Broad detection and quick differentiation of bovine viral diarrhea viruses 1 and 2 by a reverse transcription loop-mediated isothermal amplification test.

For broad detection of pestivirus A (bovine viral diarrhea virus 1: BVDV1) and pestivirus B (BVDV2) by a reverse transcription loop-mediated isothermal amplification (RT-LAMP) test, the P25 primer set was designed using nucleotide sequences of 5'-UTR region of 1454 BVDVs. The base coverage of each primer against diverse BVDVs were more than 99% in each base position. The one step LAMP test with the P25 primer set could detect both BVDV1 (TK) and BVDV2 (KZ), but did not amplify 5 other bovine viruses. Detection limit of the LAMP test was 103 copies of synthesized DNAs, and 10-3 and 10-4 dilutions of viral RNAs of TK and KZ strains, respectively, whereas that with current Aebischer's primer set was 10-2 dilution and negative of these RNAs, respectively. All of the 63 viral RNA samples of persistently infected (PI) cattle, consisting of the 1a (12), 1b (31), 1c (11), and 2a (9) subgenotypes, were broadly detected with the P25, while only 65% of them were positive with Aebischer's primer set. The validation study showed that the RT-LAMP test with the P25 had 100% sensitivity and 100% specificity against that with updated Vilcek's PCR primers. Also, by using the P26 primer set which contained 3 species-specific primers, all 63 RNA samples were clearly distinguished from BVDV1 or BVDV2 by the typing RT-LAMP test. These results indicate that the one step RT-LAMP test using P25 or P26 primer sets would be useful for broad detection and rapid differentiation of BVDV1 and BVDV2.

Day of prostaglandin F2α administration after natural ovulation affects the interval to ovulation, the type of ovulated follicle, and the failure to induce ovulation in cows.

The factors that affect the interval to ovulation, the type of ovulated dominant follicle (DF), and the cause of anovulation after prostaglandin (PG) treatment were investigated. Nine cows were assigned to six groups (54 cows in total) but the group size was later fixed at eight cows (48 in total). They received 25 mg tromethamine dinoprost as dinoprost on Day 6 (Group D6), Day 7 (Group D7), Day 8 (Group D8), Day 9 (Group D9), Day 10 (Group D10), or Day 11 (Group D11) after natural ovulation (Day 0). If the DF did not ovulate, then the cow was assigned to Group NO. In Group D6, the 1st DF ovulated in all cows 4 days after PG treatment, whereas in Groups D9, D10, and D11, the 2nd DF ovulated in all cows 4 to 7 days after PG treatment. In 10 cows, the DF did not ovulate, and late anovulation was significantly higher in Group D6 cows than in Group D11 cows. The progesterone (P4) levels decreased to less than 1 ng/ml in all groups on the day after PG treatment. The estradiol-17ß (E2) levels began to increase after PG treatment and peaked at 2 days before ovulation in the cows that ovulated. In anovulated cows, E2 tended to be higher and there was no clear E2 peak in some cows. These results indicated that the number of days to ovulation, the type of ovulated DF, and anovulation were affected by factors that were associated with the DF when it was producing E2.