Analysis of the breast milk of giant pandas (Ailuropoda melanoleuca) and the preparation of substitutes.

The first milk substitute for giant panda cubs was developed in 1988 based on limited data about giant panda breast milk and that of certain types of bear. Mixtures of other formulas have also been fed to cubs at some facilities. However, they are not of sufficient nutritional quality for promoting growth in panda cubs. Here, we report analysis of giant panda breast milk and propose new milk substitutes for cubs, which were developed based on the results of our analysis. The Chengdu Research Base of Giant Panda Breeding obtained breast milk samples from three giant pandas. Up to 30 ml of breast milk were collected from each mother by hand. Then, the milk samples were frozen and sent to Nihon University. The levels of protein, fat, carbohydrates, ash, moisture, vitamins, minerals, total amino acids, fatty acids, lactose and other carbohydrates in the milk were analyzed. The breast milk samples exhibited the following nutritional values: protein: 6.6-8.5%, fat: 6.9-16.4%, carbohydrates: 2.5-9.1%, ash: 0.9-1.0% and moisture: 67-83%. We designed two kinds of milk substitutes based on the data obtained and the nutritional requirements of dogs, cats and rodents. The nutritional composition of the milk substitutes for the first and second stages was as follows: protein: 38 and 26%, fat: 40 and 40%, carbohydrates: 13 and 25%, ash: 6 and 6% and moisture: 3 and 3%, respectively. In addition, the substitutes contained vitamins, minerals, taurine, docosahexaenoic acid, lactoferrin, nucleotides and other nutrients.

Reduced production of polymeric immunoglobulin receptor in murine dextran sodium sulfate-induced colitis.

Polymeric immunoglobulin receptor (pIgR) plays an intrinsic role in protecting the intestinal tract from invading pathogens. In the present study, we observed a decrease in pIgR in colon lysate from mice with dextran sodium sulfate (DSS) colitis. A decrease in pIgR was detected in both mRNA and protein levels. Histologic examinations revealed marked destruction of intestinal epithelial cells (IECs), and only a small number of regenerating IECs expressed pIgR. These results suggest that the decrease in pIgR was due to the destruction of IECs. Because activation of toll-like receptor 3 slows the progression of DSS colitis, we injected polyriboinosinic: polyribocytidylic acid (poly I:C) intraperitoneally and observed the correlation between pIgR level and severity of DSS colitis. Poly I:C markedly decreased progression of DSS colitis, and pIgR levels significantly recovered. Furthermore, we found that expressions of IFN-γ and TNF-α were higher in DSS colitis. These results indicate that the decrease in pIgR was not compensated for by increased expression of these cytokines. In sum, our findings show that pIgR levels vary according to the severity of DSS colitis and that these changes might be useful as a biomarker of the severity of inflammatory bowel disease.

Excretion of bisphenol A into rat milk.

Bisphenol A (BPA), an endocrine-disrupting chemical, is widely used in the production of polycarbonate plastic and epoxy resins. This study analyzed the BPA concentration in rat milk, in order to assess the risk of BPA transfer to the offspring via milk. The rats ingested BPA by oral administration or by drinking the water in a polycarbonate bottle, and the milk samples were collected using an automated experimental milker. The BPA concentration in the samples of milk, drinking water, and food was analyzed by LC/MS. In the case of milk samples obtained from rats injected with BPA at 2, 4, 8, and 24 h prior to milking, the BPA concentrations were 0.462 +/- 0.182 ppm, 0.138 +/- 0.0185 ppm, 0.080 +/- 0.0197 ppm, and 0.0232 +/- 0.0051 ppm, respectively. Also, in the cases of the water sample left in polycarbonate bottle and the milk sample obtained from rats provided it as drinking water, the concentrations of BPA were 0.000332 +/- 0.00015 ppm and 0.0184 +/- 0.0050 ppm, respectively. The results indicate that the BPA administered to the dams was transferred to their milk, and that BPA concentration in milk was higher at the early period after the single bolus dose. Additionally, these results reveal that sequential elution of BPA from polycarbonate containers in a much diluted form would undergo bioaccumulation in dams and likely be transferred to pups via milk in a much concentrated form.

Antifungal activity of itraconazole and voriconazole against clinical isolates obtained from animals with mycoses.

