Effect of physiological heterogeneity of E. coli popula-tion on antibiotic susceptivity test

According to the instantaneous growth rate (dN/dt) of E. coli CVCC249 growing in batch culture, the entire growth progress was distinguished into four phases: accelerating growth phase, constant growth phase, decelerating growth phase and declining phase, in each of which obvious variation in physiological and biochemical properties was detected, including total DNA, total protein, and MTT-dehydrogenase activity, etc., that led to difference in their antibiotic susceptivity. Antibiotic susceptivity of the population sampled from each phase was tested by Concentration-killing Curve (CKC) approach following the formula N=N ₀/{1+exp[r·(x-BC ₅₀)]}, showing as normal distribution at the individual cell level for an internal population, in which the median bactericidal concentration BC ₅₀ represents the mean level of susceptivity, while the bactericidal span BC ₁₋₉₉=(2lnN ₀)/r indicates the variation degree of the antibiotic susceptivity. Furthermore, tested by CKC approach, the antibiotic susceptivity of E. coli CVCC249 population in each physiological phase to gentamicin or enoxacin was various: susceptivity of the population in the constant growth phase and declining phase all increased compared with that in the accelerating growth phase for gentamicin but declined for enoxacin. The primary investigations revealed that the physiological phase should be taken into account in the context of antibiotic susceptivity and research into antimicrobial mechanism. However there are few reports concerned with this study. Further research using different kinds of antibiotics with synchronized continuous culture of different bacterial strains is required. Supported by the Natural Science Foundation of Shandong Province, China (Grant No. Y2005C58), the Natural Key Technology R&D Program of China (Grant No. 2006BAK02A03-6) and the Youth Scientific Research Foundation of Shandong Academy of Agricultural Science (2005YQ035)     

Antimicrobial Peptide JH-3 Effectively Kills Salmonella enterica Serovar Typhimurium Strain CVCC541 and Reduces Its Pathogenicity in Mice

Salmonella is an important zoonotic pathogen and is a major cause of gastrointestinal diseases worldwide. The current serious problem of antibiotic abuse has prompted the search for new substitutes for antibiotics. JH-3 is a small antimicrobial peptide with broad-spectrum bactericidal activity. In this study, we showed that JH-3 has good bactericidal activity towards the clinical isolate Salmonella enterica serovar Typhimurium strain CVCC541. The minimum inhibitory concentration (MIC) of JH-3 against this bacterium was determined to be 100 μg/mL, which could decrease the number of CVCC541 cells by 1000-fold in vitro within 5 h. The transmission electron microscopy (TEM) results showed that JH-3 can damage the cell wall and membrane of CVCC541, leading to the leakage of cell contents and subsequent cell death. To measure the bactericidal activity of CVCC541-infected mice were treated intraperitoneally 40 or 10 mg/kg JH-3 at 2 h or 3 days postinfection. Our results showed that treatment with 40 mg/kg JH-3 at 2 h postinfection had the best therapeutic effect and could significantly protect mice from a lethal dose of CVCC541. Furthermore, the clinical symptoms, bacterial burden in blood and organs, and intestinal pathological changes were all decreased and were close to normal. This study examined the therapeutic effect of the antimicrobial peptide JH-3 against S. enterica CVCC541 infection for the first time and determined the therapeutic effect of different JH-3 doses and treatment times, laying the foundation for studies of new antimicrobial agents.

