Histopathology of incidental findings in cynomolgus monkeys ( macaca fascicularis ) used in toxicity studies

The purpose of our publication is to widely communicate pictures of spontaneous findings occurring in cynomolgus monkeys. Focal lymphoplasmacytic infiltration is commonly seen in the general organs. The frequency and severity of these lesions may be influenced by the administration of drugs with an effect on the immune system. Lymphoplasmacytic infiltration in the lamina propria of the stomach is also frequently seen in cynomolgus monkeys, and it is caused mainly by a Helicobacter pylori infection. Various degrees of brown pigments are observed in various organs, and it is possible to distinguish the material of the pigments by its morphological features and site. A focal/segmental glomerular lesion is occasionally seen in a section of the kidney, and the minimal lesion has no influence on the urinalysis. We showed the common glomerular lesions in HE-stained sections, as well as in PAM- or PAS-stained sections, for understanding the details. Young and pubertal monkeys are usually used in toxicity studies; therefore, understanding various maturation stages of the genital system is important. In particular, the female genital system needs to be understood in the morphology, because their cyclic changes are different from other laboratory animals. Thus, we present the normal features of the cyclic changes of the female genital organs. Furthermore, we provide more information on spontaneous findings in cynomolgus monkeys for exact diagnoses in toxicity studies.     

Porencephaly in a cynomolgus monkey ( macaca fascicularis )

Porencephaly was observed in a female cynomolgus monkey (Macaca fascicularis) aged 5 years and 7 months. The cerebral hemisphere exhibited diffuse brownish excavation with partial defects of the full thickness of the hemispheric wall, and it constituted open channels between the lateral ventricular system and arachnoid space. In addition, the bilateral occipital lobe was slightly atrophied. Histopathologically, fibrous gliosis was spread out around the excavation area and its periphery. In the roof tissue over the cavity, small round cells were arranged in the laminae. They seemed to be neural or glial precursor cells because they were positive for Musashi 1 and negative for NeuN and GFAP. In the area of fibrous gliosis, hemosiderin or lipofuscin were deposited in the macrophages, and activated astroglias were observed extensively around the excavation area.     

ELISAs based on rNcGRA7 and rNcSAG1 antigens as an indicator of Neospora caninum activation

Bovine abortion caused by the Apicomplexa parasite Neospora caninum is a major economical problem in the livestock industry worldwide. However, the relationship between N. caninum infection and abortion is still unknown. Our study aimed to elucidate the relationship between parasite-specific antibody responses, parasite stages and abortion. In experimentally infected cattle, anti-NcGRA7 IgG1 antibody was predominantly detected during the acute infection stage, while the production of anti-NcSAG1 IgG1 antibody was observed during both acute and chronic stages. Furthermore, levels of anti-NcGRA7 IgG2 antibody were lower than those of anti-NcSAG1 IgG2 antibody. When tested on cattle with Neospora-associated abortion, positive rates of the anti-NcGRA7 IgG2 antibody were significantly lower than those of the anti-NcSAG1 antibody, although there was no difference in IgG1 antibody-positive rates between the two antigens. In addition, anti-NcGRA7 IgG2 antibodies were not detected in cattle for more than 30days after abortion. Our results suggested that anti-NcGRA7 and anti-NcSAG1 antibodies are suitable indicators for the activation stage of N. caninum infection and broad detection of the infection, respectively. In conclusion, the use of recombinant NcGRA7 and NcSAG1-based ELISAs will be useful for evaluating the abortion risk associated with N. caninum infection.     

Nodulation during vegetative growth of soybean stage does not affect the susceptibility to red crown rot caused by Calonectria ilicicola

Soybean cultivar Enrei and its derivative lines, non-nodulating En1282 and supernodulating Sakukei 4, were grown in an experiment field naturally infested with the red crown rot fungus, Calonectria ilicicola, to determine whether nodulation elicits the disease. Infection frequency by the fungus invariably increased with time and reached about 100% 6 weeks after sowing. Disease severity was invariably slight until 8 weeks after sowing. At harvest, however, disease severity was lowest in En1282 and highest in Sakukei 4. Enrei was intermediately susceptible. Sakukei 4 produced many more nodules than Enrei, while En1282 completely failed to nodulate. For three other pairs of nodulating and non-nodulating isogenic lines, all non-nodulating lines had lower severity than their nodulating counterparts at harvest. Pot experiments with Enrei and its derivatives revealed that inoculation with Bradyrhizobium sp. did not affect disease severity during the vegetative stage of plant growth, i.e., 6 weeks after sowing. These results suggested that the production of nodules in the vegetative stage is not, at least directly, related to susceptibility to the disease, but physiologic changes after anthesis vary depending on the nodulation traits in the respective soybean lines, resulting in different levels of susceptibility to red crown rot.     

Functional Analysis of an ATP-Binding Cassette Transporter Gene in Botrytis cinerea by Gene Disruption

The BMR1 gene encoding an ABC transporter was cloned from Botrytis cinerea. To examine the function of BMR1 in B. cinerea, we isolated BMR1-deficient mutants after gene disruption. Disruption vector pBcDF4 was constructed by replacing the BMR1-coding region with a hygromycin B phosphotransferase gene (hph) cassette. The BMR1 disruptants had an increased sensitivity to polyoxin and iprobenfos. Polyoxin and iprobenfos, structurally unrelated compounds, may therefore be substrates of BMR1. Received 18 December 2000/ Accepted in revised form 18 April 2001     

A Hepatocellular Adenoma in a Diet-induced Obese Mouse

A hepatic nodule was noted in a C57BL/6J mouse with diet-induced obesity at 53 weeks of age. Macroscopically, a protruding yellowish white nodule was observed on the visceral surface of the left lateral lobe. Light microscopy demonstrated clear demarcation from the compressed adjacent parenchyma, with loss of the distinct lobular pattern. The proliferating cells of the lesion varied in shape and showed cellular atypia and prominent nucleoli along with vacuoles of various sizes. Some of the cells contained various-sized eosinophilic inclusion bodies in their cytoplasm, and electron microscopy revealed the presence of lipid droplets in the rough endoplasmic reticulum. Eosinophilic inclusions were observed as electron dense granular material in the rough endoplasmic reticulum, with one or a few low density central cores. A diagnosis of hepatocellular adenoma was made based on these findings.     

