A molecular mechanism of resistance to streptomycin in <Emphasis Type="Italic">Xanthomonas oryzae</Emphasis> pv. <Emphasis Type="Italic">oryzicola</Emphasis>

Xanthomonas oryzae pv. oryzicola, the causal agent of rice leaf streak disease, was found to be sensitive to streptomycin (an aminocyclitol glycoside antibiotic), by inhibition of protein synthesis resulting from interference with translational proofreading. This study aimed to determine the molecular resistance mechanism of X. oryzae pv. oryzicola to streptomycin. Seven streptomycin-resistant mutants were obtained by UV induction or streptomycin selection. These mutants can grow at 100 μg ml⁻¹ of streptomycin while the wild-type strain (RS105) cannot grow at 5 μg ml⁻¹. Sequencing indicated that the rpsL gene encoding ribosomal protein S12 has 375 bp encoding 125 amino acid residues. In all resistant strains, a mutation in which AAG was substituted for AGG (Lys→Arg) occurred either at codon 43 or 88. Two plasmids, pUFRRS and pUFRRX, were constructed by ligating the rpsL gene into the cosmid pUFR034. The plasmids pUFRRS and pUFRRX containing the Lys→Arg mutation of the rpsL gene conferred streptomycin resistance to the sensitive wild-type strain by electroporation. Both transformants, RS1 and RS2, could grow in the medium containing 50 μg ml⁻¹ of streptomycin. A mutation at codon 43 or 88 in rpsL can result in resistance of Xanthomonas oryzae pv. oryzicola to streptomycin.     

Progesterone supplementation during multiple ovulation treatment in buffalo species (<Emphasis Type="Italic">Bubalus bubalis</Emphasis>)

The aim of this study was to evaluate the effect of exogenous progesterone supplementation on superovulatory response in buffaloes that has undergone a multiple ovulation program. Fourteen Mediterranean buffaloes were divided into two groups and received a 4-day decreasing dosage of an equal mixture of 500 IU of FSH and LH starting on day 8 of the cycle. In group A (n = 7) a progesterone-releasing intravaginal device was removed on day 8, whereas in group B (n = 7) it was left till day 10, when PGF_2a {\hbox{PG}}{{\hbox{F}}_{{2}\alpha }} was administered. Eighty hours later, buffaloes were artificially inseminated and after 6 days they undergone uterine flushing. A higher (P < 0.05) number of corpora lutea (8.3 vs. 5.7) and embryo/flushing/buffalo (2.3 vs. 1.3) were recorded in group B vs. group A if responsive buffaloes are considered (n = 12) and the number of corpora lutea was highly correlated with the number of embryos (r = 0.65; P < 0.05). In conclusion, progesterone supplementation during the first 2 days of the superovulation treatment seems to enhance the recovery rate in buffalo species. A high ovulation rate, associated with a high number of corpora lutea, can represent a parameter for estimating embryo recovery.     

Temporal changes in endogenous estrogens and expression of behaviors associated with estrus during the periovulaory period in Murrah buffaloes (<Emphasis Type="Italic">Bubalus bubalis</Emphasis>)

The objective of this study were (1) to establish the duration of behavioral estrus signs and timing of ovulation in Murrah buffaloes (n = 10) and (2) to determine relationship between behavioral estrus signs with change in plasma estrogen concentrations. Estrus and its behavioral signs were detected at hourly intervals by visual observations, per recta examination of genitalia and bull parading four times in a day for 30 minutes each. Among the behavioral signs of estrus, swollen vulva (80%) was the best indicator of estrus followed by excitement (70%). Among the duration of behavioral estrus signs the first and longest duration of estrus signs was swollen vulva which was seen upto 19.8 ± 0.8 h after onset of estrus. The mean total duration of estrus symptoms from appearance to disappearance of all the behavioral estrus symptoms was 23.5 ± 1.7 h. All the behavioral estrus symptoms were observed during the period of estrogen surge. Endocrine profile during the periestrus period showed that the mean peak concentrations of total estrogen 23.9 ± 3.9 pg/ml occurred at 8.8 ± 1.7 h after onset of estrus. The average number of estrus symptoms observed per animal during onset of spontaneous estrus was 5.7. Ovulation occurred after 37.4 ± 1.7 h after onset of estrus and 13.4 ± 1.0 h after end of total estrogen surge respectively. In conclusion, our results suggest that all signs of behavioral estrus occurred during the preovulatory rise in estrogens. The first sign of estrus to be observed was a swollen vulva and this symptom persisted the longest.     