     

Influence of agitation and aeration on growth and antibiotic production by <Emphasis Type="Italic">Xenorhabdus nematophila</Emphasis>

The effect of agitation and aeration on the growth and antibiotic production by Xenorhabdus nematophila YL001 grown in batch cultures were investigated. Efficiency of aeration and agitation was evaluated through the oxygen mass transfer coefficient (K _L a). With increase in K _L a, the biomass and antibiotic activity increased. Activity units of antibiotic and dry cell weight were increased to 232 U ml⁻¹ and 19.58 g l⁻¹, respectively, productivity in cell and antibiotic was up more than 30% when K _L a increased from 115.9 h⁻¹ to 185.7 h⁻¹. During the exponential growth phase, DO concentration was zero, the oxygen supply was not sufficient. So, based on process analysis, a three-stage oxygen supply control strategy was used to improved the DO concentration above 30% by controlling the agitation speed and aeration rate. The dry cell weight and activity units of antibiotic were further increased to 24.22 g l⁻¹ and 249 U ml⁻¹, and were improved by 24.0% and 7.0%, compared with fermentation at a constant agitation speed and a constant aeration rate (300 rev min⁻¹, 2.5 l min⁻¹).     

Low effective population size and survivorship in a grassland grouse

Assessments of census size (N _c) and effective population size (N _e) are necessary for the conservation of species exhibiting population declines. We examined two populations (Oklahoma and New Mexico) of the lesser prairie-chicken (Tympanuchus pallidicinctus), a declining lek-breeding bird, in which one population (Oklahoma) has larger clutch size and more nesting attempts per year but lower survival caused by human changes to the landscape. We estimated demographic and genetic estimates of N _e for each population and found that both populations have low N _e estimates with a risk of inbreeding depression. Although Oklahoma females produce a larger number of offspring, the proportion of females successfully reproducing is not higher than in New Mexico. Higher reproductive effort has likely reached a physiological limit in Oklahoma prairie-chickens but has not led to a higher N _e or even a larger N _c than New Mexico. We propose that future conservation efforts focus on maximizing survivorship and decreasing the variance in reproductive success because these factors are more likely than increasing reproductive output alone to yield population persistence in lek-breeding species.     

Glycoalkaloids and acclimation capacity of hybrids between <Emphasis Type="Italic">Solanum tuberosum</Emphasis> and the incongruent hardy species <Emphasis Type="Italic">Solanum commersonii</Emphasis>

F₁ and backcross hybrids between sexually incompatible species Solanum commersonii and Solanum tuberosum were characterized for glycoalkaloid content and capacity to cold acclimate. Glycoalkaloid (GA) analysis revealed that F₁ triploids and BC₁ pentaploids contained the glycoalkaloids of both parents. In BC₂ (near) tetraploids the situation was different, in that some hybrids produced the GAs of both parents, whereas others contained only the GAs of S. tuberosum. This suggested that the GAs from S. commersonii may be lost rapidly, and that they may have a simple genetic control. The total tuber GA content of BC₁ and BC₂ groups averaged quite acceptable levels (165.9 mg/kg in BC₁ and 192.8 mg/kg in BC₂), with six genotypes having a GA content <200 mg/kg fresh weight. The F₁ triploid hybrids expressed a capacity to cold acclimate similar to S. commersonii, whereas BC₁ and BC₂ genotypes generally displayed an acclimation capacity higher than the sensitive parent but lower than S. commersonii. However, one BC₁ and two BC₂ genotypes with an acclimation capacity as high as S. commersonii were identified. The polar lipid fatty acid composition in S. commersonii and its hybrid derivatives showed that, following acclimation, there was a significant increase in 18:3. Correlation analysis between the capacity to cold acclimate and the increase in 18:3 was significant, suggesting that the increase in 18:3 can be used as a biochemical marker for the assisted selection of cold-acclimating genotypes in segregating populations.Communicated by G. Wenzel     

When are genetic methods useful for estimating contemporary abundance and detecting population trends?

The utility of microsatellite markers for inferring population size and trend has not been rigorously examined, even though these markers are commonly used to monitor the demography of natural populations. We assessed the ability of a linkage disequilibrium estimator of effective population size (N_e) and a simple capture-recapture estimator of abundance (N) to quantify the size and trend of stable or declining populations (true N = 100–10,000), using simulated Wright–Fisher populations. Neither method accurately or precisely estimated abundance at sample sizes of S = 30 individuals, regardless of true N. However, if larger samples of S = 60 or 120 individuals were collected, these methods provided useful insights into abundance and trends for populations of N = 100–500. At small population sizes (N = 100 or 250), precision of the N_e estimates was improved slightly more by a doubling of loci sampled than by a doubling of individuals sampled. In general, monitoring N_e proved a more robust means of identifying stable and declining populations than monitoring N over most of the parameter space we explored, and performance of the N_e estimator is further enhanced if the N_e/N ratio is low. However, at the largest population size (N = 10,000), N estimation outperformed N_e. Both methods generally required ≥ 5 generations to pass between sampling events to correctly identify population trend.     

