Chemotherapy of Babesia divergens in the gerbil, Meriones unguiculatus

It was found that surprisingly low doses of four babesicides were effective against Babesia divergens in gerbils and it was concluded that this was due to the involvement of host resistance, which may be of a non-specific nature. The efficacy of the drugs relative to each other was the same in gerbils as in cattle and this host-parasite system is evidently more suitable for the screening of babesicides than are other rodent babesia systems. The prophylactic dose of imidocarb dipropionate required to provide a similar degree of protection in gerbils as in cattle was found to be much higher and was very close to toxic levels. Challenge infections resulted in sterile immunity. Acute babesiosis in gerbils could be cured with all four drugs if parasitaemias were below approximately 45 per cent and packed cell volumes above 18 per cent at treatment.     

Induction of protective immunity to Babesia divergens in Mongolian gerbils, Meriones unguiculatus, using culture-derived immunogens

Immunogens derived from microaerophilous stationary phase (MASP) cultures of Babesia divergens grown in bovine erythrocytes were used to inoculate the laboratory host of B. divergens, the Mongolian gerbil, Meriones unguiculatus. Animals inoculated subcutaneously twice with preparations of freeze-thawed merozoites in complete Freund's adjuvant were fully protected against homologous challenge, as were gerbils immunised with a non-viable preparation of parasite-enriched lysed infected bovine erythrocytes. Animals which had been infected with small numbers of parasitised erythrocytes from cultures cooled to 4 degrees C, allowed to recover, then challenged, also survived. All three groups had high antibody titres which dropped immediately after challenge and then rose again. Gerbils given culture supernatants containing soluble merozoite protein coat antigens were partially protected only after receiving a third inoculation. Non-immunised animals all died 4 days after challenge.     

Preliminary development of a live drug-controlled vaccine against bovine babesiosis using the Mongolian gerbil, Meriones unguiculatus.

This study investigated the practicality and potential of the gerbil, Meriones unguiculatus, as a source of live Babesia divergens vaccine and also as a model for the use of the vaccine in cattle. A series of experiments with gerbils concerning vaccine infectivity, immunogenicity and safety were carried out. It was concluded that the use of RPMI medium/40% foetal calf serum as a diluent improved vaccine infectivity, but that the parasitaemia of the blood obtained from donor gerbils had little or no effect. The immunostimulants levamisole and killed Corynebacterium parvum improved vaccine immunogenicity and it was also shown that the subcutaneous route of infection resulted in the greatest host response. Control of vaccine virulence with drugs was only possible when drugs with prophylactic properties, such as imidocarb and long-acting oxytetracycline, were used. More studies are required on all these topics, particularly with regard to their applicability to cattle, and also concerning the possible attenuation of the parasite by manipulation in the gerbil host.     

Effects of rapid passage in the gerbil (Meriones unguiculatus) on the course of infection of the bovine piroplasm Babesia divergens in splenectomised calves

The antigenicity and virulence of the cattle piroplasm Babesia divergens was compared in splenectomised calves before and after adaptation to the gerbil. It was apparent that adaptation of B divergens to the gerbil did not result in attenuation of the parasite in cattle. The gerbil-adapted parasite had a shorter prepatent period, produced higher parasitaemias, and more severe haematological and biochemical changes than the initial bovine strain. The antigenicity of the parasite was unchanged by gerbil passage. The changes in the virulence of the gerbil-adapted parasite were attributed to selection of a rapidly replicating parasite.     

Supplementation of heterologous complement induces anti-Thy-1.1 nephritis in the Mongolian gerbil (Meriones unguiculatus)

Anti-Thy-1.1 nephritis in the rat is a popular experimental model for mesangial proliferative glomerulonephritis (GN). This model is characterized by direct binding of anti-Thy-1.1 antibody with Thy-1.1 antigen expressed on mesangial cells (MCs) of glomeruli in the rat. A single injection of anti-rat thymocyte serum (ARTS) results in GN with proteinuria and extensive mesangiolysis. Development of mesangiolysis and proteinuria are complement-dependent. We previously demonstrated Thy-1.1 antigen, similar to the rat, in thymocytes, brain cells and MCs of the kidney in the Mongolian gerbil (MG). In this study, we attempted to develop a MG nephritis model, but an injection of ARTS did not induce GN. An additional injection of guinea pig serum as a complement after ARTS injection resulted in anti-Thy-1.1 nephritis in MG. Degeneration of MCs and neutrophil infiltration were observed 1 hr after GP serum injection. Mesangiolysis and fibrin exudation occurred 12 hr after the injection and MC proliferation was apparent 7 days after the injection. In the complement-dependent hemolytic test, MG serum could not hemolyze sheep erythrocytes. These results suggested low activity, or depletion of some factors, in complements of MG serum.     

