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Eperythrozoon suis in naturally infected pigs: a light and electron microscopic study.

The light and electron microscopic characteristics of natural infections with Eperythrozoon suis are reported. E. suis are enclosed by a single membrane, and they reach a length of up to 600 nm and a diameter of 375 to 500 nm. Neither distinct organelles nor nuclear structures appear in the parasites cytoplasm. They replicate by binary fission, and have close contact with red blood cells without showing any signs of membrance alteration. Massive phagocytosis of whole red blood cells not parasitized by E. suis in spleen and lymph nodes.

Anaplasmataceae Infections↗

Erythrocyte membrane alterations associated with the attachment and replication of Eperythrozoon suis: a light and electron microscopic study.

Erythrocytes from pigs with experimental porcine eperythrozoonosis were examined using light microscopy and scanning and transmission electron microscopy. Three distinct forms of Eperythrozoon suis were attached to the plasma surface of erythrocyte membrane. Erythrocytes were initially parasitized by one or several immature forms. Immature forms enlarged and developed into juvenile and mature forms. The parasite replicated by budding of small immature forms from larger immature forms, juvenile forms, and mature forms. Small immature forms attached to adjacent membrane of the same erythrocyte or a nonparasitized erythrocyte. E. suis organisms were intimately associated with, but distinctly separated from erythrocyte membrane by a 30-nm electron lucent zone. Cell membrane in this area was denser than adjacent nonparasitized membrane. Early interaction between erythrocyte membrane and small immature forms resulted in no membrane deformation, but as this form enlarged, it later became embedded in a deep cup-like membrane invagination. As the immature form developed into the juvenile form a shallow, broad-based depression was observed in the membrane. A similar depression that covered a greater surface area was observed in the membrane parasitized by the mature form. The interaction between the parasitic forms and erythrocyte membrane resulted ultimately in severe membrane deformation. Parasite-membrane interactions may play an important role in the development of misdirected immune responses in experimental porcine eperythrozoonosis.

Animals↗

Detection of Eperythrozoon suis using the polymerase chain reaction.

Eperythrozoon suis is an extracellular red blood cell parasite that causes icteroanemia in acutely ill pigs and a variety of syndromes in chronically infected pigs. Current techniques to detect E. suis infection are limited by variability of parasitemias and antibody responses in infected animals. The polymerase chain reaction (PCR) was investigated to determine its potential as a means of detecting E. suis infection in pigs. With DNA samples extracted from either purified E. suis organisms or E. suis-infected pig blood, PCR produced an amplification product 492 base pairs in length. This amplification product hybridized successfully with the fragment of the DNA probe from which the primer sequences had been selected. Sensitivity studies indicated that the PCR protocol was capable of amplifying total genomic E. suis DNA in quantities as low as 450 pg. When PCR was used with DNA from blood samples from a splenectomized pig that had been infected with E. suis, amplification products were detectable as early as 24 hours postinfection. This preliminary analysis indicates that PCR shows promise as a means of efficiently detecting E. suis infection in pigs.

Animals↗

Comparison of an improved polymerase chain reaction protocol and the indirect hemagglutination assay in the detection of Eperythrozoon suis infection.

The ability of an improved polymerase chain reaction (PCR) protocol to detect Eperythrozoon suis DNA in the blood of experimentally infected nonsplenectomized pigs was evaluated. The protocol utilizes previously described E. suis-specific primers and a proprietary DNA-releasing reagent in a 2-step amplification cycle followed by visualization of the 492-bp amplification product on agarose gels. This PCR protocol successfully amplified E. suis DNA in blood from all postinfection samples and from the preinfection samples of 2 pigs, indicating preexisting natural infections. Results of the indirect hemagglutination test on serum samples from these pigs revealed that only 1 pig developed detectable antibody titers to E. suis infection during the 43-day study; that pig was determined by PCR to have been infected naturally with E. suis prior to experimental inoculation. These results confirm previous reports of poor antibody response of young pigs to E. suis infection and demonstrate the potential of PCR as a valuable tool for the diagnosis and study of E. suis infection in pigs.

Animals↗

Development and evaluation of a polymerase chain reaction assay using the 16S rRNA gene for detection of Eperythrozoon suis infection.