Marker-assisted introgression of blackmold resistance QTL alleles from wild Lycopersicon cheesmanii to cultivated tomato (L. esculentum) and evaluation of QTL phenotypic effects

Blackmold, caused by the fungus Alternaria alternata, is a major ripe fruit disease of processing tomatoes. Previously, we found blackmold resistance in a wild tomato (Lycopersicon cheesmanii) and quantitative trait loci (QTL) for resistance were mapped in an interspecific population. Five QTLs were selected for introgression from L. cheesmanii into cultivated tomato using marker-assisted selection (MAS). Restriction fragment length polymorphism and PCR-based markers flanking, and within, the chromosomal regions containing QTLs were used for MAS during backcross and selfing generations. BC₁ plants heterozygous at the QTLs, and subsequent BC₁S₁ and BC₁S₂ lines possessing different homozygous combinations of alleles at the target QTLs, were identified using DNA markers. Field experiments were conducted in 1998 (with 80 marker-selected BC₁S₂ lines) and 1999 (with 151 marker-selected BC₁S₂ and BC₁S₃ lines) at three California locations. Blackmold resistance was assessed during both years, and horticultural traits were evaluated in 1999. The BC₁S₂ and BC₁S₃ lines containing L. cheesmanii alleles at the QTLs were associated with a large genetic variance for resistance to blackmold and moderate heritability, suggesting that significant genetic gain may be achieved by selection in this genetic material. L. cheesmanii alleles at three of the five introgressed QTLs showed a significant, positive effect on blackmold resistance. A QTL on chromosome 2 had the largest positive effect on blackmold resistance, alone and in combination with other QTLs, and was also associated with earliness, a positive horticultural trait. The other four QTLs were associated primarily with negative horticultural traits. Fine mapping QTLs using near isogenic lines could help determine if such trait associations are due to linkage drag or pleiotropy.     

PSMC analysis of effective population sizes in molecular ecology and its application to black‐and‐white Ficedula flycatchers

Climatic fluctuations during the Quaternary period governed the demography of species and contributed to population differentiation and ultimately speciation. Studies of these past processes have previously been hindered by a lack of means and genetic data to model changes in effective population size (N_e) through time. However, based on diploid genome sequences of high quality, the recently developed pairwise sequentially Markovian coalescent (PSMC) can estimate trajectories of changes in N_e over considerable time periods. We applied this approach to resequencing data from nearly 200 genomes of four species and several populations of the Ficedula species complex of black‐and‐white flycatchers. N_e curves of Atlas, collared, pied and semicollared flycatcher converged 1–2 million years ago (Ma) at an N_e of ≈ 200 000, likely reflecting the time when all four species last shared a common ancestor. Subsequent separate N_e trajectories are consistent with lineage splitting and speciation. All species showed evidence of population growth up until 100–200 thousand years ago (kya), followed by decline and then start of a new phase of population expansion. However, timing and amplitude of changes in N_e differed among species, and for pied flycatcher, the temporal dynamics of N_e differed between Spanish birds and central/northern European populations. This cautions against extrapolation of demographic inference between lineages and calls for adequate sampling to provide representative pictures of the coalescence process in different species or populations. We also empirically evaluate criteria for proper inference of demographic histories using PSMC and arrive at recommendations of using sequencing data with a mean genome coverage of ≥18X, a per‐site filter of ≥10 reads and no more than 25% of missing data.     