Animal mycosis, particularly deep mycosis, is one of the most challenging conditions encountered by veterinarians. Pathogens causing mycotic infections in animals include fungi such as Cryptococcus neoformans, Candida spp., and Aspergillus spp. The antifungal drugs used for the treatment of deep mycoses in animals as well as humans are polyenes and azoles. However, the sensitivity of clinical isolates obtained from animals toward these drugs has rarely been assayed. In this study, the antifungal activities of itraconazole and voriconazole against clinical isolates of C. neoformans, Candida spp., and A. fumigatus isolated from animals with mycoses were examined using the broth microdilution method performed according to the guidelines provided by the Clinical and Laboratory Standards Institute. The minimum inhibitory concentrations (MICs) of itraconazole toward the C. neoformans, Candida spp., and A. fumigatus isolates were 0.125 - 1, 0.125 - 2, and 0.25 - 2 microg/ml, respectively, and those of voriconazole were 0.0625 - 0.5, < or =0.0313 - 0.0625, and 0.0625 - 1 microg/ml, respectively. The results of the MIC analyses implied that the fungal isolates obtained from infected animals exhibit an equivalent degree of susceptibility to itraconazole and voriconazole, as is observed in the case of isolates obtained from humans. The appropriate antifungal therapeutic strategy for the treatment of mycoses in animals must be selected taking into consideration the host immune status and organ function as well as the in vitro sensitivity of the pathogens to antifungal drugs.

Microsatellite variability reveals the necessity for genetic input from wild giant pandas (Ailuropoda melanoleuca) into the captive population.

Recent success in breeding giant pandas in captivity has encouraged panda conservationists to believe that the ex situ population is ready to serve as a source for supporting the wild population. In this study, we used 11 microsatellite DNA markers to assess the amount and distribution of genetic variability present in the two largest captive populations (Chengdu Research Base of Giant Panda Breeding, Sichuan Province and the China Research and Conservation Center for the Giant Panda at Wolong, Sichuan Province). The data were compared with those samples from wild pandas living in two key giant panda nature reserves (Baoxing Nature Reserve and Wanglang Nature Reserve). The results show that the captive populations have retained lower levels of allelic diversity and heterozygosity compared to isolated wild populations. However, low inbreeding coefficients indicate that captive populations are under careful genetic management. Excessive heterozygosity suggests that the two captive populations have experienced a genetic bottleneck, presumably caused by founder effects. Moreover, evidence of increased genetic divergence demonstrates restricted breeding options within facilities. Based on these results, we conclude that the genetic diversity in the captive populations is not optimal. Introduction of genetic materials from wild pandas and improved exchange of genetic materials among institutions will be necessary for the captive pandas to be representative of the wild populations.

Microreview: capsule-associated genes of Cryptococcus neoformans.

Cryptococcosis, caused by Cryptococcus neoformans is a common systemic mycosis in man and animals, particularly immunocompromised patients. This pathogenic fungus produces a thick extracellular polysaccharide capsule. Four capsule-associated genes (CAP10, CAP59, CAP60, CAP64) were cloned and sequenced, and proved to be essential for capsule synthesis. However biochemical functions of CAP gene products have not been clarified yet. Recently, the relatedness of the polysaccharide capsule and four capsule-associated genes has partly been elucidated. Nucleotide sequence of four CAP gene fragments was analyzed for phylogenetic relationships, and they were in agreement with the conventional classification of varieties and serotypes within C. neoformans. Expression of four CAP genes and capsule size were examined using two media containing different amount of glucose, and the results indicated that CAP genes might play important roles in elaboration of extracellular polysaccharide capsule. Furthermore, analyses of CAP genes in various clinical samples would give the useful information to diagnose cryptococcosis in human and animals.

Serotypes and mating types of clinical isolates from feline cryptococcosis in Japan.

Most isolates of Cryptococcus neoformans (teleomorph: Filobasidiella neoformans) from human patients and from environmental materials in Japan have been identified as serotype A mating type a by the seroagglutination test and mating experiments. A PCR method using the mating type alpha allele-specific primer of the STE12 gene and the serotype- and mating type-specific primers of the STE20 gene for identification of C. neoformans has been developed. Using the PCR method, conserved strains and clinical isolates from feline cryptococcosis were examined for serotype and the mating type. The results showed that all clinical isolates examined were identified as serotype A, MATalpha, indicating that feline cryptococcsis cases in Japan are caused by C. neoformans serotype A, MATalpha, as is the case in humans.

An automated experimental milker for rat.

The development of sucking pressure was investigated with an artificial nipple in 16 male and female rat pups on postnatal days 4, 7, 10, 14, and 18. The rat pups refused to suck on the artificial nipple on postnatal day 14 or day 18 thus negative sucking pressure had to be calculated by regression analysis. As a result, the mean maximum intra-oral negative sucking pressure on day 18 was calculated to be -160.1 mmHg in the male and -103.4 mmHg in the female. Based on these results, the maximum level of negative pressure of the automated experimental rat milker was set at -160 mmHg. The automated experimental milker for rat is able to collect milk from lactating mothers by alternating negative and atmospheric pressures through two solenoid valves and a vacuum pump attached to a microcomputer. Mother rats were milked with the automated experimental milker on postpartum days 4, 7, 10, 14 and 18 of a single lactation period. The maximum mean milk yield with this machine was 3.18 +/- 1.37 g, obtained on day 14 of the lactation period. This quantity is considerably lower than previously reported values obtained by measuring differences in body weight of the offspring and mother rat before and after suckling. It is necessary to further optimize this system, but the milk yield in the present study is adequate for chemical analysis.