First molecular genotyping of voltage gated sodium channel alleles in Culex quinquefasciatus populations in Malaysia

A nationwide investigation was performed to detect the presence of 1014 mutation(s) in voltage gated sodium channel (kdr) gene of Culex quinquefasciatus from 14 residential areas across 13 states and a federal territory in Malaysia. Molecular genotyping of kdr mutation was performed via a modified three tubes allele-specific-polymerase chain reaction (AS-PCR) and direct sequencing of kdr gene. Based on the results of AS-PCR, homozygous susceptible (SS) genotype was found in nine out of 14 populations with 38 individuals from a total sample size of 140. Heterozygous (RS) genotype was most predominant (99 individuals) and distributed across all study sites. Homozygous resistance (RR) genotype was detected in Perak (one individual) and Selangor (two individuals). The resistance kdr allele frequencies ranged from 0.1 to 0.55, with the highest being detected in Cx. quinquefasciatus population from Selangor. This study has documented the first field-evolved instance of 1014F mutation in Malaysian mosquitoes and the findings of this study could be utilized in the implementation of strategic measures in vector control programs in Malaysia.     

Variation in growth rates and aggressiveness of naturally occurring self‐fertile and self‐sterile isolates of the wilt pathogen Ceratocystis albifundus

Ceratocystis albifundus is the most important fungal pathogen of black wattle (Acacia mearnsii) grown in plantations in southern and eastern Africa. It is a homothallic fungus but also undergoes unidirectional mating type switching. As a result, the ascospore progeny can be either self‐fertile or self‐sterile. The only apparent difference between these mating types is the deletion of the MAT1‐2‐1 gene in self‐sterile isolates. There is some evidence suggesting that self‐sterile isolates grow more slowly than self‐fertile isolates, but this has not been tested rigorously. The aim of this study was to determine whether self‐sterile isolates are less fit by examining growth rate, relative germination rate and pathogenicity. Five self‐sterile isolates were generated from each of five self‐fertile isolates of C. albifundus and these 30 isolates were compared. The results showed that the self‐sterile isolates grew consistently slower and were less pathogenic than the self‐fertile isolates. The germination ratio of self‐fertile to self‐sterile isolates from single ascospores collected from the ascomata of five self‐fertile isolates was on average 7:3. This could be a consequence of the self‐sterile isolates having a lower germination rate. This observation, and the lower growth and pathogenicity levels, suggests that self‐sterile isolates are not likely to compete effectively in nature, raising intriguing questions regarding their role and value to C. albifundus and other fungi having a similar mating system.     

Dietary protein requirement for juvenile mandarin fish,Siniperca scherzeri

An 8‐week feeding trial was conducted to investigate the dietary protein requirement of juvenile mandarin fish, Siniperca scherzeri, a demersal freshwater piscivore with high commercial value in East Asia. Five isolipidic (100 g/kg) and isoenergetic (20 MJ/kg) practical diets were formulated to contain graded levels of 450, 500, 550, 600, and 650 g/kg crude protein, using mackerel fishmeal as the sole source of protein. Each of the five test diets was fed to visual satiety to triplicate groups of fish (initial mean body weight ± SE; 20.1 ± 0.2 g) reared in a recirculating freshwater system over the experimental period. Fish growth performance in terms of weight gain tended to improve with increasing dietary protein level, and the highest values were observed in those fish fed the 600–650 g/kg diets. Feed intake significantly declined as the protein content of the diet increased. Feed efficiency and protein retention tended to increase with increasing dietary protein levels, from 450 to 600 g/kg, and then declined when dietary protein content further increased to 650 g/kg. A similar trend was also found for the protein efficiency ratio, although the observed changes did not reach statistical significance. Whole‐body composition and plasma biochemical parameters were not significantly affected by the dietary protein content. Based on these findings and a broken‐line model of growth, 614.4 g/kg seems to be the proper amount of protein in a practical diet for >20 g mandarin fish.     