Fine mapping of quantitative trait loci Hd-1, Hd-2 and Hd-3, controlling heading date of rice, as single Mendelian factors

 Fine mapping was carried out on three putative QTLs (tentatively designated as Hd-1 to Hd-3) of five such QTLs controlling heading date in rice that had been earlier identified using an F₂ population derived from a cross between a japonica variety, ‘Nipponbare’, and an indica variety, ‘Kasalath’, using progeny backcrossed with ‘Nipponbare’ as the recurrent parent. One BC₃F₂ and two BC₃F₁ plants, in which the target QTL regions were heterozygous and most other chromosomal regions were homozygous for the ‘Nipponbare’ allele, were selected as the experimental material. Self-pollinated progeny (BC₃F₂ and BC₃F₃) of the BC₃F₁ or BC₃F₂ showed continuous variation in days to heading. By means of progeny testing based on BC₃F₃ or BC₃F₄ lines, we determined the genotypes of each BC₃F₂ or BC₃F₃ individual at target QTLs. Their segregation patterns fitted Mendelian inheritance ratios. When the results obtained by RFLP analysis and progeny tests were combined, Hd-1, Hd-2 and Hd-3 were mapped precisely on chromosomes 6, 7 and 6, respectively, of a rice RFLP linkage map. The results demonstrated that QTLs can be treated as Mendelian factors. Moreover, these precise locations were in good agreement with the regions estimated by QTL analysis of the initial F₂ population, demonstrating the high reliability of QTL mapping using a high-density linkage map. Received: 5 November 1997 / Accepted: 10 February 1998     

Three QTLs for <Emphasis Type="Italic">Botrytis cinerea</Emphasis> resistance in tomato

Tomato (Solanum lycopersicum) is susceptible to grey mold (Botrytis cinerea). Partial resistance to this fungus was identified in accessions of wild relatives of tomato such as S. habrochaites LYC4. In order to identify loci involved in quantitative resistance (QTLs) to B. cinerea, a population of 174 F₂ plants was made originating from a cross between S. lycopersicum cv. Moneymaker and S. habrochaites LYC4. The population was genotyped and tested for susceptibility to grey mold using a stem bioassay. Rbcq1, a QTL reducing lesion growth (LG) and Rbcq2, a QTL reducing disease incidence (DI) were identified. Rbcq1 is located on Chromosome 1 and explained 12% of the total phenotypic variation while Rbcq2 is located on Chromosome 2 and explained 15% of the total phenotypic variation. Both QTL effects were confirmed by assessing disease resistance in two BC₂S₁ progenies segregating for either of the two QTLs. One additional QTL, Rbcq4 on Chromosome 4 reducing DI, was identified in one of the BC₂S₁ progenies. F₂ individuals, homozygous for the Rbcq2 and Rbcq4 alleles of S. habrochaites showed a reduction of DI by 48%. QTLs from S. habrochaites LYC4 offer good perspectives for breeding B. cinerea resistant tomato cultivars. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Genetic analysis of vertical root pulling resistance (VRPR) in maize using two genetic populations

Root traits are important in improving nutrient and water use efficiency. Vertical root pulling resistance (VRPR) has been shown to be closely related to root system characteristics in maize (Zea mays L.). In the present study, two genetic populations derived from the same parents, one containing 218 recombinant inbred lines (RILs) and the other containing 187 advanced backcross BC₄F₃ lines, were genotyped using 184 SSR markers and evaluated for VRPR, grain yield (GY), stover yield (SY), and nitrogen uptake (N_up) under field conditions over 2 years. Our aims were (1) to locate QTLs associated with VRPR, SY, GY, and N_up, (2) to determine whether QTL detection is consistent between the BC₄F₃ and RIL populations, and (3) to identify backcross lines harboring favorable VRPR QTLs for use in future breeding programs. Using composite interval mapping (CIM), 12 and 17 QTLs were detected in BC₄F₃ and RIL populations, respectively. An important QTL region in bin 6.02 within the interval umc1006-umc1257 was found to control VRPR, SY, and N_up in both populations. These favorable alleles were contributed by the large-rooted parent Ye478. A significant positive correlation was detected between VRPR, SY, and N_up, but not between VRPR and GY. Backcross lines harboring VRPR QTLs could be useful germplasm for developing near isogenic lines (NILs) and for map-based cloning of genes controlling root growth.     

