among the immunocompromised passed out w/ the feces may contaminate food & water ingested by the same or another host

Sporulated oocysts, containing 4 sporozoites, are excreted by the infected host through feces (and possibly other routes such as respiratory secretions). Transmission of Cryptosporidium spp. occurs mainly through ingestion of fecally contaminated water (e.g., drinking or recreational water) or food (e.g., raw milk) or following direct contact with infected animals or people . Following ingestion (and possibly inhalation) by a suitable host , excystation occurs. The sporozoites are released and parasitize

the epithelial cells ( , ) of the gastrointestinal tract (and possibly the respiratory tract). In these cells, usually within the brush border, the parasites undergo asexual multiplication (schizogony or merogony) ( , , ) and then sexual multiplication (gametogony) producing microgamonts (male) and macrogamonts (female) . Upon fertilization of the macrogamonts by the microgametes ( ) that rupture from the microgamont, oocysts develop and sporulate in the infected host. Zygotes give rise to two different types of oocysts (thick-walled and thin-walled). Thick-walled oocysts are excreted from the host into the environment , whereas thin-walled oocysts are involved in the internal autoinfective cycle and are not recovered from stools . Oocysts are infectious upon excretion, thus enabling direct and immediate fecal-oral transmission. Extracellular stages have been reported, but their relevance in the overall life cycle is unclear. PATHOGENESIS  was not well recognized prior to the occurrence of AIDS  villi of the intestines become blunted  infiltration of inflammatory cells into the lamina propria & elongated crypts  varying degrees of malabsorption  death may occur in disseminated infections In the 1. 2. 3. 4. 5.

immunocompetent host: Self-limiting diarrhea (2-3 weeks) Abdominal pain Fever Nausea Weight loss

In immunocompromised persons: 1. Diarrhea becomes more severe, progressive, lifethreatening 2. Bile duct & gall bladder may become heavily infected ↳ can lead to acute & gangrenous cholecystitis 3. Respiratory infections can lead to i. chronic coughing ii. dyspnea iii. bronchiolitis iv. pneumonia

4.

Excessive fluid loss

DIAGNOSIS 1. Sheather’s sugar flotation* 2. FECT (formalin ether/ethyl acetate)* 3. Kinyoun’s modified acid-fast stain ↳ oocysts appear as red-pink doughnut-shaped circular organisms in a blue background 4.

Examination of intestinal biopsy material under a light microscope ↳ stages of the parasite can be seen at the microvillus region of the infected enterocyte 5.

Recovery of parasite from the sputum ↳ for cases that involve pulmonary involvement ↳ note: transbronchial & broncheo-alveolar lavage can yield a better result 6.

Indirect fluorescent antibody, enzyme immunoassay and DNA probes specific for C. hominis have been developed.

7.

Acid-fast staining ↳ quickest & cheapest method of diagnosis

*commonly used TREATMENT There is no acceptable treatment for cryptosporidiosis. 1. Nitazoxanide

 reported effective in preliminary trials

2. Bovine colostrum, Paromomycin and Clarithromycin

 for severe diarrhea

3. Azithromycin 4. Chemotherapy, body fluid replacement, symptomatic

 recommended for both immunocompetent & immunosuppressed patients

treatment

In the PH

EPIDEMIOLOGY 1. Universal distribution ↳ infections reported worldwide

Prevalence Rates In developing countries In the PH A study done in San Lazaro Hospital attempted to describe Cyrptosporidium among diarrheic patients

2. Epidemics are unusual in North America. 3. There was a report of an epidemic involving 400,000 cases in the state of Wisconsin in the US. ↳ attributed to the use of a faulty water purification system 4. Most epidemics are associated w/ water . ↳ water in most cases was contaminated with calf feces 5. Cryptosporidium parvum of calves has been reported to cause infection among veterinary attendants & visitors in dairy farms and petting zoos. 6. Swimming in contaminated recreation water may result in accidental ingestion of infective oocysts. ↳ swimming pool disinfection w/ 3-5mg/L of chlorine does not kill the oocysts 7. The most common MOT is from one person to another. 8. Infected food handlers may transmit oocysts during handling of i. beverages ii. raw vegetables iii. other food that may be eaten raw 9. Unpasteurized milk, freshly pressed apple cider, potato salad, and sausages were found to be sources of infection. 10. Nosocomial infections have also been reported among health workers caring for AIDS patients. Prevalence Rates  in developing countries: 3-20%  in the PH: 2.6% (low)

A study done in PGH on diarrheic patients

3-20% 2.6% (low) 8.5%

1.7%

PREVENTION & CONTROL 1. Water-borne transmission is the most common source of cryptosporidiosis. 2. Chlorination does not affect the parasite. 3. The synergistic effect of multiple disinfections & combined water treatment may reduce C.hominis cysts in drinking water. 4. Natural water & swimming pool water should not be swallowed. 5. Contamination of drinking water by human & animal feces should be prevented. II. CYCLOSPORA CAYETANENSIS ✓ When first associated w/ diarrhea, this organism was thought to be a member of cyanobacteria because it showed characteristics of the blue-green algae 1. photosynthesizing organelles 2. auto-fluorescing particles PARASITE BIOLOGY ✓ Cyclospora cayetanensis was originally called a cyanobacterium-like body (CLB), but upon careful study, it was found to be a coccidian parasite. ✓ The different developmental stages of the parasite may be found in the intestinal tissue.

