Selected Aspects of Advanced Porcine Reproductive Technology

In vitro fertilization (IVF) of in vitro matured (IVM) oocytes in pigs has become the most popular method of studying gametogenesis and embryogenesis in this species. Furthermore, because of recent advances in in vitro culture (IVC) of IVM–IVF embryos, in vitro production (IVP) of embryos now enables us to generate viable embryos as successfully as for in vivo -derived embryos and with less cost and in less time. These technologies contribute not only to developments in reproductive physiology and agriculture but also to the conservation of porcine genetic resources and the production of cloned or genetically modified pigs. However, in IVP, there still remains the problem of abnormal ploidy, which is caused by performing procedures under non-physiological conditions. In recent years, unique technologies such as intracytoplasmic sperm injection (ICSI) or xenografting of gonadal tissue into immunodeficient experimental animals have been developed to help conserve gamete resources. These technologies combined with IVP are expected to be useful for the conservation of gametes from important genetic resources. Here, we discuss the developmental ability and normality of porcine IVP embryos and also the utilization of ICSI and xenografting in advancing biotechnology in pigs.     

Nucleotide sequence homology to bovine viral diarrhea virus 2 (BVDV 2) in the 5′ untranslated region of BVDVs from cattle with mucosal disease or persistent infection in Japan

Cytopathogenic and non-cytopathogenic bovine viral diarrhea viruses (BVDVs) were isolated from cattle with mucosal disease or persistent infection in Japan. These isolates were compared for antigenic properties by cross-neutralization tests with Japanese reference strains of BVDV belonging to classical type 1. Significantly low cross-reactivity to reference strains was noted, indicating the viruses to possibly represent a new serotype in Japan. Thus, to determine the genotype of the isolates, nucleotide sequences of the 5′ untranslated region were determined and compared with those of previously reported BVDV 1 and 2. The isolates were clearly shown to belong to BVDV 2, not to BVDV 1.     

Ovarian follicular dynamics and hormones throughout the estrous cycle in Thai native (Bos indicus) heifers

Relatively few studies have been reported regarding the reproductive physiology of female Thai native cattle. Therefore, the objective of the present study was to evaluate the follicular dynamics and concentrations of follicle stimulating hormone (FSH), estradiol (E2) and progesterone (P4) during the estrous cycle in Thai native heifers (TNH) and to compare obtained results with those of European and Indian cattle breeds previously reported. For the detection of estrus, ovaries of all 20 heifers were examined twice daily (12 h intervals) by ultrasonography for three consecutive estrous cycles. From data of 60 estrous cycles (n = 60 estrous cycles from 20 heifers), it was found that 14 (70%) and 6 heifers (30%) had two (42 estrous cycles collected from 14 heifers) and three follicular waves (18 estrous cycles collected from 6 heifers), respectively. The days when estrus was detected, interovulatory intervals, life‐spans of corpus lutea (CL), and days for growing and regression of CLs were shorter in the two follicular waves than those in the three follicular waves (P < 0.05). In both two and thre follicular waves, larger maximum diameters and higher growth rates of the dominant follicle (DF) in an ovulatory wave were observed than those of the preceding waves without ovulation (P < 0.05). There was a progressive increase in follicular size and FSH and E2 production during follicular growth in each follicular wave. In addition, the FSH and E2 peak concentrations during the ovulatory wave were higher than those of the anovulation waves (P < 0.05). Moreover, although the ovarian follicular dynamic patterns in Thai native heifers were similar to those previously reported for European and Indian cattle breeds, the diameter of the largest preovulatory follicle (OF), subordinate follicles (SF) and CLs were smaller than those in European and Indian cattle breeds. In conclusion, when compared with European and some breeds of Indian cattle, the length of interovulatory intervals was shorter, and the sizes of dominant SF and CLs were smaller in Thai native heifers.     

Incentive for shifts in water management systems by shrimp culturists in southern Thailand

ABSTRACT:   Water management systems and other business situations of shrimp culturists were surveyed in two districts in southern Thailand. There were three types of water management systems in southern Thailand, namely closed, semiclosed and open systems, categorized by the frequency of exchange of pond water. Shrimp culturists in those districts tended to shift their system from the open system to the closed system of their own accord, and the average net income ratio decreased due to the change of water management system in a district from more than 2 to less than 1. The reason for this shift, in spite of the decrease in the profit rate, was prevention of infectious disease caused by water exchange. In conclusion, it was proven that independent shrimp culturists, such as medium and small scale shrimp pond owners in southern Thailand, would accept short-term decreases in profit rate in order to stabilize production.     

