Methods to detect infectious agents in laboratory animals have traditionally been serological and culture based. Molecular methods to detect infectious agents in laboratory animals are being used more routinely. Confusion as to when and how to use molecular methods abounds. In this review, we present a guide to the weaknesses and strengths of using traditional and molecular methods for the detection of infectious agents in laboratory animals.

Purpose: We sought to isolate, clone, and determine the nucleic acid sequence of the guinea pig adenovirus (GPAdV) hexon gene. From this, the amino acid sequence of the cloned portion was deduced and compared with a set of mastadenovirus hexons. Methods: The DNA isolated from a histologic section of infected guinea pig lung was subjected to high-fidelity amplification, using degenerate primers complementary to a conserved nucleic acid sequence near the 3' end of the hexon gene of mastadenoviruses and a 5' primer from GenBank accession No. X95630 (GPAdV hexon gene partial sequence). The amplified product was cloned, the nucleic acid sequence was determined, and the amino acid sequence was deduced and compared with the hexon amino acid sequences of 25 mastadenoviruses. Results: The cloned fragment comprised 1,603 base pairs (bp) [∼50%]) of the hexon. Of the initial 278 nucleic acids of the clone, 276 were identical with GenBank accession No. X95630, and the deduced amino acid sequences of both were identical. The deduced GPAdV hexon amino acid sequence from the clone aligned with structural regions NT, V1, DE1, and FG1 described for human adenovirus types 2 and 5. The GPAdV hexon had < 50% similarity in amino acid sequence, compared with hexons of 25 other mastadenoviruses. Analysis of regional peptide similarities revealed the GPAdV hexon to be more similar to animal mastadenoviruses and human subgroups A, C and F than to other human subgroups. Conclusions: The cloned portion of the GPAdV hexon contained a sequence nearly identical to that of GenBank accession No. X95630. Compared with the truncated amino acid sequences of human adenovirus types 2 and 5, the deduced GPAdV hexon amino acid sequence was similar in areas structurally conserved, but different in areas associated with type-specific antigenicity.

Regional variations in intraepithelial lymphocytes (IELs) in the small intestine were examined in BALB/c +/+, nu/+, and nu/ nu mice. The small intestine was obtained from 11- to 12-week-old mice and divided equally into three (proximal, middle, and distal) parts. The IELs were isolated from each part of the intestine, and the total numbers of IELs in nu/+ and nu/ nu mice were about a fifth of those in +/+ mice. Regional variations in the distribution of the IEL , but not the T-cell subset were found by use of flow cytometry in +/+ and nu/+ mice. On the other hand, such differences were not found in nu/ nu mice, suggesting that thymus-independent development of T cells is not different among regions. Different local expansion of thymus-dependent T cells may cause the regional variations seen in the distribution of T cell IELs in +/+ and nu/+ mice.

Background and Purpose: Studying the effects of toxic and infective compounds on the respiratory system requires a reliable method for delivering inoculum into the distal region of the lung. Although transoral intratracheal inoculation methods have been well documented for adult rats, to the authors' knowledge, a reliable method has not been validated for neonatal rats. The purpose of the study reported here was to develop a simple method for transoral inoculation in rat neonates. Methods: Seven-day-old Fischer 344 rats were anesthetized with halothane, and a spinal needle was inserted in the tracheal lumen, by use of illumination and a modified otoscope. Meconium was injected into the lungs as a marker, and the neonates were kept under close observation. After euthanasia at 24 h, lungs were removed and fixed in formalin, and the microscopic distribution of the inoculum was assessed in the left, right cranial, middle, median, and caudal lung lobes. Results: Microscopic examination of lungs indicated that intratracheal inoculation was achieved in 100% of neonatal lungs and the inoculum was consistently distributed in the alveoli of all pulmonary lobes. Important complications or mortality were not observed in the neonates. Conclusions: Intratracheal inoculation of neonatal rats is possible by use of a modified otoscope for transoral illumination. This technique is simple and reproducible and ensures, without complications, widespread distribution of inoculum in the lungs of neonatal rats.

