Co-transmission of <Emphasis Type="Italic">Tomato yellow leaf curl virus</Emphasis> (TYLCV)-Mld and TYLCV-IL by the whitefly <Emphasis Type="Italic">Bemisia tabaci</Emphasis>

The ability of the whitefly Bemisia tabaci to transmit two strains of Tomato yellow leaf curl virus, the Israel and Mild strains, was studied after serial transfers of individual whiteflies that were viruliferous for both strains to tomato plants. After single whiteflies had successive acquisition feedings first on a single plant infected with one strain and then on a plant infected with the other strain, the single whiteflies later transmitted intermittently one, the other, or both strains to the test plants during serial transfers at 1-day intervals. Because both strains were found in the head, abdomen, and legs dissected from whiteflies during the retention period after the two successive acquisition feedings, both strains apparently circulate from midgut cells to salivary glands through the hemolymph.     

Seed productivity test of CMS lines of Japanese bunching onion (Allium fistulosum L.) possessing the cytoplasm of a wild species, A. galanthum Kar. et Kir.

In a previous study, we developed male sterile lines of Japanese bunching onion (Allium fistulosum L.) possessing the cytoplasm of a wild species, A. galanthum Kar. et Kir., by backcrossing. To evaluate seed productivity of the male sterile lines in practise, they were crossed with the male fertile line, cultivar 'Kujyo', using honeybees as pollinators under field conditions. The number of florets and seeds per inflorescence, seed set and seed germination of the material were investigated. Although variation was observed among the male sterile lines, there were several lines having seed productivity equal to cultivar 'Kujyo'. Our data demonstrate that the male sterile lines of A. fistulosum possessing the cytoplasm of A. galanthum are useful as seed parents for the commercial F seed production of A. fistulosum.     

A selective barrier in the midgut epithelial cell membrane of the nonvector whitefly <Emphasis Type="Italic">Trialeurodes vaporariorum</Emphasis> to <Emphasis Type="Italic">Tomato yellow leaf curl virus</Emphasis> uptake

We studied the presence of a potential transmission barrier that blocks Tomato yellow leaf curl virus in the nonvector greenhouse whitefly, Trialeurodes vaporariorum. Because T. vaporariorum can ingest and retain the virus after acquisition feeding on an infected plant, comparable to the vector whitefly Bemisia tabaci, circumstance evidence suggested that a transmission barrier presents at location(s) where the virus moves from the digestive tract lumen to the hemolymph. To provide direct evidence for the site of a transmission barrier in the nonvector insect, we compared the accumulation levels and localization of the virus between the two species of whiteflies. Quantitative real-time and conventional PCR analysis showed that accumulation of the virus during acquisition feeding and retention after a short acquisition period were indistinguishable between the two species, but the circulation of the virus within the whiteflies differed significantly between the species. In an immunofluorescence analysis using an antibody specific to the coat protein of the virus, the virus was restricted to the luminal surface of the midgut epithelial cells and did not enter their cytoplasm or that of the salivary glands in T. vaporariorum. In contrast, the virus was localized within the cytoplasm of the midgut epithelial cells and in the paired salivary glands of B. tabaci adults. This direct evidence shows that a selective transmission barrier at the luminal membrane surface of midgut epithelial cells in the nonvector whitefly blocks entrance of the virus into the midgut epithelial cells, resulting in incompetence as a vector of the virus.     

Pathogenicity of Bordetella bronchiseptica isolated from apparently healthy rabbits in guinea pig,rat, and mouse

Bordetella bronchiseptica (B. bronchiseptica) is associated with respiratory tract infections in laboratory animals. In our laboratory animal facility, B. bronchiseptica was isolated from 21 of 27 apparently healthy rabbits obtained from a breeding farm contaminated with B. bronchiseptica. Restriction fragment length polymorphism (RFLP) analysis showed that the flagellin genotype of isolates from the laboratory animal facility and breeding farm was type A, which is seen relatively frequently in rabbits in Europe. To examine its pathogenicity, guinea pigs, rats, and mice were inoculated intranasally with a representative strain isolated in the laboratory animal facility. Following inoculation of 10⁷ colony forming unit (cfu), severe inflammation was observed in the lungs of guinea pig and mice, although the inflammation was less severe in rats. The strain was recovered from the trachea and lungs of these species after inoculation with lower dose such as 10³ or 10⁴ cfu. These results suggest that the isolated strain causes respiratory tract infection in guinea pigs, rats, and mice, and that its pathogenicity higher in mice than in rats. This study extends our knowledge of interpreting the microbiologic status of laboratory animals, which will contribute to the development of reliable and reproducible animal experiments.     