Determination of polysorbates in foods by colorimetry with confirmation by infrared spectrophotometry, thin-layer chromatography, and gas chromatography

A method is presented for the detection of polysorbates (PSs) in 8 kinds of processed foods by colorimetric and thin-layer chromatographic (TLC) techniques. The PSs are extracted from processed foods with a mixture of methylene chloride and ethanol by using an Extrelut column. The extract is further purified by using a silica gel column. The PS extract is complexed with cobalt-thiocyanate (Cothiocyanate) reagent and is determined spectrophotometrically at 620 nm. The recoveries and coefficients of variation for 8 kinds of processed foods fortified with 0.1% PS 80 were 67.9-94.6% and 4.0-11.3%, respectively. The detection limit of TLC corresponded to 50 mg PS 80/kg. PS identity was confirmed by infrared spectrophotometry of PS extract, and gas chromatography of fatty acids and thin layer chromatography of POE-sorbitan residues after saponification.     

Variation in understory structure and plant species diversity influenced by silvicultural treatments among 21- to 26-year-old Picea glehnii plantations

We investigated effects of silvicultural treatments (planting and subsequent treatments) on understory structure and plant species diversity in managed Picea glehnii plantations (21–26 years old) in northern Japan. We evaluated the importance of each treatment (machinery site preparation, planting, weeding, and thinning) in 19 plantations, with considerable variation among treatments overall. The understory had 98 vascular plant species; the most dominant species was a dwarf bamboo Sasa senanensis, followed by tree species Abies sachalinensis and Betula ermanii. Multiple regression analyses showed that thinning negatively influenced plant species diversity. Planting density showed a strong positive correlation with density of seedlings (height <20 cm), but few independent variables were correlated with density of saplings (height ≥20 cm and diameter at breast height <1 cm). The negative effect of thinning and the positive effect of planting density seem to be related to the existence or reinitiation of dense cover of Sasa senanensis. We present possible mechanisms of response to treatments to generalize the results. We then suggest ways to improve current treatments to meet the goals of wood production and biodiversity conservation in the region.     

Development of a quantification assay for the cysts of the toxic dinoflagellate <Emphasis Type="Italic">Alexandrium tamarense</Emphasis> using real-time polymerase chain reaction

ABSTRACT:   The cysts of toxic dinoflagellate Alexandrium tamarense are the seed population for the bloom responsible for paralytic shellfish poisoning (PSP). However, it is impossible to identify the Alexandrium spp. cyst on the basis of morphological features. In this study, we prepared A. tamarense cysts by sexual conjugation in laboratory conditions and developed an efficient DNA extraction method for polymerase chain reaction (PCR) assay. Using the A. tamarense cysts, we established the identification and quantification method showing the species specificity and the high sensistivity for A. tamarense cysts using real-time PCR. This assay was also able to detect and quantify the A. tamarense cysts accurately when mixed with excess cysts of A. catenella (Whedon and Kofoid) Balech prepared by conjugation experiment.     

Development of a genetically modified nontoxigenic Pasteurella multocida toxin as a candidate for use in vaccines against progressive atrophic rhinitis in pigs

OBJECTIVE: To construct a genetically modified nontoxigenic Pasteurella multocida toxin (PMT) and examine its immunoprotective activity against challenge exposure with wild-type PMT in pigs. ANIMALS: 5 healthy pigs. PROCEDURE: A nontoxigenic PMT was created by replacing the serine at position 1164 with alanine (S1164A) and the cysteine at position 1165 with serine (C1165S). Toxic activity was determined by use of the guinea pig skin test and mouse lethality test. Three pigs were vaccinated twice with the modified PMT, and the remaining 2 pigs served as nonvaccinated control animals. Vaccinated and control pigs were challenge exposed with wild-type PMT. Pigs were euthanatized and necropsied on day 14 after challenge exposure. Turbinate atrophy was examined macroscopically and assigned a score. Serum anti-PMT antibodies were determined by use of an ELISA. RESULTS: The genetically modified PMT was characterized by a total lack of toxic activity. Pigs vaccinated with the modified PMT became seropositive; in contrast, control pigs remained seronegative. Necropsy revealed that the 2 control pigs had moderate and severe turbinate atrophy, respectively, whereas the 3 vaccinated pigs did not have any lesions in the turbinates or abnormalities in other organs. CONCLUSIONS AND CLINICAL RELEVANCE: Modification by use of S1164A and C1165S leads to a complete loss of toxic effects of PMT without impairment of the ability to induce protective immunity in pigs. Analysis of these results suggests that genetically modified PMT may represent a good candidate for use in developing a vaccine against progressive atrophic rhinitis in pigs.