Genetic substructure and admixture as important factors in linkage disequilibrium‐based estimation of effective number of breeders in recovering wildlife populations

The number of effective breeders (N_b) and effective population size (N_e) are population parameters reflective of evolutionary potential, susceptibility to stochasticity, and viability. We have estimated these parameters using the linkage disequilibrium‐based approach with LDNE through the latest phase of population recovery of the brown bears (Ursus arctos) in Finland (1993–2010; N = 621). This phase of the recovery was recently documented to be associated with major changes in genetic composition. In particular, differentiation between the northern and the southern genetic cluster declined rapidly within 1.5 generations. Based on this, we have studied effects of the changing genetic structure on N_b and N_e, by comparing estimates for whole Finland with the estimates for the two genetic clusters. We expected a potentially strong relationship between estimate sizes and genetic differentiation, which should disappear as the population recovers and clusters merge. Consistent with this, our estimates for whole Finland were lower than the sum of the estimates of the two genetic clusters and both approaches produced similar estimates in the end. Notably, we also found that admixed genotypes strongly increased the estimates. In all analyses, our estimates for N_e were larger than N_b and likely reflective for brown bears of the larger region of Finland and northwestern Russia. Conclusively, we find that neglecting genetic substructure may lead to a massive underestimation of N_b and N_e. Our results also suggest the need for further empirical analysis focusing on individuals with admixed genotypes and their potential high influence on N_b and N_e.     

Single leaves photosynthetic characteristics of two insect-resistant transgenic cotton (<Emphasis Type="Italic">Gossypium hirsutum</Emphasis> L.) varieties in response to light

How the photosynthetic characteristics of insect-resistant transgenic cotton (Gossypium hirsutum L.) respond to light or whether this genetic transformation could result in unintended effects on their photosynthetic and physiological processes is not well known. Two experiments were conducted to investigate the shapes of net photosynthetic rate (P _N), stomatal conductance (g _s), apparent light use efficiency (LUE_app) and water use efficiency (WUE) light-response curves for single leaves of Bt (Bacillus thuringiensis) and Bt+CpTI (cowpea trypsin inhibitor) transgenic cotton plants and their non-transgenic counterparts, respectively. Results showed that the significant difference in response of P _N and WUE to light between transgenic cotton and non-transgenic cotton occured but not always throughout the growing season or in different experiments or for all transgenic cotton lines. It was highly dependent on growth stage, culture condition and variety, but no obvious difference between any transgenic cotton and non-transgenic cotton in the shapes of g _s and LUE_app light-response curves was observed in two experiments at different growth stages. In the field experiments, transgenic Bt+CpTI cotton was less sensitive to response of P _N to high irradiance at the boll-opening stage. In pot experiments, WUE light-response curves of both Bt transgenic cotton and Bt+CpTI transgenic cotton progressively decreased whereas non-transgenic cotton slowly reached a maximum at high irradiance at boll-opening stage. We supposed that culture environment could affect the photosynthesis of transgenic cotton both directly and indirectly through influencing either foreign genes expression or growth and physiological processes.     

