Jerzy H. Czembor, Henryk J. Czembor, Rich ard Pickering,
Plant Breed ing and Ge net ics De part ment, Plant Breed ing and Ac cli ma ti za tion In sti tute, Radzików, 05-870 B³onie, Po land.
LEAF RUST RESISTANCE IN HYBRID LINES DERIVED FROM CROSSESS BETWEEN HORDEUM VULGARE
AND HORDEUM BULBOSUM
ABSTRACT
Leaf rust caused by fun gus Puccinia hordei has great eco nomic im por tance in many bar ley grow ing re gions in Eu rope, North Amer ica, Aus tra lia and West Asia and North Af rica. Bul bous bar ley grass (Hordeum bulbosum L.), is the only mem ber of the sec ond ary bar ley genepool. In pre sented study 6 re com bi nant lines ob tained from back -cross es of bar ley cultivars (back -cross ing par ents) and ac ces sions of H. bulbosum were tested with 8 dif fer en tial iso lates of leaf rust. This study showed that re sis tance to leaf rust is pres ent in 5 from to tal 6 re com bi nant lines. Out stand ing re sis tance to leaf rust was iden ti fied in line 886Z3/1/10/1/2/1, which showed re sis tance re ac tion 0 for in oc u la tion with all iso lates used. An other 4 lines were sus cep ti ble for in oc u la tion with 2 or 4 leaf rust iso lates used. In none of tested lines was pos si ble to pos tu late pres ence of known re sis tance genes for leaf rust re sis tance. Based on re sults it may be con cluded that leaf rust re sis tance iden ti fied in re com bi nant lines co mes from H.
bulbosum and may rep re sent new unique type of re sis tance. Hy brid lines with iden ti fied re sis tance to leaf rust orig
i nat ing from H. bulbosum should be used in breed ing programmes to pro vide farm ers with cultivars with highly ef -fec tive re sis tance to this dis ease.
Key words: Hordeum bulbosum, leaf rust, Puccinia hordei, re com bi nant lines, re sis tance genes
INTRODUCTION
Bar ley (Hordeum vulgare L.) is the fourth most im por tant ce real crop in the world. Bar ley breed ers and ge net i cists are tak ing into ac count three genepools in bar ley. H. spontaneum and H. vulgare are in the pri mary genepool (Bothmer et al. 1995). Wild bar ley es pe cially H. spontaneum was used in breed ing programmes to trans fer of new dis ease resistances and tol -er ance to abiotic stress (Fischbeck 2003; Pick-ering and Johnston 2005). Bul bous bar ley grass (H. bulbosum L.), is the only mem ber of the sec ond ary genepool (Pickering et al. 2004a; Bothmer et al. 2003; Pickering and Johnston 2005). It is pe ren nial and oc curs in the Med i ter ra nean re gion, West Asia, Cau ca sus Moun tains and part of Cen tral Asia in clud ing Iran, Afganistan, Turkmenistan, Uzbekistan, Kazakhstan. It oc curs as both dip -Communicated by Ed ward Arseniuk
P L A N T B R E E D I N G A N D S E E D S C I E N C E
Volume 57 2008
loid and autotetraploid cytotypes. It nor mally re quires vernalisation to flower and has a strong self-in com pat i bil ity sys tem based on two loci (Bothmer et al. 1995).
Leaf rust caused by fun gus Puccinia hordei has great eco nomic im por -tance in many bar ley grow ing re gions in Eu rope, North Amer ica, Aus tra lia and West Asia and North Af rica (WANA) (Yahyaoui and Sharp 1987; Park et al. 1992; Park 2003; Woldeab et al. 2006). In many cases more im por tant than low er ing of bar ley yield is loss of its qual ity due to in fec tion by leaf rust. This es pe cially con cerns plan ta tions of bar ley for malt ing pur pose. Re -cently the breed ers in ter est in re sis tance to bar ley leaf rust has in creased in Eu rope (Mazaraki and Grabowska 1998; Niks et al. 2000; Czembor et al. 2006). This in ter est is caused by ob ser va tions of in creases in fit ness of leaf rust pop u la tions to many cur rently grown bar ley cultivars and to cultivar Vada. Cultivar Vada is well known for pos sess ing high level of par tial re sis -tance and it was used wildly as par ent in ma jor Eu ro pean bar ley breed ing pro grams (Czembor et al. 2006).
