Effect of volume of non-surgical embryo transfer medium on ability of porcine embryos to survive to term

Non-surgical embryo transfer is a promising method for improving efficiency in the pork industry and also for biotechnology applications, such as in vitro embryo production, transgenesis and cloning. Several groups have reported successful piglet production using an artificial insemination catheter or flexible catheter designed for this procedure; however, the efficiency of the technique is still low. The critical points that need to be addressed in order to improve this procedure are (1) the embryo deposition site and (2) volume of transfer medium associated with the embryos; however, the latter has not yet been examined systematically. In the present study, we evaluated the effect of the volume of non-surgical embryo transfer medium on the ability of porcine embryos to survive to term by using a recently produced flexible catheter. The catheter consists of a guide and an injector. Blastocysts 200-230 mum in diameter were collected from donor gilts and transferred to recipient gilts. The time required for the completion of embryo transfer using this catheter was 14.6 +/- 3.9 min. The tip of the injector was determined by laparotomy to be located in a uterine horn 20-30 cm anterior from the branching point of the uterus body. We transferred 17.0-17.3 embryos with different volumes of medium (1.6, 3.2 and 10 ml) into each of 5, 4 and 4 recipients, respectively, and pregnancy was confirmed in 4, 3 and 1 of these recipients, respectively. Three recipients in the 1.6 ml group farrowed a total of 19 piglets (4, 5 and 10 piglets, respectively). These results suggest that successful non-surgical embryo transfer is affected by the volume of transfer medium.     

Ontogeny of digestive and immune system organs of larval and juvenile kelp grouper Epinephelus bruneus reared in the laboratory

ABSTRACT:   A histological examination was made of the ontogenetic development of the digestive and immune systems of the larval and juvenile kelp grouper Epinephelus bruneus reared in the laboratory. The liver, gall bladder, pancreas and the demarcating region between the intestines and rectum were formed within 3 days post-hatch (dph). During the preflexion phase (within 16 dph), revolution of the intestine concluded, and pharyngeal teeth and the mucous cells of the esophagus were differentiated. In the transitional period to the juvenile stage (25 dph), the blind sac of the stomach, gastric glands and pyloric caeca began to form. From the viewpoint of the differentiation phase of the adult-type digestive system, the kelp grouper is similar to Heterosomata, hitherto reported. The primordial thymus, kidney and spleen were present at 12, 1 and 6 dph, and the small lymphocytes in these lymphoid organs appeared at 21, 30 and 33 dph, respectively. The developmental sequence of the lymphoid organs and the appearance ages of the lymphoid organs and small lymphocytes in the lymphoid organs in the kelp grouper are similar to those of other marine fish previously reported, except for the Pacific bluefin tuna Thunnus orientalis .     

Isolation and characterization of sulfur oxidizing bacteria from cattle manure compost

Hydrogen sulfide causes offensive odors. We attempted to isolate sulfur oxidizing bacteria (SOB) from cattle manure compost. A most probable number assay could not detect SOB by using a Cha medium which had been applied in order to isolate the SOB from active sewage. Cultivation using a Cha plate medium revealed 5.75 × 10⁷ colony forming unit/g of bacteria. A single strain of SOB was isolated from a colony formed on the plates and identified as Halothiobacillus neapolitanus. This is the first report that H. neapolitanus has been found in cattle manure compost.     

Identification of dimethylarsinic acid as the major arsenic compound in fish sauce by liquid chromatography/electrospray ionization-mass spectrometry

ABSTRACT:   Arsenobetaine, the major arsenic compound in marine animals, is substantially non-toxic. It is, however, possible that arsenobetaine undergoes bacterial transformation during manufacturing of fermented fishery products. In the present study, therefore, three types of fish sauce were examined for arsenic concentrations and species compared with those in the raw materials (sardine, Japanese sandfish and Japanese common squid). Arsenic concentrations of the three types of fish sauce were almost equivalent to those in their raw materials, suggesting no accumulation of arsenic during fermentation. Arsenic speciation was performed by a combination of cation-exchange liquid chromatography (LC) and electrospray ionization (ESI)-single quadrupole mass spectrometry (MS). Optimal conditions of LC/ESI-MS were established to analyze seven arsenic compounds (arsenate, monomethylarsonic acid, dimethylarsinic acid, arsenobetaine, trimethylarsine oxide, arsenocholine and tetramethylarsonium ion) found in biological samples. When analyzed by LC/ESI-MS, the major arsenic compound in the raw materials was arsenobetaine as expected, while not arsenobetaine but dimethylarsinic acid was identified as the major arsenic compound in the three types of fish sauce. These results suggest that arsenobetaine in the raw materials is converted to dimethylarsinic acid but not to arsenate with high toxicity by bacterial actions during manufacturing of fish sauce.     