Sporulated



contains 2 sporocysts w/ 2

oocyst Meronts



Oocyst

   

1. 2.

3. 4. 5. 6.

sporozoites each contain 8-12 merozoites in the 1st gen contain 4 merozoites in the 2nd gen undergo complete sporulation within 7-12 days in a warm environment assumed to be the infective stage when ingested, sporozoites are released and enter intestinal cells to go through schizogony & gametogony

LIFE CYCLE

Life cycle begins with the ingestion of sporulated oocyst. The released sporozoites invade the epithelial cells of the small intestines, although the site of predilection was found to be the jejunum. Multiple fissions of sporozoites take place inside the cells to produce meronts. Some develop into male (micro) & female (macro) gametes. Microgametes fertilize the macrogametes to produce oocysts. Oocysts are passed out w/ feces when host cells are sloughed off from the intestinal wall.

When freshly passed in stools, the oocyst is not infective (thus, direct fecal-oral transmission cannot occur; this differentiates Cyclospora from another important coccidian parasite, Cryptosporidium). In the environment , sporulation occurs after days or weeks at temperatures between 22°C to 32°C,

resulting in division of the sporont into two sporocysts, each containing two elongate sporozoites . The sporulated oocysts can contaminate fresh produce and water which are then ingested . The oocysts excyst in the gastrointestinal tract, freeing the sporozoites, which invade the epithelial cells of the small intestine . Inside the cells they undergo asexual multiplication into type I and type II meronts. Merozoites from type I meronts likely remain in the asexual cycle, while merozoites from type II meronts undergo sexual development into macrogametocytes and microgametocytes upon invasion of another host cell. Fertilization occurs, and the zygote develops to an oocyst which is released from the host cell and shed in the stool . Several aspects of intracellular replication and development are still unknown, and the potential mechanisms of contamination of food and water are still under investigation.

DIAGNOSIS 1. Direct microscopic examination of fecal smears under high magnification (400x) 2. Various concentration techniques & acid-fast staining (Kinyoun’s stain) 3. Safranin staining & microwave heating 4. Oocysts are autofluorescent & under fluorescent microscopy

 appear blue/green circles depending on the filter (365-450 DM)  useful for screening

✓ Infections are usually self-limiting. ✓ Immunity may result w/ repeated infections. ✓ No reported deaths.

5. PCR

 used to differentiate Cyclospora from closely related Eimeria species

1. Malaise and low grade fever

 

initial symptoms occur 12-24 hrs after exposure

2. Chronic & intermittent watery diarrhea



occurs early in the infection may alternate w/ constipation may last 6-7 weeks w/ 6 or more stools per day

TREATMENT 1. The disease is self-limiting & treatment is not necessary if symptoms are mild. 2. If pharmacologic treatment is warranted, the only effective drug is trimethoprim-sulfamethoxazole 160/800mg twice daily for 7 days. 3. There is no alternate treatment if patients are unable to tolerate sulfamethoxazole. 4. Oocysts disappear from the stools a few days after treatment. 5. Recurrence of symptoms was noted in about 40% patients within 1-3 months posttreatment.

PATHOGENESIS & CLINCAL MANIFESTATIONS

 

4. D-xylose malabsorption

5. Fatigue, anorexia, weight loss, vomiting, abdominal pain, flatulence, bloating, dyspnea



found to develop in some patients

EPIDEMIOLOGY Nepal

  

appeared in late May and June and continued until October to November most cases were reported in expatriates & tourists more recently reported in Nepali children & adults



leafy vegetables have been found to contain oocysts it is believed that cabbage became contaminated when watered w/ irrigation water

 

commonly reported in children leafy vegetables have been found to contain oocysts

Haiti



affect more homosexual males

United States

 

epidemics were reported in 1996 & 1997 raspberries imported from Guatemala were incriminated in the infections in the US lettuce & basil-pesto salad has been incriminated in other cases



Peru

 Guatemala

Philippines





it was believed that raspberries exposed to oocysts in places where creek water was used to dilute insecticides sprayed on the plants a study of diarrheic stools from children at the College of Public Health UPM revealed a prevalence of 3.1% using safranin staining heated in a microwave (2005)

✓ No animal reservoirs have been found dead, therefore cyclosporidiosis is presently considered mainly as a human disease. PREVENTION & CONTROL 1. Since the direct source is unknown, good sanitary practices should be followed. 2. Only water that has been subjected to adequate treatment procedures should be consumed. 3. Boil water. (Chlorination is NOT effective) 4. Fruits & vegetables should be washed w/ clean water and avoid eating if exposed to natural untreated water.



