Mitochondrial DNA analysis of Nepalese domestic dwarf cattle Lulu*

Dwarf Lulu cattle, the only Bos Taurus type of cattle in Nepal, are raised under severe environments in the mountainous zone of that country. In the present study, the body measurement traits, cytogenetic and molecular genetic characteristics of the Lulu cattle are investigated. Blood samples were collected from 31 animals in four villages (altitudes 2590–3550 m) in the southern part of Mustang. The Lulu cattle had a normal karyotype with 2n = 60, XY or XX. Only one male examined had a large submetacentric X‐chromosome and a small submetacentric taurine type Y‐chromosome. The mitochodrial DNA (mtDNA) genotypes were analyzed by PCR mediated restriction fragment length polymorphisms, displacement (D)‐loop region PCR mediated single strand conformation polymorphisms, and D‐loop region sequences. Many base substitutions were found in the D‐loop region, suggesting that the Lulu cattle originated from at least 10 maternal lines. Three types of mtDNA from these cattle were found, the Bos taurus type (n = 23), the Bos indicus type (n = 6), and the Bos grunniens type (n = 2). In the village at the lowest altitude, four of the five cows were of the Bos indicus type. These results indicated that mtDNA types of the Lulu cattle mostly belong to Bos taurus, but have been hybridized with Bos indicus cattle in lower‐elevation regions in their maternal lineage.     

Pathological analysis of batch safety testing of veterinary vaccines using small laboratory animals

Batch safety tests (BSTs) of veterinary vaccines are conducted using small laboratory animals to assure the safety of vaccines according to several criteria, including clinical signs and change in body weight. Although the latter is used as an evaluation index in BSTs, there have been no reports on the internal changes that affect body weight during the test period. Therefore, we analyzed BST via pathological examination of the tested animals. Here, BSTs were performed for 176 batches using mice and 126 batches using of guinea pigs. Most of the gross findings could be classified into four lesion types (nodules, adhesions, ascites, no apparent signs), with only one vaccine inducing lesions that could not be classified into any of these four types. Histopathological examination revealed that the reactions caused by BST were pyogenic and/or granulomatous inflammation. Nodular or adhesive lesions comprised more severe pyogenic granulomatous inflammation than ascites or cases with no apparent gross lesions. These nodular or adhesive lesions were more frequently induced by vaccines that contained an adjuvant than by vaccines that did not contain an adjuvant. The cases with “exceptional” gross findings histologically presented severe necrosis of the hematopoietic system. Additional testing showed that these “exceptional” lesions were induced when a specific type of light liquid paraffin was injected along with other vaccine additives. Our results show that body weight loss and/or lesions during BST were induced by proinflammatory properties of the tested vaccines and that BST is a sensitive method for detecting unexpected effects of vaccine components.     

Growth hormone suppresses the expression of IGFBP-5, and promotes the IGF-I-induced phosphorylation of Akt in bovine mammary epithelial cells

Growth hormone (GH) plays a specific role to inhibit apoptosis in the bovine mammary gland through the insulin-like growth factor (IGF)-I system, however, the mechanism of GH action is poorly understood. In this study, we show that GH dramatically inhibits the expression of IGFBP-5, and GH along with IGF-I enhanced the phosphorylation of Akt through the reduction of IGF binding protein (IGFBP)-5. To determine how GH affects Akt through IGF-I in bovine mammary epithelial cells (BMECs), we examined the phosphorylation of Akt in GH treated BMECs and found that IGF-I induced phosphorylation of Akt was significantly enhanced by the treatment with GH. We demonstrated that GH reduces mRNA and protein expression of IGFBP-5 in BMECs, but it does not affect the expression of IGFBP-3. To determine that the enhanced effect of the Akt phosphorylation by the treatment of GH is due to the inhibition of the expression of IGFBP-5, we examined the effect of IGFBP-3 and -5 on the phosphorylation of Akt through IGF-I in the GH-treated BMECs. The phosphorylation of Akt was inhibited in a dose-dependent manner when IGFBP-5 was added at varying concentrations and was also inhibited in the presence of IGFBP-3. The results of this study suggest that GH plays an important role on mammary gland involution in bovine mammary epithelial cells.     

Synthesis,insecticidal activity and pyrethroidal mode of action of new tin ether derivatives

A novel range of trimethylstanniomethyl ethers of well known pyrethroid alcohols were synthesised, and their insecticidal activities and modes of action as insecticides were investigated. Among them, ethers from three types of alcohol (3-phenoxybenzyl, 4-fluoro-3-phenoxybenzyl and 6-phenoxy-2-pyridylmethyl) showed remarkable insecticidal activities against rice stem borers, houseflies and German cockroaches. According to electrophysiological studies on the abdominal nerve cords of German cockroaches, trimethylstanniomethyl 6-phenoxy-2-pyridylmethyl ether induced a rapid decline in spontaneous firing similar to that from tetramethrin. However, insecticidal trimethyltin chloride caused an entirely different response. These observations suggest that the present tin ether derivatives resemble pyrethroids, rather than the insecticidal tin compounds known so far.     