Evaluation of Seed Vigor Tests for Predicting Seedling Establishment at Low Temperature in Rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

This study was carried out to develop an evaluation method to predict rice seedling establishment (SE) under low-temperature conditions. Two Korean-bred japonica cultivars, Shindongjin and Hopum were used in the experiment. Fresh seeds were treated with an accelerated aging (AA) at 40°C and 100% RH for 1-15 days. The SEs of the fresh and AA seeds were evaluated in nursery beds at 17°C, and their correlation coefficients with seed vigor values measured by 9 test methods including standard germination test (SGT), cool germination test (CLT), cold germination test (CDT), seedling growth rate test (SGRT), ascorbate peroxidase (APX), peroxidase (POX), superoxide dismutase (SOD), catalase (CAT), and α ‐amylase (AMY) activities. The percentage of SE decreased slowly from 75 to 0% with an increasing of AA period from 0 to 15 days. The result of nine vigor tests showed different correlations with the SE. SGT, CLT, SGRT, and POX were significantly correlated with the SE. In the correlation analysis with only short-term aging seeds (1-7 days), the SE was very highly correlated with SGT, CLT, CDT, SGRT, POX, and CAT. These results suggest that seed vigor values measured by several methods including SGT and POX could be used as a reference value to secure SE at low temperatures in nursery bed rice seedling culture.     

Three AFLP markers tightly linked to the genic male sterility <Emphasis Type="Italic">ms</Emphasis><Subscript><Emphasis Type="Italic">3</Emphasis></Subscript> gene in chili pepper (<Emphasis Type="Italic">Capsicum annuum</Emphasis> L.) and conversion to a CAPS marker

Genic male sterility (GMS) has long been used as a tool for hybrid seed production in chili pepper (Capsicum annuum L.). We developed DNA markers linked to the GMS ms ₃ gene in a segregating population using bulked segregant analysis (BSA) and amplified fragment length polymorphism (AFLP) techniques. The segregating population was subjected to BSA-AFLP with 512 primer combinations. Three AFLP markers (Eagg/Mccc₂₇₆, Eagc/Mctt₁₇₈, and Ecag/Mtgc₂₀₄) were identified as tightly linked to the ms ₃ locus. Among them, we converted the AFLP marker Ecag/Mtgc₂₀₄ to the cleavage amplified polymorphic sequence (CAPS) marker, named GMS3-CAPS, based on sequencing analysis of internal and flanking regions for the markers between male-fertile and sterile plants. This marker will be useful for pepper breeding using the GMS system.     

Genetic, agronomic and quality comparisons of two 1AL.1RS. wheat-rye chromosomal translocations

The 1AL.1RS wheat-rye chromosomal translocation originally found in ‘Amigo’ wheat possesses resistance genes for stem rust, powdery mildew and greenbug biotypes B and C, but also has a negative effect on wheat processing quality. Recently, a second 1AL.1RS translocation carrying Gb6, a gene conferring resistance to greenbug biotypes B, C, E, G and I, was identified in the wheat germplasm line ‘GRS1201′. Protein analytical methods, and the DNA polymerase chain reaction were used to identify markers capable of differentiating the 1RS chromosome arms derived from ‘Amigo’ and ‘GRS1201′. The secalin proteins encoded by genes on 1RS chromosome arms differed in ‘Amigo’ and ‘GRS1201′. A 70 kDa secalin was found in the ‘Amigo’1AL.1RS, but did not occur in the ‘GRS1201’1AL.1RS. Polymorphisms detected by PCR primers derived from a family of moderately repetitive rye DNA sequences also differentiated the two translocations. When ‘GRS1201’was mated with a non-1RS wheat, no recombinants between 1RS markers were observed. In crosses between 1RS and non-1RS parents, both DNA markers and secalins would be useful as selectable markers for 1RS-derived greenbug resistance. Recombination between 1RS markers did occur when 1RS from ‘Amigo’ and 1RS from ‘GRS1201’were combined, but in such intermatings, the molecular markers described herein could still be used to develop a population enriched in lines carrying Gb6. No differences in grain yield or grain and flour quality characteristics were observed when lines carrying 1RS from ‘Amigo’ were compared with lines with 1RS from ‘GRS1201′. Hence, differences in secalin composition did not result in differential quality effects. When compared with sister lines with 1AL.1AS derived from the wheat cultivar ‘Redland’, lines with ‘GRS1201’had equal grain yield, but produced flours with significantly shorter mix times, weaker doughs, and lower sodium dodecyl sulphate sedimentation volumes.     