AlgU contributes to the virulence of <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> pv. <Emphasis Type="Italic">tomato</Emphasis> DC3000 by regulating production of the phytotoxin coronatine

Pseudomonas syringae pv. tomato DC3000 (Pst DC3000), which causes bacterial speck disease of tomato, has been used as a model pathogen to investigate the molecular basis of plant–pathogen interactions. The function of many potential virulence factors encoded in the Pst DC3000 genome and their modes of action are not fully understood. P. syringae is known to produce the exopolysaccharide alginate. Although AlgU, a sigma factor, is known to regulate the expression of genes such as algD related to alginate biosynthesis, the molecular mechanisms of AlgU in the virulence of Pst DC3000 is still unclear. To investigate the function of AlgU and alginate in plant–bacterial pathogen interactions, we generated ΔalgU and ΔalgD mutants. After inoculation with ΔalgU but not ΔalgD, host plants of Pst DC3000 including tomato and Arabidopsis had milder disease symptoms and reduced bacterial populations. Expression profiles of Pst DC3000 genes revealed that AlgU can regulate not only the expression of genes encoding alginate biosynthesis, but also the expression of genes related to type III effectors and the phytotoxin coronatine (COR). We also demonstrated that the ΔalgU mutant showed full virulence in the Arabidopsis fls2 efr1 double mutant, which is compromised in the recognition of PAMPs. Further, the application of COR was able to restore the phenotype of the ΔalgU mutant in the stomatal response. These results suggest that AlgU has an important role in the virulence of Pst DC3000 by regulating COR production.     

Effect of <Emphasis Type="Italic">Metarhizium guizhouense</Emphasis> infection on mating competition and mate choice of <Emphasis Type="Italic">Bactrocera latifrons</Emphasis> (Diptera: Tephritidae)

Metarhizium guizhouense PSUM02 treated males of Bactrocera latifrons were investigated for the mating competition among males and mating choice by female flies to develop an auto-dissemination for the control of B. latifrons. In the present study, on day 1–4 of experiment, M. guizhouense–treated male flies were equally competitive with the normal male flies as we did not observe any differences in mating by treated and normal male flies of B. latifrons. Further, mating competitiveness were found low in treated adult male B. latifrons than normal male B. latifrons from 5th days of treatment until death. Kaplan-Meier survival analysis of treated male flies gave average survival times (AST) of 4.3?±?0.1 days, while the healthy female and male flies in the same cage showed AST of 9.3?±?0.3 and 8.3?±?0.4 days, respectively. The AST of untreated flies in control experiment ranged from 14.2–14.5 days. In mating preference experiment, M. guizhouense–treated male flies were chosen by virgin female than gravid female flies for mating. The treated male flies caused mortality in both virgin and gravid female flies in the same cage with AST of 4.4?±?0.1, 5.6?±?0.1 and 7.4?±?0.2 days, respectively, while untreated flies showed AST ranged from 13.9–14.3 days in control. The treated male flies could transmit the fungus infection to both untreated female and male flies as well as in virgin and gravid female flies by mating and contact. Our experiments showed the potentiality of M. guizhouense PSUM02 in management of B. latifrons by auto-dissemination with treated male flies, which transmit the fungus to a healthy population to reduce insect pest infestations.     