Characterization of a plasmid-encoded adhesin of an avian pathogenic Escherichia coli (APEC) strain isolated from a case of swollen head syndrome (SHS)

An avian pathogenic Escherichia coli (APEC) strain designated SHS4, isolated from a chicken with clinical signs of swollen head syndrome (SHS), adhered to but did not invade Hep-2 and tracheal epithelial cells. The PCR amplified fimA, csgA and tsh gene sequences. It produced Ia, Ib, E1, E3, K, and B colicins, but not colicin V and aerobactin. It harboured two plasmids of 60 and 98MDa and was resistant to streptomycin and tetracycline. Conjugation with a nalidixic acid (Na) resistant K-12 recipient strain (MS101) showed that the 98MDa plasmid did not transfer, whereas transfer of the 60MDa plasmid resulted in concomitant transfer of adhesion to Hep-2 and tracheal epithelial cells, production of the colicins Ia, E1, E3, and K, and the tsh-related DNA sequence. Transposon (TnphoA) mutagenesis of strain TR4 gave rise to strain Mut23, which lost its adhesive capacities, but was still able to express the same colicins as did strain TR4. PCR was able to amplify the tsh-related DNA sequence in this strain and a molecular probe based on transposon TnphoA indicated that the transposon was inserted in the 60MDa plasmid. Based on these results, we suggest that the 60MDa plasmid have adhesion genes, which may be responsible for the initial colonization of the upper respiratory tract of chickens.     

Frequency of enteropathogen detection in suckling and weaned pigs with diarrhea in Japan.

Fecal samples from suckling (n=153) and weaned (n=116) piglets with diarrhea in Japan were examined for shedding of viral, bacterial, and parasitic pathogens using culture, microscopic, and polymerase chain reaction methods. In suckling piglets, diarrhea was attributed to infection with a single etiologic agent in 60.8% of cases and with combinations of agents in 22.2%. In weaned piglets, diarrhea was attributed to a single etiologic agent in 43.1% and to combinations of agents in 47.4% of cases. Rotavirus was the most prevalent agent in suckling (67.3%) and weaned (65.5%) piglets. The detection of other pathogens was associated with age of the animals examined. Coccidia were predominantly isolated from suckling piglets, whereas Escherichia coli was found predominantly in weaned piglets. Although a relationship was not observed between detection rate of rotavirus and age of piglets, a single group of rotavirus was detected in 87.5% of suckling piglets whereas multiple groups were detected in 51.6% of weaned piglets. The results of this study confirm that diarrhea in piglets can, to a variable degree, be causally associated with multiple agents. Additionally, these results suggest reasons why this syndrome can be difficult to control.     

Performance of a foot-and-mouth disease virus reverse transcription-polymerase chain reaction with amplification controls between three real-time instruments.

The foot-and-mouth disease virus (FMDV) is a member of the picornavirus family, possessing an 8-kb single-stranded RNA genome of positive polarity. It is highly contagious among several livestock species and can lead to severe economic consequences, as evidenced by the UK outbreak in 2001. The usage of real-time polymerase chain reaction has facilitated rapid detection of FMDV. Several real-time PCR instruments are available with various capabilities, such as portability and high sample volume analysis. Primers and a dual-labeled TaqMan probe were optimized to detect a single, highly conserved 88-bp segment of the FMDV 3D (RNA polymerase) gene. To increase the confidence of the RT-PCR result, a positive amplification control was synthesized to detect potential false-positive results due to contamination if a wild-type virus is used as positive control. In addition, a preventative measure against false-negative results was developed in which endogenous beta actin mRNA is coamplified by RT-PCR. Assay performance was compared on the LightCycler1.2 (Roche), the SmartCyclerII (Cepheid), and the SDS 7900HT (ABI). These assays successfully identified the FMDV genome and beta actin mRNA from several sources of infected nasal and oral swabs, as well as probang samples.     

Durability evaluation of particleboards by accelerated aging tests

Summary Specimens of four commercially produced particleboards were exposed to various accelerated aging treatments: 1) 6-cycle ASTM D-1037 exposure, 2) 6-cycle WCAMA exposure, 3) BS 5669 test, 4) cyclic soak-dry test, and 5) cyclic boil-dry test. Thickness swelling (TS) and internal bond strength (IB) were measured after each step of each cycle. Phenol and phenol (face)/isocyanate (core)bonded boards are more durable than urea and melamine urea bonded boards. The effect of treatment 1) is most severe on TS and IB retention, followed in order by treatments 2), 5), 4) and 3). When the value of springback reached about 35%, IB became equal to zero.     

Fatigue of structural plywood under cyclic shear through thickness I: fatigue process and failure criterion based on strain energy

The fatigue behavior of plywood specimens under shear through thickness was examined on the basis of strain energy to obtain common empirical equations for the fatigue process and failure criterion under various loading conditions. Specimens were cut from commercial plywood panels of 9-mm thickness. Loading conditions were set as follows: a square waveform at a loading frequency of 0.5 Hz, a triangular waveform at 0.5 Hz, and a triangular waveform at 5.0 Hz. Peak stress applied was determined to be 0.5, 0.7, and 0.9 of static strength, that is, stress levels of 0.5, 0.7, and 0.9. The stress-strain relationships were measured throughout the fatigue test, and the strain energy was obtained at each loading cycle. Loading conditions apparently affected the relationship between stress level and fatigue life. On the other hand, the relationship between mean strain energy per cycle and fatigue life was found to be independent of loading conditions. Mean strain energy per cycle obtained as the fatigue limit was 5.85 kJ/m³ per cycle. Assuming that the accumulation of strain energy is a fatigue indicator, the fatigue process and failure criterion for the plywood specimens under the three loading conditions were commonly expressed by the relationship between cumulative strain energy and loading cycles.