Development and mapping of AFLP markers linked to the sorghum fertility restorer gene <Emphasis Type="Italic">rf4</Emphasis>

The restoration of male fertility in the sorghum IS1112 C (A3) male-sterile cytoplasm is through a two-gene gametophytic system involving complementary action of the restoring alleles Rf3 and Rf4. To develop markers suitable for mapping rf4, AFLP technology was applied to bulks of sterile and fertile individuals from a segregating BC₃F₁ population. Three AFLP markers linked to rf4 were identified and subsequently converted to STS/CAPS markers, two of which are co-dominant. Based on a population of 378 BC₁F₁ individuals, two STS/CAPS markers, LW7 and LW8, mapped to within 5.31 and 3.18 cM, respectively, of rf4, while an STS marker, LW9, was positioned 0.79 cM on the flanking side of rf4. Markers LW8 and LW9 were used to screen sorghum BAC libraries to identify the genomic region encoding rf4. A series of BAC clones shown to represent a genomic region of linkage group E were identified by the rf4-linked markers. A contig of BAC clones flanking the LW9 marker represent seed clones on linkage group E, from which fine mapping of the rf4 locus and chromosome walking can be initiated. Received: 20 June 2001 / Accepted: 3 August 2001     

DNA Marker Transmission and Linkage Analysis in Populations Derived from a Sugarcane (Saccharum spp.) x Erianthus arundinaceus Hybrid

Introgression of Erianthus arundinaceus has been the focus of several sugarcane breeding programs in the world, because the species has desirable traits such as high biomass production, vigour, ratooning ability and good resistance to environmental stresses and disease. In this study four genetic maps were constructed for two intergeneric populations. The first population (BC₁) was generated from a cross between an Erianthus/Saccharum hybrid YC96-40 and a commercial sugarcane variety CP84-1198. The second population (BC₂) was generated from a cross between YCE01-116, a progeny of the BC₁ cross and NJ57-416, a commercial sugarcane cultivar. Markers across both populations were generated using 35 AFLP and 23 SSR primer pairs. A total of 756 and 728 polymorphic markers were scored in the BC₁ and BC₂ populations, respectively. In the BC₁ population, a higher proportion of markers was derived from the Erianthus ancestor than those from the Saccharum ancestor Badila. In the BC₂ population, both the number and proportion of markers derived from Erianthus were approximately half of those in the BC₁ population. Linkage analysis led to the construction of 38, 57, 36 and 47 linkage groups (LGs) for YC96-40, CP84-1198, YCE01-116, and NJ57-416, encompassing 116, 174, 97 and 159 markers (including single dose, double dose and bi-parental markers), respectively. These LGs could be further placed into four, five, five and six homology groups (HGs), respectively, based on information from multi-allelic SSR markers and repulsion phase linkages detected between LGs. Analysis of repulsion phase linkage indicated that Erianthus behaved like a true autopolyploid.     

Detection and eradication of endophytic bacteria from micropropagated mint plants

Summary Liquid medium and an enriched agar were used to detect endophytic bacteria in micropropagated mints (Mentha spp.) within 2 to 6 d of inoculation. Bacteria isolated from the cultures were screened on several antibiotic concentrations to determine bactericidal doses. Antibiotics were also tested for phytotoxic effects. Shoot tips from infected plants were treated by immersion in liquid MS medium containing antibiotics either singly or in combination. Streptomycin applied at 1000&#x00B5;g/ml for a period of 10 d was effective and less phytotoxic in a larger number of cases than gentamicin (50&#x00B5;g/ml), neomycin (500&#x00B5;g/ml), or rifampicin (30&#x00B5;g/ml). Mint cultures that tested negative for bacteria after antibiotic treatment were multiplied, retested, and cold-stored for 1 yr or longer. Upon regrowth after storage, 25 of 30 treated cultures (83%) tested negative for bacteria. Of the 25, 8 were successfully treated with streptomycin, 1 with gentamicin, 2 with neomycin, 1 with rifampicin, and 1 with streptomycin and gentamicin; 12 required more than one treatment. An early detection system, initial trial treatment with streptomycin for infected plants, and monitoring of treated cultures successfully reduced the spread of bacterial contamination. Antibiotic treatment in liquid MS medium at pH 6.9 resulted in enhanced bactericidal activity over that seen at pH 5.5.     