Poorly differentiated cutaneous carcinoma of non-sebaceous origin in a 3-year-old Mongolian gerbil (Meriones unguiculatus)

A 3-year-old female gerbil developed a non-healing skin wound due to a malignant neoplasm. Histology, immunohistochemistry (cytokeratin 19 positive; vimentin, estrogen, and progesterone receptor negative), and electron microscopy (no desmosomes or melanosomes) revealed an undifferentiated carcinoma with pulmonary metastasis. Unlike in previous reports, it did not arise from the abdominal pad’s sebaceous gland.     

Cerebellar ataxia due to Toxocara infection in Mongolian gerbils,Meriones unguiculatus

We assessed the usefulness of gerbils as an experimental model for neurologic toxocarosis. Mongolian gerbils, Meriones unguiculatus, infected with Toxocara canis or Toxocara cati (1000 eggs/gerbil) showed progressive neurologic disorders from 50 days after infection in T. canis-infected gerbils or from 120 days after infection in T. cati-infected gerbils. The incidence of the onset was 6 of the 13 gerbils (49%) in the T. canis-gerbils and 5 of the 7 gerbils (71%) in the T. cati-gerbils. Histopathologically, the cerebellum was the most affected in both groups. We observed loss of Purkinje cells, glial nerve fibers, and nerve sheaths. We also found foci consisting of aggregated macrophages scattered in the white matter of the cerebellum. The affected gerbils showed ataxia and ultimately died of cachexia. Our findings suggest that irreversible neurologic toxocarosis in gerbils can be induced by infection with either T. canis or T. cati.     

In vitro progesterone production of corpus luteum in pregnant Mongolian gerbils (Meriones unguiculatus)

The present study was conducted to determine the in vitro progesterone (P₄) production by Mongolian gerbil (Meriones unguiculatus) corpus luteum (CL) on days 9, 18 and 22 of pregnancy in comparison with rat CL on days 9 and 16. The culture medium of one CL per well of a 96‐well plate was changed after 4 h incubation and the CL were further cultured for 24 h. The P₄ in serum and in the medium were measured by enzyme immunoassay system. Gerbil CL weight measured after 24 h incubation was at the same level for all 3 days of pregnancy and significantly different from that of the rat CL for the 2 days of pregnancy. The serum P₄ level in gerbils was high in early pregnancy and decreased in advanced days of pregnancy. In contrast, the P₄ in rats was higher in the latter half of pregnancy. In vitro P₄ production by a single CL after 4 h incubation decreased significantly during the days of pregnancy in both the gerbil and the rat. P₄ values produced by the gerbil CL for 24 h incubation were significantly higher than after 4 h incubation in all 3 days of pregnancy, however, there was no difference between the two incubation time groups in the rat. In vitro P₄ production by CL correlated well with the serum P₄ level on the 3 days of pregnancy in the gerbil. In the rat, however, the relationship was inversed between the two parameters. The correlation in this experiment suggested that CL is the sole source of P₄ for the maintenance of the gerbil pregnancy, with the result of abortion after an ovariectomy on day 20 of pregnancy as described in our previous report.     

Efficacy of paraherquamide against immature Trichostrongylus colubriformis in the gerbil (Meriones unguiculatus)

Paraherquamide was 98 to 100 per cent effective against six-day-old Trichostrongylus colubriformis infections in gerbils when given as single oral doses of 1.56 mg kg-1 and above. Doses of 0.78 or 0.39 mg kg-1 were 96 and 66 per cent effective, respectively. A single oral dose of 200 mg kg-1 was well tolerated.     