The 16S ribosomal RNA (rRNA) gene of Eperythrozoon suis was amplified using gene-specific primers developed from GenBank sequence accession U88565. The gene was subsequently cloned and sequenced. Based on these sequence data, 3 sets of E. suis-specific primers were designed. These primers selectively amplified 1394, 690, and 839 base-pair (bp) fragments of the 16S rRNA gene from DNA of E. suis extracted from the blood of an experimentally infected pig during a parasitemic episode. No polymerase chain reaction (PCR) products were amplified from purified DNA of Haemobartonella felis, Mycoplasma genitalium, or Bartonella bacilliformis using 2 of these primer sets. When the primer set amplifying the 690-bp fragment was used, faint bands were observed with H. felis as the target DNA. No PCR products were amplified from DNA that had been extracted from the blood of a noninfected pig or using PCR reagents without target DNA. The detection limits for E. suis by competitive quantitative PCR were estimated to range from 57 and 800 organisms/assay. This is the first report of the utility of PCR-facilitated diagnosis and quantitation of E. suis based on the 16S rRNA gene. The PCR method developed will be useful in monitoring the progression and significance of E. suis in the disease process in the pig.

Amino Acid Sequence↗

Light and electron microscopic features of eperythrozoon-like parasites in a North American opossum (Didelphis virginiana).

Epierythrocytic parasites associated with a severe anemic episode have not been previously reported in the opossum. A Wright-Giemsa-stained peripheral blood smear from an anemic North American opossum (Didelphis virginiana) revealed numerous organisms attached to red blood cells either singularly or in chains. Ring forms of the organism were common and could be found free in the plasma. Electron microscopy revealed that these organisms were attached to the intact plasma membrane in depressions on the surface of red blood cells. Delicate fibrils between the organism and adjacent membrane were observed. The organisms were round to oval with a diameter of 300-750 nm and were enclosed by a single limiting membrane. The light and electron microscopic features of these epierythrocytic organisms are similar to those reported for Eperythrozoon and Haemobartonella species.

Anemia↗

Use of a polymerase chain reaction assay to detect infection with Eperythrozoon wenyoni in cattle.

OBJECTIVE: To determine whether a polymerase chain reaction (PCR) assay could be used to detect Eperythrozoon wenyoni in the blood of cattle. DESIGN: Prospective study. ANIMALS: 95 cattle from various herds in Alabama and Georgia and 96 bulls enrolled in Auburn University's Alabama Beef Cattle Improvement Association Bull Test program. PROCEDURE: Blood samples were collected by means of venipuncture of the median caudal vein and submitted for a CBC and PCR assay. Blood smears were made immediately after blood collection and examined by means of light microscopy. RESULTS: Three of 95 cattle from herds in Alabama and Georgia and 5 of 96 bulls enrolled in the Bull Test program had positive PCR assay results. Organisms were seen in blood smears from only 5 of these 8 animals. Organisms were not seen in blood smears from any animals for which results of the PCR assay were negative. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that a PCR assay may be an effective method for detecting E wenyoni infection in cattle and that the PCR assay may be a more sensitive test than evaluation of blood smears.

Alabama↗

[The effect of Eperythrozoon suis infection on the osmotic fragility of erythrocytes].

Osmotic fragility of erythrocytes was tested in weaned pigs experimentally infected with Eperythrozoon (E.) suis. Acute eperythrozoonosis of splenectomized pigs led to an increase of osmotic fragility. It is supposed that E. suis infection causes a structural change in erythrocyte membrane. Possible mechanisms of this cell membrane injury are discussed.

Animals↗

Evaluation of an enzyme-linked immunosorbent assay for detection of Eperythrozoon suis antibodies in swine.

An ELISA was developed and tested to detect antibodies to Eperythrozoon suis in swine. Results were compared with those of the indirect hemagglutination (IHA) test. Antigen isolated from swine heavily infected with E suis was used for both tests. Comparison of the ELISA with the IHA test revealed a significant (P less than 0.001) correlation between results. Of 114 samples obtained from 9 swine infected with E suis, 87.7% were seropositive (titer greater than or equal to 200) via the ELISA, and 80.7% were seropositive (titer greater than or equal to 20) via the IHA test. The sensitivity of the ELISA was greater than that of the IHA test. All blood samples obtained from specific-pathogen-free swine tested negative for E suis antibody. Cross-reactions were not observed between E suis antigen and antisera against various swine and cattle disease agents using ELISA. We concluded that the ELISA may be used for rapid and effective diagnosis of infection with E suis in swine.

