561 by xiangpeng


									                                                                                         SCIENTIFIC CORRESPONDENCE

Management of nursery wilt of black pepper (Piper nigrum L.)
with antagonistic bacteria

Black pepper is an important agricultural        and host plant, which resembled the field         015, PN-026, PN-032 and PN-033, were
produce of Kerala, that contributes sub-         conditions in a better way than the dual          selected for the in vivo biological control
stantially to the foreign exchange earning       culture plate method, where there is no           assay, based on their differential antago-
of India. Black pepper cultivation in            involvement of the host plant. Disease-           nism on the dual culture assay and the
India has been under threat due to severe        free, single-node cutting (approximately          shoot lesion assay.
infestation of foot-rot disease incited by       8 cm length) obtained from runner shoots             Planting medium consisting of sieved,
Phytophthora capsici, which has been             of black pepper cv. Panniyoor-1 were              autoclaved sand and soil (pH 6.2) in the
spreading at an alarming rate. Infection         surface-sterilized with 0.1% sodium hypo-         ratio of 1 : 1 was filled in plastic pots
in the nursery stage results either in the       chlorite solution for 10 min, rinsed with         (11 cm dia) and two-node runner shoot cut-
wilting of the cuttings, i.e. the planting       sterile distilled water and spread in a           tings collected from a garden at the
material or rotting of the newly emerging        laminar airflow chamber. Bacterial cells          Regional Agricultural Research Station
leaves. The disease is often carried to the      were harvested from 48-h-old KMB                  (RARS), Wayanad were planted after
main field through the infected rooted           plates by drenching 10 ml sterile distilled       surface sterilization, as described earlier.
cutting and the rooting medium.                  water and scrapping with a glass spreader.        Antagonists were applied to the cuttings
   Biological control, especially that involv-   Stem cuttings were completely dipped in           by dipping the 5 cm basal portion in bac-
ing Pseudomonas spp., has emerged as             the bacterial suspension for 30 min and           terial suspension for 30 min. The fungal
an important alternative in managing soil-       spread in a laminar airflow chamber for           pathogen P. capsici was artificially inocu-
borne plant diseases in recent years1–3.         another 30 min for drying. A pinprick             lated into the planting hole prior to plant-
However, very little work has been done          was given on the middle portion of the            ing, by applying 2 ml zoospore suspen-
on biological control of diseases in vege-       cuttings using a sterile needle, and the          sion prepared in sterile distilled water
tatively propagated crops, except potato4,5.     pricked area inoculated with a small myce-        (approximately 105 propagules/ml) from
Trichoderma harzianum and Tricho-                lial bit of P. capsici collected from a           ten-day-old culture grown on CA plates.
derma viride have been used as biologi-          seven-day-old culture on PDA medium.              Three replications with 15 plants each
cal control agents against P. capsici            The inoculated cuttings were kept in              were maintained in a glasshouse (25 ±
attacking black pepper, both in the main         150 mm dia petri dishes. Moist filter             2°C). Irrigation was given daily with ster-
field and nursery6. Only limited attempts        paper was kept on the inner side of the           ile distilled water. P. capsici-inoculated
have been made on the biological control         lid to provide high humidity, and the             control, uninoculated healthy control and
of foot-rot of black pepper using bacte-         plates were incubated at 25°C under dark-         chemical control (0.2% copper oxy chlo-
rial antagonists7. We report here a rapid        ness. Length of black lesion developed            ride (COC) drenching at 15 days inter-
screening assay for the selection of effi-       around the inoculated region on the stem          val) were maintained for comparison.
cient bacterial antagonists which can            cuttings due to the fungal invasion was              Disease incidence in the in vivo condi-
colonize and protect the planting mate-          recorded at 24 h intervals for a period of        tion was recorded by counting the wilted
rial against P. capsici-induced wilt of          96 h (Figure 1). Inoculation with P. cap-
black pepper in the nursery.                     sici alone and pinprick alone served as
   Fluorescent pseudomonads were iso-            controls. All the isolates showing antago-
lated on King’s medium B (KMB) from              nism on both CA and PDA were tested
the underground shoot portions of three-         for suppression of lesion development.
month-old healthy rooted cuttings of                Bacterial isolates differed in their ability
black pepper cv. Panniyoor-1 raised in           to suppress the lesion development in the
the nursery. Virulent black pepper strain        assay. Development of lesion was moni-
of P. capsici was isolated from the              tored for four days and a conducive envi-
infected nursery plants and maintained           ronment for the fungal infection, such as
on either carrot agar (CA) or potato dex-        high humidity and darkness was provi-
trose agar (PDA) slants at 25°C.                 ded. However, in a few treatments lesion
   All the bacterial isolates were tested        development was less or was delayed.
against P. capsici on PDA and CA by              This could be attributed to the ability of
dual culture technique, and inhibition of        the bacterial isolates to survive on the
fungal growth was noted after five to seven      stem, at least for a limited period in the
days7. Among the 64 different isolates           test conditions, and thereby preventing
tested, 11 were found to have antagonism         the disease development. Black pepper
on both CA and PDA with varying                  being a vegetatively propagated crop,
degrees, as evidenced by difference in           any biocontrol agent that is used for sup-
the inhibition zone. A screening on the          pressing the soil-borne infection of the          Figure 1. Suppression of lesion on black
                                                                                                   pepper shoots during screening of bacterial
stem cuttings was done further. This             planting material by P. capsici should            antagonists P. capsici after 48 h of inocu-
assay was performed on the basis of the          have the ability to survive on the stem           lation. PN-026, PN-015, PN-033, PN-032,
interaction of the pathogen, antagonist          cuttings. Four bacterial antagonists, PN-         P. capsici control and pinprick control.