Clinical comparison of isoflurane and sevoflurane anaesthesia in the gerbil (Meriones unguiculatus)

The objective of this study was a comparison of the volatile anaesthetics isoflurane and sevoflurane in terms of their clinical effects in gerbils (Meriones unguiculatus) (n=12 each). Induction of anaesthesia was performed in a body chamber with an anaesthetic concentration of 4.0 Vol.% at an oxygen flow of 500 ml/min for isoflurane and 8.0 Vol.% at an oxygen flow of 1000 ml/min for sevoflurane, respectively. Anaesthesia was maintained via nose cone with an anaesthetic concentration of 2.8 to 3.2 Vol.% at an oxygen flow of 200 ml/min for isoflurane and 5.0 to 5.2 Vol.% at an oxygen flow of 400 ml/min for sevoflurane. Those anaesthetic concentrations ensured reflex status conform with surgical tolerance. In spite of its higher blood-gas coefficient induction time was slightly faster for isoflurane. Recovery time was significantly longer in the isoflurane group than it was in the sevoflurane group. Both inhalants caused respiratory depression. Respiratory rate was lower in sevoflurane animals compared to isoflurane. The animals were positioned on a heating pad immediately after induction, thus a decrease of the body temperature could be prevented. Both inhalants can be recommended for usage in gerbils. Sevoflurane showed no clinical benefit compared to isoflurane.     

Cellular Composition of the Pancreatic Islets in Mongolian gerbils (Meriones unguiculatus)

In Mongolian gerbil, morphological changes with age in the content of endocrine cells in the pancreatic islets were analyzed and their relation evaluated by an oral glucose tolerance test. The glucose level 2 hours after glucose administration was 125 ± 5 mg./dl in the young group and 103 ± 4 mg./dl in the old group. In the dorsal portion, B cells were mainly observed in the central area of the islets, surrounded by circular layers of A cells in the peripheral area. Between the A cells and B cells, D cells were scattered or present in layers. A few PP cells were present in the peripheral area of the islets. In the ventral portion, only a few A cells were observed in the peripheral area of the islets, and B cells were surrounded by PP cells. Secretory granules of A cells generally had an electron-dense spherical core in the limiting membrane. The halo between the limiting membrane and core in A cells was narrower than that in B cells. Secretory granules of B cells were larger than those in A cells, and the core was less electron-dense, and the halo was wider. Secretory granules of D cells were similar in size to those of A cells; the core showed low electron density, and the halo was very narrow. Granules of PP cells resembled those of A cells, but the electron density of the core was slightly lower. The gerbils showed changes in glucose tolerance, the size of the pancreatic islets, the percentage of B cells, and of A cells in the dorsal portion with age. The content of endocrine cells in Mongolian gerbils was similar to that in humans, rats, mice, and hamsters.     

Undifferentiated Sarcoma of the Salivary Gland in a Mongolian Gerbil (Meriones unguiculatus)

A subcutaneous mass was found in the lower ventral neck region of a 55-week-old male Mongolian gerbil (Meriones unguiculatus). Histopathologically, the mass involved salivary glands and featured diffuse proliferation of pleomorphic neoplastic cells with large necrotic foci. The lesion was well demarcated from the surrounding tissue, although invasive growth to fibrous septa was occasionally observed. The neoplastic cells were mainly arranged in irregular sheets with severe cellular atypia, round to oval nuclei and varying amounts of eosinophilic cytoplasm. Mitotic figures and multinucleated giant cells were frequent. Immunohistochemical analysis revealed that the neoplastic cells were strongly positive for vimentin and S-100 and negative for NSE, cytokeratin, α-SMA, c-kit, factor VIII, CD34, α-1-antitrypsin, lysozyme and MSR-A. Based on the results, the mass was diagnosed as an undifferentiated sarcoma of the salivary gland. To the best of our knowledge, this is the first report of such a tumor in Mongolian gerbils.     

Comparative studies on distribution and fine morphology of the intestinal Peyer's patches in Mongolian gerbils (Meriones unguiculatus) and mice.

