This study detected a total of 52 cases of CDV either tested by RT-PCR (73%, 38/52) only or IHC (63%, 33/52). The positive rate of RT-PCR diagnosis was 100%
for the 38 cases for which specimens were available. The positive rate of immunohistochemical labeling was 97% (33 of 34) for 34 cases with specimens available for IHC. For cases combined tested both by IHC and RT-PCR, what was 95
% (19/20) agreement, only one case showed negative ( Table 7).
Of the 52 cases, 65% (34/52) had CNS demyelination confirmed by H&E or LFB-CEV. Of these demyelination cases, 68% (23/34) were RT-PCR positive for CDV, and 82% (28/34) were IHC positive for CDV. Of the demyelination cases, 97%
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interpreted for diagnosis of CDV than was routine H&E staining (Fig. 1A,C). In general, the cerebrum (Fig. 1B), cerebellar white matter, 4th ventricle, lung(Fig. 1D), urinary bladder, and spleen were likely to be IHC-positive in CDV-infected dogs, and characteristic of IHC labelling as reported previously14 and had at least six of pathology findings consistent with canine distemper (Table. 2).
PATHOLOGY
All cases were examined by histology. Totally, the most common CDV diagnostic lesions of the dogs were lymphoid depletion, INIB/ICIB, interstitial pneumonia, and CNS demyelination in decreasing order. The positive rates were 79% (41/52), 75%
(39/52), 71% (37/52), and 65% (34/52), respectively. However, the distribution of the nine CDV lesions varied considerably between the two groups of dogs. In the clinic dogs, the three most common lesions were lymphoid depletion (96% of the dogs), intranuclear inclusion bodies (87%), and interstitial pneumonia (81%). In the shelter dogs, these three lesions were also the most common, although the percentage of affected dogs was lower in each case, 50%, 55%, and 55%, respectively. In both groups of dogs, the least common lesion was foot pad hyperkeratosis, which affected 28% of the clinic dogs and 20% of the shelter dogs (Table 2).
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(Table 3, 4). Two types of inclusion bodies were noted. One type was characterized as one, large, round or ovoid, distinct, homogenous, intensely eosinophilic INIB. The most common sites were cerebral astrocytes (Figure 2A), cerebellar white matter astrocytes, ependymal cells lining the 4th ventricle, the periphery of the splenic central arteries (Figure 2B), macrophages and lymphocytes in the lymph nodes, tonsil lymphoid tissues, pulmonary macrophages and bronchiolar epithelium, renal pelvis, urinary bladder epithelium (Figure 2C) and gastric glandular cells (Figure 2D).
Another type was one or several smaller eosinophilic ICIB, seen chiefly in the renal pelvis, urinary bladder epithelium (Figure 2C) and skin epidermis.
The 29 co-infections and associated lesions with CDV infection are recorded and compared in two groups (Table 6). Most common co-infections were Cestodiasis, and Dirofilariasis. However, the co-infections included the basophilic adenovirus inclusion bodies (type II) (Figure 3A), adenovirus inclusion bodies (type I), coccidiosis, babesiosis, hemobartonellosis, aspergillosis and suppurative bronchitis were found in clinic group only. The parasitic infestation was more common in the older shelter group. Other common associated lesions included: 13.
meningoencephalitis, 16 suppurative bronchopneumonia 22. extramedullary hematopoiesis (EMH) and 25. interstitial nephritis showed high incidence in shelter
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The microfilariae of Dirofilaria immitus were very common in both groups. They were found in pulmonary capillaries but were also noted in the interstitium with mixed leukocytic aggregates and erythrocytes (Figure 3B). Some rarely found infections included adenovirus infection in hepatic cells (type I) (Figure 3C), babesiosis as well as hemobartonellosis were also found in this study. One case showed disseminated pyogranulomatous pneumonia with numerous, intralesional banana-shaped protozoal tachyzoites strongly suggested toxoplasma-like protozoa (Figure 3D) infection. One case showed disseminated pyogranulomatous lesions in the cerebrum, kidney, liver and serosa of gastrointestinal tract with intralesional PAS-positive thin septae hyphae, and Y-shaped branching, strongly suggested Aspergillosis infection.
DATA ANALYSIS
The clinic group showed significantly difference of younger age than the shelter group. There were significant differences in lesion frequencies, or disease pattern, between the clinic and shelter groups. The occurrence of 6 of the 9 diagnostic CDV lesions was significantly higher in the clinic dogs than in the shelter dogs: intranuclear inclusion bodies; demyelination in the cerebrum, cerebellum and brain stem;
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lesions, catarral enteritis, hyperkeratosis, and syncytium, there was no significant difference in frequency of occurrence between the two groups of dogs.