Diagnosis of histoplasmosis by detection of the internal transcribed spacer region of fungal rRNA gene from a paraffin-embedded skin sample from a dog in Japan

The lesions of histoplasmosis in dogs in Japan differ from those in dogs in North America. Affected dogs in Japan have had multiple granulomatous or ulcerated foci in skin or gingiva and have not had pulmonary or gastrointestinal lesions. The present report introduces a polymerase chain reaction (PCR) diagnosis of canine histoplasmosis and the characteristic of disease in Japan. The surgically removed skin ulcerate samples from a 5-years-old female Shiba-inu native to Japan without traveling out of the country were evaluated. Tissue samples had many yeast-like organisms in the macrophages. DNA was extracted from paraffin-embedded tissue samples. A nested PCR technique was applied. The detected sequence of the internal transcribed spacer of ribosomal RNA gene had 99.7% in homology with Ajellomyces capsulatus (the teleomorph of Histoplasma capsulatum). Clinical manifestations, historical background of equine epizootic lymphangitis in Japan, and a human autochthonous case of histoplasmosis farciminosi indicated that this dog might have been infected with H. capsulatum var. farciminosum as a heteroecism.     

Effect of isotonic and hypotonic lactated ringer's solutions with dextrose intravenously administered to dehydrated heifers

To determine the effects of rapid infusion of essential fluids in a volume of hypotonic lactated Ringer's solution, the central venous pressure (CVP) and acid-base equilibrium were investigated in to mildly dehydrated heifers. Mild dehydration was induced in 9 Holstein heifers by withholding food and water until 7.0+/-5.7% of plasma volume had been lost. The heifers were randomly assigned to the ILG (lactated Ringer's + 5% dextrose), HLG (1/2 lactated Ringer's + 2.2% dextrose) or HRG (1/2 Ringer's + 2.5% dextrose) groups with 3 heifers in each group. Heifers received 30 ml/kg of one of the fluids, at a flow rate of 20 ml/kg/hr. The rapid intravenous (IV) infusions of HLG and HRG used in this study were found to be safe and effective in increasing plasma volume without increasing CVP, even though the infusion was given to the jugular vein at a dosage of 30 ml/kg. However, ILG infusion induced progressive increases in CVP, reaching 9.0+/-2.0 mmHg. No clinical signs, such as moist rales on auscultation, moist cough, jugular vein congestion, ophthalmoptosis, salivation or arrhythmia, were observed throughout the fluid infusion. The relative changes in base excess (rBE) for the ILG and HRG groups were significantly decreased until the end of fluid infusion. As for the HLG group, rBE slightly decreased until the end of the fluid infusion. Then the values significantly increased and exceeded the pre-infusion value at the end of the experiment. While IV infusion of HLG inhibited acidification caused by dilution, HRG infusion induced diluted acidification. It is suggested that HLG infusion should be examined as a treatment for cattle with dehydration and moderate metabolic acidosis, since rapid infusion of HLG may be more beneficial for rehydrating cattle with metabolic acidosis than current treatment.     

A new ENU-induced mutant mouse with defective spermatogenesis caused by a nonsense mutation of the syntaxin 2/epimorphin (Stx2/Epim) gene

Repro34 is an N-ethyl-N-nitrosourea (ENU)-induced mutation in mice showing male-specific infertility caused by defective spermatogenesis. In the present study, we investigated pathogenesis and molecular lesions in relation to spermatogenesis in the repro34/repro34 homozygous mouse. Histological examination of the testis showed that the seminiferous epithelium of the repro34/repro34 mouse contained spermatogonia and spermatocytes but no round and elongating spermatids. Instead of these haploid cells, multinucleated giant cells occupied the niche of the seminiferous tubules. Immunohistochemical staining for Hsc70t, an elongating spermatid specific protein, confirmed the absence of elongating spermatids. Furthermore, RT-PCR showed that there were significantly reduced expressions of the marker genes specifically expressed in the spermatid and that there was no difference in the expressions of the spermatocyte specific marker genes. These findings indicated interruption of the spermatogenesis during transition from the spermatocyte to spermatid in the repro34/repro34 mouse. The repro34 locus has been mapped on a 7.0-Mb region of mouse chromosome 5 containing the Syntaxin 2/Epimorphin (Stx2/Epim) gene, and targeted disruption of this gene has been reported to cause defective spermatogenesis. We therefore sequenced the entire coding region of the Stx2/Epim gene and found a nucleotide substitution that results in a nonsense mutation of this gene. The expression pattern of the Stx2/Epim gene during the first wave of spermatogenesis, increased expression at later stages of spermatogenesis, was in agreement with the affected phase of spermatogenesis in the adult repro34/repro34 testis. We therefore concluded that the male infertility of the repro34/repro34 mouse is caused by the interruption of spermatogenesis during transition from the spermatocyte to spermatid and that the nonsense mutation of the Stx2/Epim gene is responsible for the interruption of spermatogenesis.