The intestinal Peyer's patches (PP) were significantly different in distribution and fine morphology between Mongolian gerbils and mice. The PP of gerbils were evenly distributed from the duodenum to ileum, whereas the PP of mice were more densely distributed in the lower ileum than other parts of the intestine. In gerbils, each PP had a much greater number of lymphoid follicles than in mice, although the total number of PP was smaller. Electron microscopy revealed that the PP dome of gerbils was covered with two types of epithelial cells, one with shorter microvilli and the other with longer ones, whereas the epithelial cells of the murine PP dome was uniform-shaped with numerous medium-sized microvilli. Some dome absorptive cells of gerbils were morphologically similar to poorly differentiated crypt cells of mice, suggesting to be immature M cells.     

In vivo transfer of an Escherichia coli enterotoxin plasmid possessing genes for drug resistance.

Experiments were conducted to study transfer of an enterotoxin (Ent) plasmid from a porcine enteropathogenic Escherichia coli to an E coli K12 strain in the intestine of newly weaned pigs. The Ent plasmid carried genes for resistance to tetracycline, streptomycin, and sulfonamides, thereby permitting a selection for tetracycline-resistant exconjugants in the feces of the pigs. In vivo transfer of the Ent plasmid was demonstrated to occur when the pigs were given large oral inocula of donor and recipient cultures, 1 hour apart. Differences in extent of transfer were not detected in pigs given antibiotic-free feed compared with littermates on feed containing oxytetracycline at 50 g/ton. In one experiment, tetracycline-resistant Ent- exconjugants were found which appeared to have received an R plasmid from an enteropathogenic type of E coli resident in the intestine.     

In vivo assessment of drug sensitivity of African trypanosomes using the akinetoplastic induction test.

Following treatment of mice infected with Trypanosoma congolense or T brucei brucei with various doses of isometamidium chloride or diminazene aceturate, the induction of akinetoplastic (AK) forms was observed in the trypomastigotes of both species within 10 hours of drug administration. The levels of AK-induction were closely correlated with the levels of resistance to each compound found using a standard in vivo drug assay in mice. In general, ineffective doses of either compound conferred AK-induction rates of less than 30 per cent; relapsing cases had between 30 and 50 per cent while curative doses had AK-induction rates of 50 per cent or more. In vivo determination of AK-induction rates using ordinary light microscopy is thus a potentially feasible alternative indicator to the conventional use of mice infection and treatment methods for assessing drug sensitivity in African trypanosomes.     

Individual heterogeneity in erythrocyte susceptibility to Babesia divergens is a critical factor for the outcome of experimental spleen-intact sheep infections

Susceptibility of sheep erythrocytes to Babesia divergens was investigated in vitro and a high inter-individual variability in their ability to support parasite population development was demonstrated, with some individuals having refractory red blood cells (RBC). As neither changes in growth conditions nor the use of different B. divergens strains influenced the level of susceptibility, the main factor postulated for this variability is the erythrocyte itself. Sheep therefore represent an excellent in vitro model to study the parasite-erythrocyte interaction. In addition, the existence of refractory RBC should help in the identification of the erythrocyte components required for B. divergens development. Experimental infections were carried out on spleen-intact sheep characterized by refractory or fully susceptible erythrocyte types. These differences translated into the successful infection of only those animals with susceptible erythrocytes: infected animals showed no clinical signs, but maintained an asymptomatic persistent infection, as usually observed in the natural bovine host. Sheep therefore represent model organisms that can allow us to study interactions between B. divergens and its vertebrate host at different levels of biological organisation, from the target cell to the intact animal, and represent an experimental infection model of concomitant immunity. Only a low percentage (13%) of the sheep population tested possessed susceptible erythrocytes and the potential role of sheep as a natural host or reservoir of B. divergens is discussed.     

Detection of macrolide resistance genes,ermC and ermB,in Japanese honey using real-time PCR assays

American foulbrood (AFB) is a honeybee disease caused by Paenibacillus larvae, and tylosin is used as the prophylactic in Japan. Honey contains macrolide-resistant bacteria that are a potential source of genes that may confer tylosin resistance to P. larvae. To investigate the potential risk of such genes in Japanese honey, we developed real-time PCR assays for the detection of important macrolide resistance genes, ermC and ermB, and analyzed 116 Japanese honey samples with known contamination status of P. larvae. Consequently, 91.38% of samples contained ermC and/or ermB, and 71.55% of samples contained both ermC and P. larvae, suggesting the possible emergence of tylosin-resistant P. larvae in Japan. Therefore, judicious use of the prophylactic is essential in maintaining its effectiveness.