The Detection of Hamster Connexins: A Comparison of Expression Profiles with Wild-Type Mouse and the Cancer-Prone Min Mouse

The open reading frames of 17 connexins from Syrian hamster (using tissues) and 16 connexins from the Chinese hamster cell line V79, were fully (Cx30, Cx31, Cx37, Cx43 and Cx45) or partially sequenced. We have also detected, and partially sequenced, seven rat connexins that previously were unavailable. The expression of connexin genes was examined in some hamster organs and cultured hamster cells, and compared with wild-type mouse and the cancer-prone Min mouse. Although the expression patterns were similar for most organs and connexins in hamster and mouse, there were also some prominent differences (Cx29 and 30.3 in testis; Cx31.1 and 32 in eye; Cx46 in brain, kidney and testis; Cx47 in kidney). This suggests that some connexins have species-specific expression profiles. In contrast, there were minimal differences in expression profiles between wild type and Min mice. Species-specific expression profiles should be considered in attempts to make animal models of human connexin-associated diseases.     

Temperature,deltamethrin-resistance status and performance measures of Plutella xylostella: complex responses of insects to environmental variables

1. Worldwide, the excessive use of insecticides has resulted in field-evolved insecticide-resistant populations of diamondback moth (DBM), Plutella xylostella. A deltamethrin-resistant DBM population from the field was divided into two subpopulations in the laboratory. One population (S-strain) was maintained with no further exposure to insecticides, whereas the other population (R-strain) was maintained under a regime of intermittent selection with deltamethrin. 2. Individuals from both strains were reared at constant temperatures in the range 10–35 °C in the absence of deltamethrin, and the effects of rearing temperature on various traits were investigated. At the time of experimentation, the R-strain was 20-fold more resistant to deltamethrin than the S-strain. 3. Temperature differentially affected developmental time, adult life span, pupal weight, and fecundity of both strains. Although both strains laid eggs after being reared at 10 °C, few of these eggs were fertile. The R-strain developed significantly faster than the S-strain. The integrated performance of the S-strain and R-strain was greatest at 25 and 15 °C, respectively. 4. The present study provides important information on the complexities of the outcomes of the interactions between ectotherms and temperature. Specifically, temperature-trait relationships may not be unimodal, and ectotherm genotypes (in this case insecticide-resistance status) and abiotic stresses can interact with unpredictable outcomes. 5. Current models predicting DBM population dynamics and relative abundance in different locations do not consider different thermal biologies of different genotypes. The present study shows the dramatic effects of environment on many parameters used in these models and will help to enhance their accuracy, and thus their utility.     

A combined transcriptional,biochemical and histopathological study unravels the complexity of Alternaria resistance and susceptibility in Brassica coenospecies

Alternaria blight is one of the most devastating diseases of rapeseed-mustard caused by a necrotrophic fungus Alternaria brassicae. Lack of satisfactory resistance resource in Brassica is still a main obstruction for developing resistance against Alternaria. In this study, we have selected Brassica juncea, Sinapis alba and Camelina sativa to understand and unravel the mechanism of disease resistance against Alternaria. Histopathological studies showed early onset of necrosis in B. juncea (1 dpi) and delayed in S. alba (2 dpi) and C. sativa (3 dpi) respectively. Early and enhanced production of hydrogen peroxide (H₂O₂) was observed in C. sativa and S. alba (6 hpi) when compared to B. juncea (12 hpi). An increase in catalase activity was observed in both C. sativa (36 % at 6 hpi) and S. alba (15 % at 12 hpi), whereas it significantly decreased in B. juncea at 6 hpi (23 %), 12 hpi (30 %) and 24 hpi (8 %). Gene expression analysis showed induction of PR-3 and PR-12 genes only in C. sativa and S. alba when compared to B. juncea suggesting their vital role for Alternaria resistance. In contrast, SA marker genes were significantly expressed in B. juncea only which provides evidence of hormonal cross talk in B. juncea during Alternaria infection thereby increasing its susceptibility.     

Gene expression changes associated with cytotoxicity identified using cDNA arrays

In order to investigate gene expression changes associated with cytotoxicity, we used cDNA arrays to monitor the expression of over 5,000 genes in response to toxic stress in the HepG2 liver cell line. Cells were treated with cytotoxic doses of acetaminophen, caffeine or thioacetamide for nine time points ranging from 1 to 24 h. Samples of mRNA from each time point were used to prepare radiolabeled cDNA, which was hybridized to nylon-membrane-based cDNA arrays. High-stringency washes were applied to reduce cross-hybridization. Analysis of spot intensities revealed that each compound led to approximately 150-250 gene expression changes that were sustained over at least three adjacent time points. The affected genes could be classified into clusters based on their temporal patterns of differential expression. A common set of 44 genes showed similar expression changes in response to all three compounds. Of these changes, 90% could be confirmed by quantitative RT-PCR analysis. The results indicate that detailed array-based time-course studies, coupled with a sensitive and highly specific confirmation assay, provide a powerful means of identifying cytotoxicity-associated gene expression changes. Electronic Publication     

Association of the colistin resistance gene mcr-1 with faecal pollution in water environments in Hanoi,Vietnam