Proteome analysis of whole and water‐soluble proteins in masseter and semitendinosus muscles of Holstein cows

To assess both quantitative and qualitative differences between the slow‐ and fast‐type muscles, masseter (slow) and semitendinosus (fast) from four Holstein cows were analyzed by two‐dimensional difference gel electrophoresis (2D DIGE) and mass spectrometry. The proteome analysis identified 27 spots as 20 proteins in the whole protein fraction extracted with 8 mol/L urea solution, and 16 spots were identified as 11 proteins in the water‐soluble protein fraction. Two slow‐type myofibrillar proteins (myosin light chain‐1 slow‐b and myosin light chain‐2 slow), and aconitase‐2 mitochondria were present at higher levels in the masseter muscle (P < 0.05). Four fast‐type myofibrillar proteins (myosin light chain‐1 fast, myosin light chain‐2 fast, myosin light chain‐3 fast and tropomyosin‐1), and three enzymes of glycolytic pathway (enolase‐3, aldolase‐A and triosephosphate isomerase), were present at higher levels in the semitendinosus muscle (P < 0.05). Our proteome analysis showed that the composition of sarcoplasmic proteins as well as myofibrillar proteins was clearly different between slow‐ and fast‐type muscles.     

QTL analysis of morphological traits and pseudostem pigmentation in bunching onion (<Emphasis Type="Italic">Allium fistulosum</Emphasis>)

Pseudostems of bunching onion (Allium fistulosum L.) show wide variation in morphological traits and skin color. However, despite being one of the most important agronomic traits, molecular studies of bunching onion pseudostems remain limited. In this study, six morphological traits (plant height, leaf length, pseudostem length, leaf width, pseudostem width and number of leaf sheaths) along with pseudostem pigmentation indices were evaluated in two field trials using an F_2:3 population derived from a single F₁ cross between a white single pseudostem (non-tillering) and a red tillering bunching onion. Plant height was highly correlated with both leaf length and pseudostem length, but not the number of leaf sheaths. In contrast, the number of leaf sheaths was significantly negatively correlated with both leaf width and pseudostem width. A total of 27 QTLs for the six morphological traits were detected in 16 regions of 11 linkage groups, with a major QTL for the number of leaf sheaths repeatedly detected on Chr. 8. Meanwhile, two QTLs associated with pseudostem pigmentation were repeatedly detected on linkage groups Chr. 4a and Chr. 5a-2. The latter (qPig5a-2) was considered a major QTL, and its location estimated by marker genotyping of the F₂ population around the qPig5a-2 region as being within a 7.6 cM interval.     

Inheritance mode of male sterility in bunching onion (<Emphasis Type="Italic">Allium fistulosum</Emphasis> L.) accessions

Cytoplasmic male sterility (CMS) is an indispensable trait for F₁ hybrid seed production in bunching onion (Allium fistulosum L.). Expansion of the cytoplasmic diversity of F₁ hybrid cultivars by introduction of various CMS resources has great potential to eliminate vulnerability to cytoplasm type-specific diseases. This study aimed to evaluate appearance frequency of male sterile plants in several bunching onion accessions and to identify CMS resources. In eight (‘Nogiwa Aigara’, ‘Bansei Hanegi’, ‘Amarume’, ‘Kimnung’, ‘Zhangqiu’, ‘INT/CHN/1990/GOTOU’, ‘Natsunegi’ and ‘Guangzhou’) of 135 accessions collected from Japan, China, Mongolia, Korea and Taiwan, male sterile plants appeared with varied frequencies from 1.7% (‘Nogiwa Aigara’ and ‘Bansei Hanegi’) to 24.5% (‘Zhangqiu’). The inheritance mode of Zhangqiu- and Guangzhou-derived male sterility was confirmed to be CMS by sib-crossings and interbreed crossings. Microscopic examination of microsporogenesis in the CMS plants revealed that microspore protoplasm rapidly degenerated without mitotic division after the release of microspores from tetrads. The CMS germplasm described here would be useful for the development of “A” lines to be used in F₁ hybrid seed production of bunching onion. Male fertility in ‘Nogiwa Aigara’, ‘Bansei Hanegi’, ‘Kimnung’, ‘INT/CHN/1990/GOTOU’ and ‘Natsunegi’ was verified to be controlled by a single fertility restoration locus.