Putative morphology of Neoehrlichia mikurensis in salivary glands of Ixodes ricinus.

Neoehrlichia mikurensis is an emerging tick-borne intracellular pathogen causing neoehrlichiosis. Its putative morphology was described in mammalian, but not in tick cells. In this study, we aim to show the presumptive morphology of N. mikurensis in salivary glands of engorged females of Ixodes ricinus. To accomplish this, we collected I. ricinus ticks in a locality with a high N. mikurensis prevalence, allowed them to feed in the artificial in vitro feeding system, dissected salivary glands and screened them by PCR for N. mikurensis and related bacteria. Ultrathin sections of salivary glands positive for N. mikurensis but negative for other pathogens were prepared and examined by transmission electron microscopy. We observed two individual organisms strongly resembling N. mikurensis in mammalian cells as described previously. Both bacteria were of ovoid shape between 0.5-0.8 µm surrounded by the inner cytoplasmic and the rippled outer membrane separated by an irregular electron-lucent periplasmic space. Detection of N. mikurensis in salivary glands of I. ricinus suggests that this bacterium uses the "salivary pathway of transmission" to infect mammals.

Putative phage hyperparasite in the rickettsial pathogen of abalone, "Candidatus Xenohaliotis californiensis".

Studies on the ecology of microbial parasites and their hosts are predicated on understanding the assemblage of and relationship among the species present. Changes in organismal morphology and physiology can have profound effects on host-parasite interactions and associated microbial community structure. The marine rickettsial organism, "Candidatus Xenohaliotis californiensis" (WS-RLO), that causes withering syndrome of abalones has had a consistent morphology based on light and electron microscopy. However, a morphological variant of the WS-RLO has recently been observed infecting red abalone from California. We used light and electron microscopy, in situ hybridization and16S rDNA sequence analysis to compare the WS-RLO and the morphologically distinct RLO variant (RLOv). The WS-RLO forms oblong inclusions within the abalone posterior esophagus (PE) and digestive gland (DG) tissues that contain small rod-shaped bacteria; individual bacteria within the light purple inclusions upon hematoxylin and eosin staining cannot be discerned by light microscopy. Like the WS-RLO, the RLOv forms oblong inclusions in the PE and DG but contain large, pleomorphic bacteria that stain dark navy blue with hematoxylin and eosin. Transmission electron microscopy (TEM) examination revealed that the large pleomorphic bacteria within RLOv inclusions were infected with a spherical to icosahedral-shaped putative phage hyperparasite. TEM also revealed the presence of rod-shaped bacteria along the periphery of the RLOv inclusions that were morphologically indistinguishable from the WS-RLO. Binding of the WS-RLO-specific in situ hybridization probe to the RLOv inclusions demonstrated sequence similarity between these RLOs. In addition, sequence analysis revealed 98.9-99.4 % similarity between 16S rDNA sequences of the WS-RLO and RLOv. Collectively, these data suggest that both of these RLOs infecting California abalone are "Candidatus Xenohaliotis californiensis," and that the novel variant is infected by a putative phage hyperparasite that induced morphological variation of its RLO host.

Ultrastructure and phylogenetic analysis of 'Candidatus Neoehrlichia mikurensis' in the family Anaplasmataceae, isolated from wild rats and found in Ixodes ovatus ticks.

A novel bacterium that infects laboratory rats was isolated from wild Rattus norvegicus rats in Japan. Transmission electron microscopy of the spleen tissue revealed small cocci surrounded by an inner membrane and a thin, rippled outer membrane in a membrane-bound inclusion within the cytoplasm of endothelial cells. Phylogenetic analysis of the 16S rRNA gene sequence of the bacterium found in R. norvegicus rats and Ixodes ovatus ticks in Japan revealed that the organism represents a novel clade in the family Anaplasmataceae, which includes the Schotti variant found in Ixodes ricinus ticks in the Netherlands and the Ehrlichia-like Rattus strain found in R. norvegicus rats from China. The novel clade was confirmed by phylogenetic analysis of groESL sequences found in R. norvegicus rats and Ixodes ovatus ticks in Japan. No serological cross-reactivity was detected between this bacterium and members of the genera Anaplasma, Ehrlichia or Neorickettsia in the family Anaplasmataceae. It is proposed that this new cluster of bacteria should be designated 'Candidatus Neoehrlichia mikurensis'.

Description of Aegyptianella botuliformis n. sp. (Rickettsiales: Anaplasmataceae) from the helmeted guineafowl, Numida meleagris.