III. CYSTOISOSPORA BELLI causative agent of cystoisosporiasis ↳ medical condition affecting the small bowel ↳ the other known species Isospora hominis is now taxonomically grouped under the genus Sarcocystis

PARASITE BIOLOGY Oocysts  translucent, oval, 20-33um x 10-19um 1. 2.

3. 4. 5. 6. 7.

The sporulated oocyst contains 2 sporocysts each, containing 4 sporozoites (infective stage). When ingested via contaminated water/food, the sporozoites excyst in the small intestine releasing sporozoites. Sporozoites penetrate the epithelial cells, staring the asexual stage (schizogonic phase) of the life cycle. Sporozoites develop in the epithelial cell to form a schizont. Schizont ruptures the host epithelial cell, liberating merozoites into the lumen. Merozoites infect new epithelial cells, and the process of asexual reproduction in the intestine continues. ↳ may continue for weeks or months Some of the merozoites undergo gametogony to produce macrogametes & microgametes (sexual stages), which fuse to form a zygote that eventually matures to form an unsporulated oocyst.

one week, the sexual stage begins with the development of male and female gametocytes . Fertilization results in the development of oocysts that are excreted in the stool . PATHOGENESIS & CLINICAL MANIFESTATIONS ✓ The mechanism by which the parasite produces these lesions is still not clear. Immunocompetent 1. Generally asymptomatic or may present as a self-limiting gastroenteritis 2. Severe diarrhea & malabsorption



may occur in more severe infections

2. Severe diarrheal illness



resembles that of cryptosporidiosis, giardiasis or cyclosporiasis

3. Flattened mucosa & damaged villi



revealed in mucosal bowel biopsy

3. Low-grade fever, anorexia, vomiting, general body malaise, weight loss, flatulence 4. Stools usually contain undigested food, mucus, and Charcot-Leyden crystals Immunocompromised 1. Self-limiting enteritis

At time of excretion, the immature oocyst contains usually one sporoblast (more rarely two) . In further maturation after excretion, the sporoblast divides in two (the oocyst now contains two sporoblasts); the sporoblasts secrete a cyst wall, thus becoming sporocysts; and the sporocysts divide twice to produce four sporozoites each . Infection occurs by ingestion of sporocysts-containing oocysts: the sporocysts excyst in the small intestine and release their sporozoites, which invade the epithelial cells and initiate schizogony . Upon rupture of the schizonts, the merozoites are released, invade new epithelial cells, and continue the cycle of asexual multiplication . Trophozoites develop into schizonts which contain multiple merozoites. After a minimum of

4. Infiltration of lamina propria w/ lymphocytes, plasma cells and eosinophils

DIAGNOSIS 1. Direct microscopy or FECT

 

detection of oocysts presence of CharcotLeyden crystals

 

2. Other concentration techniques

1. 2.

Zinc sulfate Sugar flotation

 

3. Fecal smear stained by a modified Ziehl-Neelsen method



oocysts stain granular red color against a green background

4. Phenol-auramine, iodine staining



help visualize the organism

5. Acid-fast stain (Kinyoun’s stain or auramine-rhodamine stain) 7. Blood examination





peripheral eosinophilia is common

8. String capsule (Enterotest) 9. Duodenal aspirate examinations 10. Molecular-based techniques TREATMENT For asymptomatic infections 1. Bed rest 2. Bland diet For symptomatic infections (e.g. those occurring in AIDS patients) 1. Trimethoprim-sulfamethoxazole  160/800 mg 4x per day for 10 days  2x per day for 3 weeks 2. Combination therapy w/ pyrimethamine & sulfadiazine  for 7 weeks EPIDEMIOLOGY

 

  

unlike other coccidians, humans are the only known hosts of C.belli worldwide distribution more common in tropical & subtropical countries w/ poor sanitary conditions actual incidence is not known has been tagged as the causative agent of diarrheal episodes in day care centers and mental institutions common among patients w/ AIDS also reported among those with 1. lymphoma 2. leukemia 3. organ transplants has been reported in both adults & children severe diarrhea is common among infants both sexes were found susceptible to infection

In Africa, 2-3% of those with AIDS were infected.  South America: 10%  Haiti & Africa: 7-20% Endemic in: 1. Africa 2. Australia 3. Carribean Islands 4. Latin America 5. Southeast Asia PREVENTION & CONTROL 1. Good sanitary practices 2. Thorough washing & cooking of food 3. Drinking safe water

  

IV. TOXOPLASMA GONDII belongs to Phylum Apicomplexa worldwide distribution infects humans & many species of animals

PARASITE BIOLOGY Infective stages

1. Tachyzoite 2. Bradyzoite 3. Oocyst

Definitive host Complete life cycle Intestinal stages

cat family (Felidae) occurs only in members of the cat family 1. Schizogony 2. Gametogony 3. Sporogony  

Cysts Extraintestinal stages

Asexual stages: 1. Tachyzoites 2. Bradyzoites

Stages present in humans & other intermediate hosts

1. 2.

Tachyzoites Bradyzoites

Endodyogeny



variation of binary fission that allows asexual ...