Animals↗

Recombinant DNA probe detecting Eperythrozoon suis in swine blood.

A genomic library to Eperythrozoon suis DNA was constructed in lambda gt11, and from this library, E suis clone KSU-2 was identified as a potential diagnostic probe. In hybridization experiments that used 100-microliters samples of blood collected in chaotropic salt solutions, the KSU-2 probe hybridized strongly with purified E suis organisms and blood samples from splenectomized swine that were parasitized with E suis. However, the probe under stringent conditions did not give radiographic indications of hybridizing with equine blood DNA, bovine blood DNA infected with Anaplasma marginale, canine blood DNA infected with Ehrlichia canis, feline blood DNA infected with Haemobartonella felis, or uninfected swine blood DNA.

Animals↗

An Eperythrozoon-like parasite in llamas.

An Eperythrozoon-like RBC parasite was found in a herd of llamas in western Kentucky. The light microscopic and electron microscopic features of the organism were similar to those of E suis. Results of an indirect hemagglutination test for E suis were positive with sera from some parasitemic and nonparasitemic llamas. Parasitemic llamas were usually less than 1 year old and had more severe clinical signs of disease than did older llamas. Poor weight gain and growth, anorexia, and prolonged recumbency were the principal signs of disease. Anemia was usually mild and not accompanied by icterus. Hypoglycemia was associated with parasitemia. Some response to oral or parenteral treatment with tetracycline was noticed, but recrudescence of the infection in some treated llamas was observed. Llamas in nearly every region of the United States have had titers in approximately 12% of samples tested, suggesting that infection with the organism may be widespread. Parasitemic llamas have been recognized in at least 8 states.

Animals↗

[Serologic progression studies using CF and ELISA for the detection of antibodies against Eperythrozoon suis infection of swine].

Serological studies by means of a CF-Test and Elisa were undertaken for evaluation of antibody titers against Eperythrozoon suis infection in swine. The animals were partly splenectomized and underwent artificial infection. It could be shown that the splenectomized animals showed higher and longer lasting antibody titers than the non splenectomized swine. It is stated, that the above mentioned tests can be used in diagnosis on a herd basis but not on a single animal basis.

Anaplasmataceae Infections↗

[Eperythrozoon infection in swine: effect on the acid-base balance and the glucose, lactate and pyruvate content of venous blood].

The influence of latent and of splenectomy-induced clinically manifest Eperythrozoon suis infection on the following parameters of the carbohydrate metabolism and the acid-base status was tested in venous blood of German Landrace pigs: Levels of glucose, lactic and pyruvic acid, blood-pH, base excess, actual bicarbonate concentration, standard bicarbonate concentration, pCO2, pO2. The latent E. suis infection resulted in a consistent decrease of blood glucose level. 23 days after infection, blood glucose was reduced by 25% of the initial value. The other parameters were not changed by latent E. suis infection. Acute Eperythrozoonosis induced severe hypoglycaemia (means Gluc, = 39.7 mg/dl and blood acidosis (means pH = 7.13). In vitro experiments showed that break-down of glucose in E. suis infected blood occurs very rapidly. There was no significant reduction of the glucose concentration in control blood that had been treated accordingly. There was an increase of lactic acid (means = 62.7 mg/dl), pyruvic acid (means = 1.86 mg/dl), and pCO2 (means = 82.1 mm Hg). The concentrations of actual bicarbonate (means = 24.8 mmol/l) and standard bicarbonate (means = 20.9 mmol/l) were lowered, and there was a negative base excess (means = -3.56 mmol/l). The ratio of lactic and pyruvic acid changed from 11:1 to 30:1. It seems likely that E. suis itself is able to metabolize glucose. Acidosis is considered to result from both the increase of lactic acid (metabolic component) and an impairment of pulmonary gas exchange (respiratory component).

Acid-Base Equilibrium↗

Eperythrozoon wenyonii infection in dairy cattle.