Aegyptianella botuliformis n. sp. (Rickettsiales: Anaplasmataceae) isolated from helmeted guineafowls Numida meleagris from the Kruger National Park is described. The rickettsia occurs within a membrane-bound vacuole in the cytoplasm of erythrocytes with up to 8 organisms in a mature inclusion. The initial body resembles that of Aegyptianella pullorum. The tightly packed, sausage-shaped intermediate forms are a distinctive morphological feature, seen as irregular, pleomorphic forms under light microscopy. While more larvae and nymphs of Amblyomma hebraeum and Amblyomma marmoreaum were found on the birds than larvae of an Argas sp., it is believed that the latter are the vectors of A. botuliformis n. sp. In addition to the Kruger National Park, positive blood smears were obtained from guineafowls at other localities in the Transvaal.

The morphological features of Aegyptianella bacterifera: an intraerythrocytic rickettsia of frogs from Corsica.

Study of ultrathin sections and freeze-fracture replicas of erythrocytes containing Aegyptianella bacterifera (Rickettsiales; Anaplasmataceae) revealed that the organisms develop within a membrane bound vacuole in the erythrocyte cytoplasm. The organisms divide by binary fission to produce about 12 rickettsiae in a mature inclusion. The organisms have gram-negative cell envelopes. The distribution of intramembranous particles within the outer and plasma membranes of A. bacterifera is similar to that of other rickettsiae and gram-negative bacteria such as Escherichia coli. The definition of the genus Aegyptianella should be broadened to include rickettsiae measuring up to 5 microns in length prior to division which develop within membrane bound inclusions in erythrocytes of their hosts.

Isolation and identification of Aegyptianella pullorum (Rickettsiales, Anaplasmataceae) in wild turkeys from North America.

Isodiagnosis of blood from Rio Grande wild turkeys from southern Texas revealed a small, intraerythrocytic rickettsia, Aegyptianella pullorum Carpano, 1928, in 24 of 300 samples. Identification of this first isolate from North America was made using both light and transmission electron microscopy. It is suggested that the translocation of wild turkeys from Texas to other states could spread this pathogen to both wild birds and domestic poultry.

Hemobartonellosis in squirrel monkeys (Saimiri sciureus) in a domestic breeding colony: case report and preliminary study.

Infection with Hemobartonella sp was diagnosed in a colony-born squirrel monkey with normocytic, normochromic anemia and pronounced punctate erythrocytic basophilic stippling on Wright's-Giemsa stained blood films. The diagnosis was confirmed using transmission electron microscopy. Two randomly selected colony-born squirrel monkeys were splenectomized in an effort to activate and detect possible latent hemobartonellosis . One monkey became parasitemic 12 days following splenectomy. The second monkey was inoculated on day 14 with 1 ml of whole blood from an infected, but nonparasitemic monkey and developed overt parasitemia 3 days later (day 17 following splenectomy). Infections in the latter two monkeys were confirmed using scanning electron microscopy.

Sequestration and phagocytosis of Haemobartonella felis in the spleen.

Spleens of two cats infected with Haemobartonella felis were examined by electron microscopy to determine the means by which the organism was sequestered in this organ. The principal means of sequestration occurred when H felis, located on the erythrocytes was removed by phagocytosis by a cordal macrophage, apparently preceded by the adhesion of extended processes of the macrophage to H felis. The second and least frequent means of removal of H felis was by pitting, a process that did not cause destruction of the host erythrocyte. The H felis was pitted from the parasitized erythrocyte when H felis passed through gaps between reticular cells or when the parasitized erythrocyte passed among the cytoplasmic processes of the reticular cells in the splenic cords. Some H felis were closely associated with the plasmalemma of cordal reticular cells and also were located in intracytoplasmic vacuoles of the cells without being influenced by the phagocytic process.

Morphologic alteration of Paranaplasma caudata following treatment of calves with oxytetracycline.

The morphologic features of Paranaplasma caudata were determined in whole blood, blood clots, and liver biopsy specimens after treating infected calves with oxytetracycline. Following staining of blood smears with new methylene blue, light microscopy showed that paranaplasma bodies in erythrocytes of treated calves were vacuolated, but tails were normal in appearance. By electron microscopic examination of parasitized erythrocytes of treated calves, paranaplasma bodies were found to be degenerated (aggregation of nucleoprotein granules in initial bodies and condensation and coalescence of initial bodies), but tails were normal. The identical degenerative changes of paranaplasma bodies and lack of alteration of tails were evident in erythrocytes in sinusoids of liver biopsy specimens from treated calves.