Dynamic expression of green fluorescent protein and Bacillus thuringiensis Cry1Ac endotaxin in interspecific hybrids and successive backcross generations (BC1 and BC2) between transgenic Brassica napus crop and wild Brassica juncea

The persistence and stability of a transgene encoding a Bacillus thuringiensis (Bt) Cry1Ac insecticidal protein was investigated in hybrids between crop Brassica napus and a recurrent wild Brassica juncea population. Interspecific hybrids (F₁) and backcross progenies (BC₁, BC₂) containing green fluorescent protein (GFP) and Bt genes were successfully produced in the greenhouse. Stable Bt toxin levels were found in hybrid and advanced backcross progenies formed in wild B. juncea. Bt Cry1Ac concentration was significantly lower in BC₂ plants than in transgenic B. napus, F₁, BC₁, while no significant differences were detected among the latter three plant genotypes. A GFP marker gene was used as a scorable marker and indicator of Bt transgene expression. GFP fluorescence intensity was significantly correlated with Bt Cry1Ac concentration at the flowering stage and the pod formation stage in both transgenic oilseed rape hybrids and backcrossed progenies (BC₁, BC₂). It was demonstrated that GFP was a suitable marker for Bt protein in the backcross of B. juncea, which could facilitate the detection of gene flow and is useful in biosafety management.     

Photosynthesis and plant growth response of transgenic Bt cotton (<Emphasis Type="Italic">Gossypium hirsutum</Emphasis> L.) hybrids under field condition

Field experiments were conducted under rain-fed conditions to study the growth and photosynthetic efficiency of transgenic Bt cotton hybrids during 2002–03 and 2003–04 seasons. Three Bt cotton hybrids (Bollgard 1) and their non-Bt (NBt) counterparts viz. MECH 12, MECH 162, and MECH 184 were grown along with a local hybrid NHH44. Growth parameters such as plant height, main-stem nodes, biomass accumulation, and physiological processes like stomatal conductance (g _s), and rates of transpiration (E) and photosynthesis (P _N) did not differ significantly between Bt and NBt hybrids up to 80 DAS (d after sowing). Squaring commenced at 50 DAS both in Bt and NBt. The loss of young fruiting forms by the entomological factors was three times less in Bt than NBt. As a consequence, Bt had more early formed bolls on the lower canopy which contributed to higher biomass and seed cotton yield. On the other hand, bolls distributed intermittently in NBt. Heavy boll load altered the growth and physiological processes, and as a result Bt had higher g _s, E, and P _N than NBt. Since developing bolls (sink) divert the saccharides and nutrients from other organs, Bt plants with heavy boll load senesced early and stopped the production of new squares and bolls. Thus, the boll load influenced the change in growth and physiological processes of Bt from NBt.     

More nitrogen partition in structural proteins and decreased photosynthetic nitrogen-use efficiency of <Emphasis Type="Italic">Pinus massoniana</Emphasis> under in situ polluted stress

Masson pine (Pinus massoniana L.) trees in the Pearl River Delta have shown growth decline since late 1980s, particularly those around industrially polluted regions. As nitrogen is an important nutritional element composing functional proteins, structural proteins and photosynthetic machinery, investigation on nitrogen allocation is helpful to understand nutrient alteration and its regulation mechanism in response to pollution stress. Current year (C) and 1-year old needles (C + 1) of five mature trees were sampled in industrially polluted site and unpolluted natural reserve for bioassay. Needles of declining trees had significantly higher leaf nitrogen per unit area (N_L) but lower photosynthetic capacity (P _max), which resulted in lower photosynthetic nitrogen use efficiency (PNUE) than those of healthy trees. Nitrogen fraction to the photosynthetic apparatus in the C and C + 1 needles at polluted site was 27 and 22%, significantly lower than the corresponding healthy needles (48 and 32%). The content of structural proteins was positively correlated with N_L in C and C + 1 needles. Moreover, the C and C + 1 needles of declining trees had about 1.8 times structural protein as those of healthy trees, suggesting that more nitrogen allocation to structural protein are needed for stronger structural defenses under polluted stress. Decreases in PNUE of declining pine trees could be partially explained by increases in structural protein nitrogen.