For many years the use of dis easere sis tant bar ley cultivars has been an ef -fi cient means for con trol ling ma jor dis eases and pre vent ing yield losses (Czembor 1996, 2005; Brooks et al. 2000; Finckh et al. 2000; Fischbeck 2003; Weibull et al. 2003). How ever, bar ley breed ers, ge net i cists and plant pa thol o gists are con stantly look ing for new ef fi cient sources of re sis tance to ma jor dis eases in clud ing leaf rust to com bine them with al ready used in mod ern cultivars in or der to in crease the re sis tance du ra bil ity (Czembor 1996, 2005; Brown and Hovmoller 2002; Bonman et al. 2005).
In sev eral stud ies H. bulbosum was de scribed as spe cies with very high level of re sis tance to bar ley patho gens in clud ing leaf rust (Pickering et al. 2004a; Pickering and Johnston 2005). De spite of these ob ser va tions, there are only few re ports on ge netic in ves ti ga tions on H. bulbosum and on trans -fer of re sis tance to ma jor patho gens from H. bulbosum to H. vulgare (Pickering et al. 2004b, 2006a; Zhang et al. 2001; Pickering and Johnston 2005). In these re ports hy brid lines of H. bulbosum x H. vulgare ex pressed re sis tance to such dis eases as leaf rust, pow dery mil dew, scald, septoria specled leaf blotch, BaYMV/BaMMV, stem rust (Pickering et al., 2004a, 2006b; Shtaya 2007).
In bar leyleaf rust hostpatho gen sys tem 19 loci with ma jor genes for re -sis tance were iden ti fied (Park and Karakou-sis 2002, Steffenson 2002, Park et al. 2003, Weerasena et al. 2004). Most of these re sis tant loci were iden ti fied in old cultivars or land races of H. vulgare (Czembor et al. 2006). How -ever, three loci Rph10, Rph11 and Rph16 were in tro duced to bar ley from H. spontaneum. In ad di tion, based on re ports of Pickering and co-work ers two re sis tance loci Rph17, Rph18 were de scribed which orig i nate from H. bulbosum (Pickering et al. 1998, Franckowiak 2000).
The ob jec tive of this study was to de ter mine leaf rust re sis tance in re com -bi nant lines ob tained in crosses be tween H. bulbosum and H. vulgare.
MATERIALS AND METHODS
Plant ma te rial
Six re com bi nant lines ob tained from crosses and back cross es of bar ley cultivars Vada, Emir, Golden Prom ise (back cross ing par ents) and ac ces sions of H. bulbosum (S1, 2920/4, HB2032) were tested (Ta ble 1). In ad di tion 3 cultivars which were back cross ing par ents for spe cific lines were tested. Re -com bi nant lines were ob tained at New Zea land In sti tute for Crop and Food Re search, New Zea land (Pickering 1987, 1988; Pickering et al. 1987, 1998, 2000a).
Patho gen
Eight dif fer en tial iso lates of P. hordei were used (Ta ble 2). These iso lates orig i nated from IHAR Radzikow col lec tion and were cho sen ac cord ing to dif fer ences in vir u lence spec tra ob served on 12 dif fer en tial cultivars. None of the iso lates used was able to dif fer en ti ate genes Rph4 from Rph8 and Rph1 from Rph10 and Rph11.
Re sis tance tests
Five to 10 plants per line were eval u ated in a green house with 8 iso lates of P. hordei. This study was car ried out in the IHAR Radzikow grow ing cham -bers. Cultivar L94, which does not carry any known genes for re sis tance to P. hordei, was used as a sus cep ti ble con trol.
Test ing pro ce dure
The plants were grown with 16 h light and tem per a ture range of 18-22°C. Urediniospores of P. hordei were sus pended in deionized wa ter with cou ple
Ta ble 1
Re com bi nant lines, their ped i grees and chro mo some lo ca tion of H. bulbosum introgression
No Recombinant line Back crossing parent(H. vulgare) H. bulbosumparent H. bulbosum introgressionChromosome location of
1 81882/83/3/2/9 Vada S1 2HS
2 4176/n/3/2/6 Vada S1 No data
3 886Z3/1/10/1/2/1 Emir 2920/4 2HL
4 38P18/5-13/1-9 Emir HB2032 2HL
5 38U4/1/3/7 Golden Promise 2920/4 5HL + 6HS
drops of “Tween 20" and in oc u lated onto oneweek old seed ling plants (pri mary leaf fully ex panded) us ing a rate 3 mg urediniospores and 10 ml of wa -ter- for 100 plants. In oc u lated plants were in cu bated for 24 hours in a cham ber in which the hu mid ity was main tained near sat u ra tion by mist from ul tra sonic hu mid i fi ers n com plete dark ness and with a tem per a ture range of 12-15oC. Then plants were trans ferred to a green house bench.