Purpose: To diagnose lung and liver tumors experimentally induced in mice in three-dimensional magnetic resonance (MR) images constructed by superimposing transversal multislice MR images of thoracic and abdominal regions taken under a high magnetic field of 7.05 tesla (T). Methods: Lung and liver tumors were induced by administration of urethane to A/J mice and implantation of transplantable colon-26 cells into BALB/c mice, respectively. Two-dimensional (2-D) multislice MR images from the thoracic to abdominal regions were taken under the proton density-weighted conditions. Each organ in the 2-D MR images was pseudocolored, and a three-dimensional (3-D) image was constructed by superimposing them on a UNIX computer, using volume-rendering software. Results: In the normal mouse, each organ in the thoracic and abdominal regions was three-dimensionally imaged and was clearly distinguished from the others. In mice with tumors in the lung or liver, the pathologic changes in the tissue could be visualized in 3-D images. Conclusions: The MR images three-dimensionally constructed by use of a method combining MR imaging under a high magnetic field of 7.05 T and a computer technique using volume-rendering software was useful for diagnosis of lung and liver tumors experimentally induced in mice.

Purpose: Mouse polyoma virus and K virus are murine polyomaviruses frequently used in carcinogenicity and cellular biology studies in mice. These viruses can cause persistent infections, which increase the likelihood of transmission through transplantation of cells from infected mice. To identify polyomavirus-infected biological samples, several diagnostic polymerase chain reaction (PCR) assays were developed. Methods: Polyomavirus-family and virus-specific PCR assays were designed and optimized for specificity and sensitivity. The generic (polyomavirus-family) PCR assay and mouse polyoma virus-specific assays were compared with the mouse bioassay for diagnosis of infected cellular samples. Results: Specificity of the PCR assays was confirmed by testing a battery of other murine viruses. The mouse polyoma virus PCR test was the most sensitive assay, detecting as few as 2,000 copies of homologous virus. The K virus PCR assay was about eightfold less sensitive, and the generic PCR test was the least sensitive. Mouse polyoma virus and generic PCR assays amplified mouse polyoma virus in the inoculum and tissues from experimentally infected mice, and performed better than did the mouse bioassay. Conclusions: Results of this study confirm that PCR is a specific and sensitive method for detection of murine polyomaviruses in biological samples.

The objective of the study reported here was to induce obesity in the female Göttingen minipig to establish a model of the human metabolic syndrome. Nine- to ten-month-old female Göttingen minipigs received a high-fat high-energy (HFE) diet or a low-fat, low-energy (LFE) diet. The energy contents derived from fat were 55 and 13 %, respectively. After 5 weeks, animals were subjected to dual energy x-ray absorptiometry (DEXA) scanning, intravenous glucose tolerance testing (IVGTT), and 6-h growth hormone profile recording. After treatment, mean body weight of pigs of the LFE group was 21.0 ± 0.4 kg, and was 26.8 ± 0.2 kg in pigs of the HFE group (P < 0.0001). The DEXA scanning indicated that the fat content of the LFE group was 10.0 ± 1.2 % versus 15.2 ± 0.7 % in the HFE group (P < 0.003). Triglycerides concentration was significantly (P < 0.05) increased in pigs of the HFE group (0.24 ± 0.03 m M), compared with that in pigs of the LFE group (0.13 ± 0.04 m M). Preprandial plasma glucose and insulin concentrations were not affected, but insulin area under the curve during IVGTT was significantly high in the obese animals. Growth hormone (GH) secretion was low in both groups of pigs. The obese minipig shares some of the metabolic impairments seen in obese humans, and may thus serve as a model of the metabolic syndrome.