Dietary eicosapentaenoic acid requirement of juvenile rock bream,Oplegnathus fasciatus

A feeding trial was conducted to evaluate the optimum dietary level of eicosapentaenoic acid (EPA, 20:5n‐3) based on growth and non‐specific immune responses in juvenile rock bream. A basal diet without EPA supplementation was used as a control, and six other diets were prepared by supplementing with 4, 8, 12, 16, 20 or 40 g of EPA per kg diet. The actual EPA concentrations of the diets were 0.5, 4.3, 8.5, 13.0, 16.8, 21.0 and 41.2 g of EPA per kg diet, and the diets were abbreviated as EPA_0.5, EPA_4.3, EPA_8.5, EPA_13.0, EPA_16.8, EPA_21.0 and EPA_41.2, respectively. Triplicate groups of fish averaging 1.06 ± 0.01 g (mean ± SD) were fed one of the seven experimental diets at the apparent satiation for 8 weeks. At the end of the feeding trial, weight gain, specific growth rate and feed efficiency of fish fed EPA_16.8, EPA_21.0 and EPA_41.2 diets were significantly higher than those of fish fed EPA_0.5, EPA_4.3, EPA_8.5 and EPA_13.0 diets (p < .05). Superoxide dismutase activity of fish fed EPA_16.8, EPA_21.0 and EPA_41.2 diets were significantly higher than those of fish fed EPA_0.5, EPA_4.3 and EPA_8.5 diets. Fish fed EPA_21.0 and EPA_41.2 diets showed significantly higher lysozyme activity than did fish fed EPA_0.5, EPA_4.3, EPA_8.5 and EPA_13.0 diets. The broken‐line analysis of weight gain indicated that the optimum dietary EPA level was 16.7 g/kg diet. These results suggested that the optimum dietary EPA level in juvenile rock bream could be greater than 16.7 g/kg diet but less than or equal to 16.8 g/kg diet based on the broken‐line analysis and the ANOVA test of weight gain.     

山西猪场猪蓝耳病并发猪胸膜肺炎放线杆菌的病毒鉴定及细菌分离鉴定以及治疗方案

2020年2月份山西某大型规模猪场育肥猪突发呼吸困难,无过多症状表现,即突然死亡,发病率为2%,死亡率1.5%左右。实验室以病死猪为研究对象,综合死亡猪只脏器病变状况观察,无菌采集病死猪的肺脏、肝脏、脾脏病料,进行RT-PCR检测猪蓝耳病原;用含1%NAD和5%胎牛血清的TSA琼脂培养基进行细菌分离培养和纯化,经形态学鉴定、PCR鉴定、16Sr RNA基因序列进化树构建等方法,结果显示:该发病猪群为猪繁殖与呼吸障碍综合征并发猪胸膜肺炎放线杆菌感染。药敏试验结果显示:该猪胸膜肺炎放线杆菌对安普霉素、硫酸黏菌素、替米考星+氟苯尼考、替米考星+阿莫西林、氟苯尼考、头孢噻呋钠+硫酸黏菌素、头孢喹肟、头孢噻呋钠+新霉素、替米考星+头孢噻呋钠、多西环素+氟苯尼考、头孢噻呋钠+阿莫西林、阿莫西林+克拉维酸钾、头孢噻呋钠+恩诺沙星、新霉素、头孢噻呋钠、安普霉素+头孢噻呋钠敏感。对恩诺沙星耐药。本实验室根据临床及检测结果制定治疗方案,为该猪场的猪繁殖与呼吸障碍综合征并发猪胸膜肺炎的临床诊断及用药提供了理论依据。