In the clinic group (n = 32), inclusion bodies were most common in urinary bladder (68%), lymph node or spleen (65%), lung (62%) and cerebellum (56%), with 50%
positive rate of detection overall in this group. In the shelter dogs, the most common sites for the inclusion bodies were the lymphoid tissues and cerebellum (both 35%), followed by the brain stem (30%); the least common sites were the alimentary tract and the skin (both 10%). The inclusion bodies of shelter group were identified in the same tissues but were significantly less common, with overall 23% detection rate (Table 5).
The occurrence of the INIB/ICIB in four of the nine sites was significantly higher in the clinic dogs than in the shelter group, namely in the urinary bladder, lymphoid tissues, lung, cerebellum, and alimentary tract. For the remaining tissues, there were no significant differences in occurrence between the two groups of dogs. Furthermore, the mean number of inclusion body positive tissues per dog in the clinic group (4.6 ± 0.6), was significantly higher than that in the shelter group (1.8 ±0.6) by two-tailed Student’s t test.
However, there was no significant difference in the mean number of co-infections
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1.5 in the clinic group). Furthermore, the occurrence of 28 of the 29 co-infections and associated lesions was not significantly different between the two groups by two-tailed Student’s t test; the exception was interstitial nephritis, which was significantly more common in the shelter dogs than in the clinic dogs.
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To the best of our knowledge, this study is a first comprehensive, retrospective study of the pathology of CDV infection, that includes statistical analysis of the distribution of lesion, inclusion bodies, and co-infections in two contrasting groups of dogs. The principal findings were: (i) The CDV diagnostic lesions including lymphoid depletion, frequency of INIB/ICIB, pneumonia and demyelination, were significant different between younger clinic and older shelter group, (ii) The high occurrence of INIB/ICIB of CDV in Taiwan was noted in spleen, lymph nodes, lung and gastric mucosa epithelium. However, its positive rates were high in younger clinic group. In this study, most of the cases were “non-enteritis group” which showed pneumonia (55 to 81%) and degenerative demyelinating lesions in the CNS (20-78%) depending on sheler or clinic group (Table 2). These results were quite similar as “non-enteritis group” of canine distemper in Japan. Those cases were suspected different antigenically from the vaccine strain (Okita et al.,1997). However, the high occurrence of INIB/ICIB in urinary bladder (26-68%), spleen and lymph nodes (35-65%), lung (15-62%) and gastric mucosa epithelium(10-43%) in this study (Table 5) were similar to “enteritis group”of CDV in Japan (Okita et al.,1997). (iii) The positive rates of RT-PCR, IHC labeling, CNS demyelination and INIB/ICIB were 73%, 63%, 65%, 75%, respectively. The positive rates of either IHC labeling only
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case blocks are damage (35% not available, n =18), so we did not test IHC and few cases did not had RT-PCR results (27% not available, n=14) (Table 7).
When all 52 dogs were considered together, the positive rates of distemper inclusion bodies was 75%, which was slightly higher than previously reported;
previous studies have ranged from 17 to 72%.8,19,23,29 However, intranuclear inclusion bodies were chiefly finding in this study rather than intracytoplasmic inclusion bodies in neuron.25 The occurrence of inclusion bodies in 4 of 9 tissues was significantly higher in the clinic dogs than in the shelter dogs between our two groups of dogs (Table 5). This may reflect the fact that the clinic group was acute stage of CDV infection, and younger, contrasting with the chronic stage and older shelter group.3,27
However, 75% of these dogs in clinic group are under 6-month-old were noted in the present Taiwanese study. The antigenic genetic variation of field CDV in Taiwan may account for these cases. Because the H gene of field isolated CDV in Taiwan had 10% amino acid variation from the vaccine Onderstepoort strain.13 The present study showed that the immunohistochemical detection of CDV antigen in tissue sections was superior to the demonstration of inclusion bodies or syncytial cells for the diagnosis of canine distemper. This finding is consistent with previous reports.6,14,19,20 Among our findings of co-infections and associated lesions of particular note, was
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dogs (Table 6). A similar high value was also reported.5,6 The primary CDV infection, which might causes immunosuppression, and have predisposed the dog to secondary Tyzzer’s disease and intestinal coccidiosis;9 other viruses or with Bordetella