Colistin is one of the antibiotics of last resort for human health. However, the dissemination of the plasmid-mediated colistin resistance gene mcr-1 is of great concern globally. In the One Health framework, the environment is an important component for managing antimicrobial resistance. However, little information is available concerning the prevalence of mcr-1 in water environments. We aimed to reveal the prevalence of mcr-1 in different water environments in Hanoi, Vietnam. Quantitative PCR was applied to detect mcr-1 in four urban drainages receiving untreated domestic wastewater, three rivers, five lakes and two groundwater samples. Urban drainages contained higher concentrations of mcr-1, suggesting that urban residents carry the gene. The class 1 integron-integrase gene was identified as a good surrogate of antibiotic resistance genes including mcr-1. A significant correlation was found between the levels of mcr-1 and the human-specific cross-assembly phage, which is an indicator of human faecal pollution. These results indicated that the primary source of mcr-1 in urban water environments is human faeces, which is consistent with the fact that most domestic wastewater is untreated in Hanoi. The control of untreated wastewater is critical for alleviating the spread of mcr-1 in water environments in Vietnam.     

Genetics and Mechanism of Resistance to Meloidogyne arenaria in Peanut Germplasm

Segregation of resistance to Meloidogyne arenaria in six BC₅F₂ peanut breeding populations was examined in greenhouse tests. Chi-square analysis indicated that segregation of resistance was consistent with resistance being conditioned by a single gene in three breeding populations (TP259-3, TP262-3, and TP271-2), whereas two resistance genes may be present in the breeding populations TP259-2, TP263-2, and TP268-3. Nematode development in clonally propagated lines of resistant individuals of TP262-3 and TP263-2 was compared to that of the susceptible cultivar Florunner. Juvenile nematodes readily penetrated roots of all peanut genotypes, but rate of development was slower (P = 0.05) in the resistant genotypes than in Florunner. Host cell necrosis indicative of a hypersensitive response was not consistently observed in resistant genotypes of either population. Three RFLP loci linked to resistance at distances of 4.2 to 11.0 centiMorgans were identified. Resistant and susceptible alleles for RFLP loci R2430E and R2545E were quite distinct and are useful for identifying individuals homozygous for resistance in segregating populations.     

Identification of up-regulated genes in human uterine leiomyoma by suppression subtractive hybridization

In searching for differentially expressed genes in human uterine leiomyomas (ULs), suppression sub-tractive hybridization was used to construct an UL up-regulated library, which turned out to represent 88genes. After two rounds of screening by reverse Northern analysis, twenty genes were proved to be up-regulated, including seventeen known genes and three genes with unknown function. All these genes werefirstly associated with UL. Three genes with notable difference were selected for Northern confirmationOur results proved the authenticity of the twenty genes. One gene named Phospholipase A2 (PLA2) showedup-regulation in 4/6 of the patients and investigation of tissue distribution indicated that it had obviousexpression in prostate, testis, liver, heart and skeletal muscle.     

Embryo culture of Medicago scutellata and M. sativa

Resistance to the alfalfa weevil (Hypera postica (Gyllenhal)) and the potato leafhopper (Empoasco fabae (Harris)) is lacking in cultivated alfalfa. However, a closely related annual Medicago, Medicago scutellata, possesses dense glandular stem and leaf hairs which provides a mechanism for resistance. Several attempts have been made at transfering the glandular haired traint from M. scutellata to perennial alfalfa with limited success. Earlier studies have shown that one reason for the lack of success is embryo abortion. Therefore, this study was initiated to observe zygotic embryo-genesis and to develop an embryo rescue technique for M. scutellata and M. sativa. Observations of zygotic embryogenesis showed that the two species are similar in morphology and can be described from youngest to oldest as globular, heart, torpedo, and hook shaped embryos. M. sativa embryos are smaller than M. scutellata embryos and develop three to four days later. Self pollinated M. scutellata (PI 307446) and sib mated M. sativa (Saranac AR) embryos were cultivated on Murashige and (2,4-D), indolacetic acid (IAA), 6-benzylaminopurine (BAP), and kinetic (KIN). Embryos from both species were also cultured on Schenk and Hildebrandt's (SH) basal medium with the addition of L-glutamine and L-proline. The experimental design was a completely randomized factorial for each experiment. Heart and torpedo shaped embryos from M. scutellata grew best (27.5% plantlet recovery) when cultured on MS medium with 0.05 mgl^-1 of both IAA and BAP. After 15 to 30 days on this medium, the embryos had only developed shoots. Therefore, it was necessary to transfer the shoots to MS basal medium without phytohormones for rooting. Rooting occurred in 15 to 30 days and the plantlets could be acclimatized to soil within 2 to 4 weeks. M. sativa embryos grew best (31% plantlet recovery) on SH medium with 50 mM L-glutamine. M. sativa embryos developed both shoots and roots on this medium. This information may now be applied to the development of an embryo culture method for recovering insect resistant hybrids between M. scutellata and M. sativa. Disclaimer statement: Mention of a trademark or proprietary product does not constitute a guarantee or warranty of the product by the USDA, and does not imply its approval to the exclusion of other products that may also be suitable.