Dis ease as sess ment
Re ac tions of each ac ces sion were eval u ated af ter an in cu ba tion pe riod of 12-14 days in a green house at 20-24°C. Dis ease symp toms were as sessed on the pri mary leaf of the seed lings ac cord ing to 0-4 scale adapted from Le vine and Cherewick (1952). In fec tion types 0, 0;, 1 and 2 were con sid ered in dic a -tive of in com pat i bil ity whereas in fec tion types 3 and 4 of com pat i bil ity.
Pos tu la tion of leaf-rust re sis tance genes
Hy poth e ses about the spe cific re sis tance genes pres ent were made by com par ing the re ac tion spec tra of the tested lines with those of dif fer en tial lines.
Ta ble 2
Dif fer en tial iso lates and their in fec tion types on dif fer en tial set.
Accession name Accessionnumber Gene
Isolates Ph- 9 Ph-5 Ph-4 Ph-6 Ph-31 Ph-21 Ph-17 Ph-25 Sudan CIho 6489 Rph1 4 4 4 4 4 4 4 4 Peruwian CI 935 Rph2 4 4 4 4 4 4 2 4 Estate CI 3410 Rph3 0 4 0 4 4 0 4 4 Gold CI 1145 Rph4 4 4 0 4 4 4 4 4 Magnif CI 13860 Rph2+Rph5 4 1 4 0 0 0 1 4 Bolivia CI 1257 Rph2+Rph6 4 4 4 4 0 4 4 4 Cebada Capa CI 6193 Rph7 0; 0; 0; 0; 0; 0; 0; 0; Egypt 4 CI 6481 Rph8 4 4 0 4 4 4 4 4 HOR 2596 CI 1243 Rph9 4 4 4 4 4 1 4 4 Cliper C8 None Rph10 4 4 4 4 4 4 4 4 Cliper C67 None Rph11 4 4 4 4 4 4 4 4 Triumph PI 290195 Rph12 4 4 4 4 4 0; 4 4
RESULTS
Line 4176/n/3/2/6 was sus cep ti ble for in fec tion with all iso lates used. Most prob a bly this line do not pos sess re sis tance to leaf rust (Ta ble 3). An other 5 lines tested showed re sis tance re ac tion to in fec tion with leaf rust. The most re sis tant line for in oc u la tion with leaf rust was line 886Z3/1/10/1/2/1. This line showed re sis tance re ac tion 0 for in oc u la tion with all iso lates used. An -other 4 lines were sus cep ti ble for in oc u la tion with 2 or 4 leaf rust iso lates used. The most com mon re sis tance re ac tion was 0. In none of tested lines was pos si ble to pos tu late pres ence of known re sis tance genes for leaf rust re sis -tance.
Bar ley cultivars used as back cross ing par ents showed lack of re sis tance or very lim ited one for in oc u la tion with 8 dif fer en tial iso lates.
DISCUSSION
Pre sented study con firmed find ings of other in ves ti ga tors that hy brid lines of H. bulbosum x H. vulgare pos sess re sis tance to ma jor patho gens of bar ley in clud ing leaf rust (Pickering et al. 1999, 2004b; 2006b; Pickering and Johnston 2005; Shtaya 2007). Based on screen ing tests it may be con -cluded that re sis tance to leaf rust is pres ent in 5 from to tal 6 re com bi nant lines ob tained from crosses and back cross es of bar ley cultivars and ac ces -sions of H. bulbosum. Out stand ing re sis tance to leaf rust was iden ti fied in line 886Z3/1/10/1/2/1, which showed re sis tance re ac tion 0 for in oc u la tion with all iso lates used. An other 4 lines were sus cep ti ble for in oc u la tion with
Ta ble 3
Re ac tion of 6 re com bi nant lines and 3 cultivars to in fec tion with 8 iso lates of Puccinia hordei.