We studied the allelic and genotypic distribution of the major histocompatibility class-II locus DQA1 observed in a random sample of Indian rhesus macaques (Macaca mulatta) from a major breeding facility in the United States. The DNA was isolated from whole blood samples collected between 1991 and 1994 from 65 Indian rhesus monkeys. Polymerase chain reaction-restriction fragment length polymorphism analysis (PCR-RFLP), which involves use of specific amplification of DQA1 exon 2 and subsequent restriction digestion of the 242-base pair fragment, was used to genotype the animals for the 20 known macaque (Mamu)-DQA1 alleles. Frequencies for four alleles (DQA1*240x, *2502, *2503 and *0102) differed significantly from those reported in a smaller sample of rhesus macaques from the German Primate Center. The modest genetic survey of Mamu-DQA1 genotypes presented here will be particularly useful in designing epidemiologic studies that investigate associations between immunogenetic background and disease susceptibility in macaque models of human disease.

Purpose: To develop a pig model that would enable repeated biopsy specimen collection and endoscopic monitoring of the gut. This would increase precision of the experiment and reduce the number of experimental animals required. Methods: Six 10-week-old Yorkshire pigs underwent surgery, and a cannula was inserted in the cecum. Two pigs served as non-operated controls. The health status of the animals was monitored by clinical, hematologic, and biochemical examinations and by studies of gut motility and microbial flora. The experimental period lasted for eight weeks and approximately 45 biopsy specimens were obtained from each animal. Results: Repeated endoscopy was performed and biopsy specimens were taken. Adverse effects on the animal's health were not apparent, and differences were not evident in transit time of digesta or in diversity of the gut microbial flora. After surgery there was a transient increase in the concentrations of haptoglobin, serum amyloid A, and plasma cortisol, and in body temperature and white blood cell count. Conclusions: It is possible to use an intestinal cannula in the cecum both for endoscopy and biopsy specimen collection. The procedures did not influence health status of the pigs, nor alter gut function. The method will be useful in experimental infection studies as well as in other physiologic investigations.

Japanese medaka, (Oryzias latipes), small, freshwater, tropical cyprinodonts, are principally used for toxicologic and carcinogenicity assays, but are finding more applications in developmental genetic and biological research. An increase in mortality began in brood stock of adult medaka that had been shipped and housed separately by sex. Initially, mortality averaged one fish daily and began in females two weeks after they were received. Cohabitation began eight weeks after arrival. After four to six weeks of cohabitation in different spawning aquaria, mortality was observed in males. Clinical signs of disease included loss of scale luster and color, with subsequent blanching of dorsal flank musculature, small raised nodules on various external surfaces, emaciation, fraying of fin tips, and equilibrium disturbances. Histologic examination of affected adults revealed multi-organ granulomatous inflammation with intracellular acid-fast bacilli. Specimens from 46 juvenile medaka that were spawned from affected adults, were submitted for culture and histologic evaluation. Of 18 fish, two had lesions similar to those of adults. The organism isolated from the remaining fish was identified as Mycobacterium fortuitum. Due to atypical rapid progression of disease, spread of M. fortuitum to progeny, and poor prognosis, the entire colony was euthanized.

A spontaneous focal polar anterior subcapsular lenticular opacity characterized by focal epithelial proliferation was found in Charles River Sprague-Dawley rats from various breeding facilities around the world (F rance, Japan, and the United States). The incidence of this change slightly increased with age up to a maximal incidence of 9.8% in 28- to 35-week-old male rats (French source). Over that period, there was little change in the size of the opacity; however some rats that were examined over longer periods (more than 2 years of age) developed secondary anterior cortical changes, and rarely, histologic findings of pigmentation and/or mineralization. The lenticular change was present throughout the life of the animals and had no sex predilection; mode of inheritance was not investigated. Due to its small size, this lens opacity is more easily identified by use of slit lamp biomicroscopy than by use of indirect ophthalmoscopy, and serial sections of the eye aid in locating it for histologic evaluation.