Approximately 10 of 100 young heifers that had recently delivered their first calf--members of a large Colorado dairy herd--had a syndrome of swollen teats and distal portions of the hind limbs, prefemoral lymphadenopathy, transient fever, rough coat, decreased milk production, and subsequent weight loss and reproductive inefficiency. Acute clinical signs of disease were associated with large numbers of Eperythrozoon wenyonii seen on blood smears, and resolution of signs correlated with reduction or disappearance of the parasite. Other known causes of peripheral edema could not be documented. The parasite was transmitted to 4 of 7 nonlactating dairy cows destined to be culled and a splenectomized calf via IV inoculation of blood from parasitemic heifers, but clinical signs of infection were not induced.

Anaplasmataceae Infections↗

[The acid-base equilibrium and carbohydrate metabolism during infection with Eperythrozoon suis].

Following on from clinical observations which point to severe metabolic disturbances in association with acute Eperythrozoon (E.) suis infection, the parameters of acid-base balance (pO2, pCO2, pH, actual bicarbonate, standard bicarbonate, base excess) as well as the glucose-, lactate- and pyruvate levels, were measured in venous blood during the course of eperythrozoonotic infection. Glucose consumption was investigated in in vitro experiments with differing numbers of pathogens. Acute E. suis infection is accompanied by a severe acidosis and hypoglycaemia. In vitro experiments showed that a rapid breakdown of glucose follows in E. suis infected blood. No significant reduction in glucose concentration was established in control blood in a comparable time period. The results give rise to the assumption that E. suis is capable of independent glucose breakdown. Both the increase in lactate concentration (metabolic component) and a disturbance of pulmonary gaseous exchange (respiratory component) are regarded as the cause of the acidosis.

Acid-Base Equilibrium↗

[Eperythrozoon infection in swine as a disease factor].

Infection with Eperythrozoon suis (E. suis) normally takes a latent course and depends on various factors. Only animals suffering extreme stress show clinically manifest attacks with anemia, fever and occasionally jaundice. Stress in the form of high animal-concentration in the stables, poor climatic conditions, change of stables or food or chronic general diseases predispose the pigs to clinical illness. The diagnosis of E. suis infection is made difficult by the prevalence of general diseases in the stocks. Experimental infections have shown that also latent infections with E. suis cause partly significant deviations in the red and white blood cell count as well as in the glucose and acid-base balance and lead to serologically detectable immune reactions which could be used as a basis for stock-diagnosis. It is still unknown to what extent infection with E. suis represents a factor in the manifestation of other complex diseases. Some observations, however, give reason to believe that Eperythrozoonosis as a basis infection is itself a precursor for the manifestation of widespread infectious diseases.

Anaplasmataceae Infections↗

Arthropod transmission of Eperythrozoon coccoides in mice.

The association of Eperythrozoon coccoides infection in laboratory mice, with commonly encountered external parasites, was examined. Transmission of E. coccoides was achieved by infesting recipient mice with as few as 10 live Polyplax serrata immediately after they were obtained from donor mice infected with E. coccoides. Polyplax serrata removed from such hosts and fasted for 24 hours also were able to initiate E. coccoides infections in recipient mice. Transovarial transmission of E. coccoides through P. serrata was not demonstrated. Transmission of E. coccoides by Myocoptes musculinus, Myobia musculi and Radfordia affinis was not observed.

Anaplasmataceae Infections↗

The significance of Eperythrozoon ovis in ill-thrift in sheep in the eastern Cape coastal areas of South Africa.

Ill-thrift in sheep in the coastal region of the Eastern Cape is described. It is shown to be a complex problem attributable to many causes, most of which can be eliminated by supplementary feeding, drenching, vaccination, dipping and management. However, studies on the aetiology of ill-thrift in young sheep indicate that arthropod-borne anaemia-producing pathogens are an important contributing factor, which cannot readily be diagnosed and controlled. Experiments were conducted on various farms to determine whether Eperythrozoon ovis occurred in sheep in the coastal areas of the Eastern Cape and to what extent infection affected these animals. Other blood parasites found in sheep in the Eastern Cape included Borrelia theileri, Anaplasma ovis, Ehrlichia ovina, Cytoecetes phagocytophila and Theileria ovis. E. ovis, either alone or in combination with one or more of these parasites, caused a severe prolonged anaemia accompanied by the development of ill-thrift.

Anaplasmataceae Infections↗