No lines and cultivarsRecombinant
Isolates Postulated
genes Possible genes 9 5 4 6 31 21 17 25 1 81882/83/2/9 4 4 4 1 0; 1 4 2 ?* 2 4176/n/3/2/6 4 4 4 4 4 4 4 4 None Rph1 or Rph10 or Rph11 3 886Z3/1/10/1/2/1 0 0 0 0 0 0 0 0 ? 4 38P18/5-13/1-9 0 0 0 0 4 0 4 0 ? 5 38U4/1/3/7 4 4 4 2 4 2 2 2 ? 6 102C2/18 0; 0 0 4 4 0; 0 0 ?
7 Golden Promise 4 4 4 4 4 4 4 4 None Rph1 or Rph10 or Rph11
8 Emir 4 4 4 4 4 2 4 4 ? Rph9
2 or 4 leaf rust iso lates used. In none of tested lines was pos si ble to pos tu late pres ence of known re sis tance genes for leaf rust re sis tance. How ever based on re sis tance re ac tion we can con clude that line 81882/83/2/9 may have more than one re sis tance gene be cause it ex pressed 3 dif fer ent re sis tance re -ac tions: 0;, 1 and 2. Lines 886Z3/1/10/1/2/1 and 38P18/5-13/1-9 may have one or more re sis tance genes ex pressed as re sis tance re ac tion 0. In line 38U4/1/3/7 we can pos tu late pres ence of one or more re sis tance genes ex pressed as re sis tance re ac tion 2. In ad di tion, ex pres sion of re sis tance re ac -tion 2 is show ing also pos si bil ity for the pres ence of some level of par tial re sis tance in this line. Bar ley cultivars used as back cross ing par ents showed lack of re sis tance or very lim ited one for in oc u la tion with 8 dif fer en tial iso -lates. It con firms that re sis tance loci pres ent in tested re com bi nant lines orig i nate from H. bulbosum par ents.
In pre sented study seed ling re sis tance tests were used in or der to de scribe in fec tion types ex pressed by bar ley plants af ter in oc u la tion with dif fer en tial iso lates of P. hordei. This kind of test ing as suf fi cient for screen ing for dis ease re sis tance and it is used com monly in many breed ing pro grams to pos -tu late the pres ence of spe cific genes for re sis tance in mod ern cultivars and to screen for new sources of ef fec tive re sis tance (Czembor 1996, 2005; Czembor and Czembor 2001). How ever, by us ing this kind of tests it is not pos si ble or at least it is dif fi cult to iden tify and de scribe par tial re sis tance. For de scrip tion of this kind of re sis tance we need to con duct ad di tional to in fec tion type of mea sure ments of char ac ter is tics. In ad di tion, par tial re sis -tance is gen er ally better ex pressed at the adult plant stage (Ochoa and Parlevliet 2007). It will be very in ter est ing if fur ther stud ies of de scribed hy brid lines will also in clude pa ram e ters de scrib ing par tial re sis tance and ex -ten sion of re sis tance stud ies to plants at adult stage. Fi nal de ter mi na tion of the num ber of re sis tance genes and the type of their ac tion in tested hy brid lines may be es tab lished by crosses and back cross es among ap pro pri ate ge -no types (Czembor 1996, 2005).
De scrip tion of new sources of re sis tance are im por tant be cause fu ture strat e gies for the con trol of bar ley patho gens will have to fo cus in creas ingly on more eco log i cally ac cept able meth ods. Any us age of chem i cals (pes ti cides, fun gi cides, her bi cides, and min eral fer til iz ers) in ag ri cul ture is in -creas ingly crit i cized in so ci et ies of many coun tries. Breed ing for re sis tance rep re sent such eco log i cally safe method. In ad di tion to eco log i cal also eco -nom i cal ar gu ments (use of fuel, la bour, spe cial ma chines) are in fa vour of breed ing for re sis tance ver sus chem i cal con trol (Nierobca et al. 2003). Hy -brid lines with iden ti fied re sis tance to leaf rust orig i nat ing from H. bulbosum should be used in breed ing programmes to pro vide farm ers with cultivars with highly ef fec tive re sis tance to this dis ease.
ACKNOWLEDGEMENT
Au thors thank Dr Rich ard Pickering (New Zea land In sti tute for Crop and Food Re search Lim ited, Pri vate Bag 4704, Christchurch, New Zea land) for seed sam ples of re com bi nant lines tested in this study.
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