Feeding potential of <Emphasis Type="Italic">Cryptolaemus montrouzieri</Emphasis> against the mealybug <Emphasis Type="Italic">Phenacoccus solenopsis</Emphasis>

Phenacoccus solenopsis Tinsley (Hemiptera: Pseudococcidae) is an exotic species native to the USA, damaging cotton and other plant families. The feeding potential of different development stages of Cryptolaemus montrouzieri Mulsant, a biological control agent against mealybugs, was investigated on different development stages of P. solenopsis. Fourth instar grubs and adults of C. montrouzieri were the most voracious feeders on different instars of mealybug. The number of 1^st instar nymphs of mealybug consumed by 1^st, 2^nd, 3^rd and 4^th instar larvae and adult beetles of C. montrouzieri was 15.56, 41.01, 125.38, 162.69 and 1613.81, respectively. The respective numbers of 2^nd and 3^rd instar nymphs of mealybug consumed were 11.15 and 1.80, 26.35 and 6.36, 73.66 and 13.32, 76.04 and 21.16, 787.95 and 114.66. The corresponding figures for adult female mealybugs were 0.94, 3.23, 8.47, 12.71 and 73.40, respectively. The results indicate that C. montrouzieri has the potential to be exploited as a biocontrol agent in North India; inoculative releases of 4^th instar larvae and adults may provide instant control of P. solenopsis. Field experiments should be conducted to determine the efficiency of the ladybird on this mealybug.     

Advanced lentil lines screened for resistance to <Emphasis Type="BoldItalic">Fusarium oxysporum</Emphasis> f. sp. <Emphasis Type="BoldItalic">lentis</Emphasis> under greenhouse and field conditions

Fusarium oxysporum f. sp. lentis is the most important pathogen of lentil plants, and most areas under lentil cultivation are reported to have a fusarium wilt disease background. The plants are infected in the seedling stage and later stages of their development. Fusarium wilt disease, which has appeared at high incidence rates during recent years, has caused sharp drops in the yield, especially in Moghan, in the northwest of Iran. Forty-five isolates of the pathogen were collected from different regions of the country with two isolates from ICARDA in the summer of 2008 and identified using Nelson’s key. The pathogenicity of the collected isolates was studied on a sensitive line (ILL 4605) under greenhouse conditions and significant differences in pathogenicity were found among them. The most pathogenic isolates from three provinces, East Azerbaijan (EA 30), Ardebil (Ar 3) and Khorasan (Kh 45), were selected and used in screening of 55 developed lines under greenhouse and field conditions. In the greenhouse, test plants were inoculated by immersing root tips in spore suspension and sowing seeds in pre-infested pot soil. Field tests were carried out in a naturally highly infested farm. At all stages, the plant response to the disease was based on the percentage of dead plants. Cluster analyses of the greenhouse and field data led to the selection of three lines (81S15, FLIP2007-42 L and FLIP2009-18 L) that were resistant under greenhouse and field conditions.     

Pathogenicity and mycotoxin production by <Emphasis Type="Italic">Fusarium proliferatum</Emphasis> isolated from onion and garlic in Serbia

Fusarium proliferatum can occur on a wide range of economically important vegetable plants but its role in disease is not always well established. In 2000 and 2001, from forty-one field samples of wilting onion and garlic plants in Serbia, F. proliferatum as the predominant fungal species was isolated from root and bulbs. Seventy isolates were firstly characterized for their sexual fertility and were shown to be mostly members of Gibberella intermedia (sixty-seven of seventy isolates, the remaining three isolates were unfertile), the sexual stage of F. proliferatum (syn. mating population D of G. fujikuroi complex). A selected set of eleven F. proliferatum isolates from both hosts were also tested for their pathogenicity and toxigenicity. Although onion and garlic plants were susceptible to all isolates, onion plants showed a significantly higher disease severity index. Six of the eleven isolates of F. proliferatum produced fumonisin B₁ from 25 to 3000 μg g⁻¹, and beauvericin from 400 to 550 μg g⁻¹; ten isolates produced fusaric acid from 80 to 950 μg g⁻¹ and moniliformin from 50 to 520 μg g⁻¹. Finally, all isolates produced fusaproliferin up to 400 μg g⁻¹. These results confirm F. proliferatum as an important pathogen of garlic and onion in Europe and that there is a potential mycotoxin accumulation risk in contaminated plants of both garlic and onion.