Molecular Monitoring of hrpB-disrupted Mutant of Ralstonia solanacearum in Tomato Plants

A nonpathogenic mutant of Ralstonia solanacearum was produced by the insertion of transposon Tn4431. The mutagenized gene was then cloned from a genomic DNA library by the gene tagging method, using the labeled lux operon located on Tn4431 of pUCD623 as a hybridization probe. From nucleotide sequence analysis of the transposon-inserted genomic clone, the hrpB gene was shown to be disrupted by the inserted transposon. Tomato plants were inoculated with the hrpB-disrupted mutant bacteria, for which multiplication and translocation were then monitored using the colony hybridization method. In addition, the original pathogenic bacteria in which the lux operon had been functionally ligated with the genomic promoter were also used for inoculation and traced by their bioluminescence. Multiplication of the hrpB-disrupted mutant was suppressed initially in the invaded root tissues and then in upper hypocotyl after translocation, suggesting that the pathogenic strain of R. solanacearum overcomes at least two steps of host responses expressed in root and hypocotyl tissues. Thus, our approach for molecular monitoring of the bacteria enabled us to precisely analyze the infection behavior of the pathogenic bacteria in planta. Received 16 April 1999/ Accepted in revised form 10 August 1999     

A case of spontaneous Zymbal’s gland carcinoma with lung metastasis in an aged Fischer 344 rat

Zymbal’s gland neoplasms are induced in rats through the administration of various carcinogens, but spontaneous neoplasia is rare. This report describes a spontaneous Zymbal’s gland carcinoma with lung metastasis found in an aged male Fischer 344 rat. Macroscopically, the dome-like tumor nodule, approximately 30 mm in diameter with ulceration, was located near the ear canal of the rat. No healthy tissue or structure of Zymbal’s gland was identified on the corresponding side, while the normal salivary glands and a lacrimal gland were observed. Histologically, a large part of the tumor mass was occupied by poorly differentiated neoplastic cells, the shapes of which were oval to polygonal or fusiform. Additionally, clusters of sebaceous-like foamy cells and squamous metaplasia with prominent keratinization were observed. Tumor cells were found to metastasize to the lung; these cells displayed histological similarities, including a sebaceous gland-like pattern, to those in the primary site. The tumor cells were immunohistochemically positive for cytokeratin AE1/AE3 or vimentin but negative for CD68, S100, α-smooth muscle actin, von Willebrand factor, and desmin. Our results indicate that the tumor was a poorly differentiated Zymbal’s gland carcinoma with lung metastasis.     

Methods of evaluating the spermatogenic ability of male raccoons (Procyon lotor)

Feral raccoons (Procyon lotor) have been growing in number in Japan, and they are becoming a problematic invasive species. Consequently, they are commonly captured and killed in pest control programs. For effective population control of feral raccoons, it is necessary to understand their reproductive physiology and ecology. Although the reproductive traits of female raccoons are well known, those of the males are not well understood because specialized knowledge and facilities are required to study them. In this study, we first used a simple evaluation method to assess spermatogenesis and presence of spermatozoa in the tail of the epididymis of feral male raccoons by histologically examining the testis and epididymis. We then evaluated the possibility of using 7 variables—body weight, body length, body mass index, testicular weight, epididymal weight, testicular size and gonadosomatic index (GSI)—to estimate spermatogenesis and presence of spermatozoa in the tail of the epididymis. GSI and body weight were chosen as criteria for spermatogenesis, and GSI was chosen as the criterion for presence of spermatozoa in the tail of the epididymis. Because GSI is calculated from body weight and testicular weight, this model should be able to be used to estimate the reproductive state of male raccoons regardless of season and age when just these two parameters are known. In this study, GSI was demonstrated to be an index of reproductive state in male raccoons. To our knowledge, this is the first report of such a use for GSI in a member of the Carnivora.