CURRENT SCIENCE, VOL. 83, NO. 5, 10 SEPTEMBER 2002                                                                                         561
                  Table 1.   Percentage wilt of black pepper cuttings in the nursery                   7. Jubina, P. A. and Girija, V. K., J. Mycol.
                                                                                                          Plant Pathol., 1998, 28, 147–153.
                                         Percentage wilt of black pepper cuttings

              Treatment                        60 DAP                 90 DAP                           Received 22 October 2001; revised accepted
                                                                                                       18 July 2002
              PN-026                        40.60 (39.44)          45.00 (41.92)
              Pathogen control              95.83 (79.78)          97.92 (83.99)
              Healthy control                6.25 (14.00)           6.25 (14.00)
              0.2% COC drenching            27.08 (30.83)          27.08 (30.83)
                                                                                                                                  K. N. ANITH†,#,*
              CD (P = 0.05)                      6.15                   6.02
                                                                                                                          N. V. RADHAKRISHNAN‡
               Mean of three replications having fifteen cuttings each; bFigures in                                       T. P. MANOMOHANDAS‡
              parenthesis are arc-sine transformed values.

                                                                                                         Department of Plant Pathology,
cuttings. The isolate PN-026, which                 borne infection by P. capsici in black             College of Agriculture,
showed maximum suppression of lesion                pepper nursery, as it gives a direct rela-         Kerala Agricultural University,
development in the screening (8.22 mm               tionship between the lesion suppression            Vellayani,
compared to 59.22 mm in the pathogen                and in vivo biological control activity.           Thiruvananthapuram 695 522, India
control after 96 h of incubation), reduced                                                               Regional Agricultural Research Station,
the incidence of the disease significantly                                                             Kerala Agricultural University,
compared to other bacterial antagonists              1. Bowen, G. D. and Rovira, A. D., Adv.           Ambalavayal,
(Table 1). In general, isolates giving                  Agron., 1999, 66, 1–120.                       Wayanad 673 593, India
                                                     2. Weller, D. M., Annu. Rev. Phytopathol.,        #
greater suppression of lesion develop-                                                                   Present address:
ment in the shoot assay, irrespective of                1998, 26, 379–407.                             North Florida Research and
their antagonism on the dual culture plate           3. Whipps, J. M., Adv. Bot. Res., 1997, 26,          Education Center,
assay, showed higher degree of biocon-                                                                 Institute of Food and Agricultural Sciences,
                                                     4. Chambers, A. M. and Scott, E. S., J. Phy-
trol efficiency. Our study suggests that                topathol., 1995, 143, 471–477.
                                                                                                       University of Florida,
the screening procedure described in the             5. Schippers, B., Philos. Trans. R. Soc. Lon-     155 Research Road,
present investigation can be used as a rapid            don, 1998, 318, 283–293.                       Quincy, FL 32351, USA
and more accurate technique for select-              6. Anandaraj, M. and Sarma, Y. R., J. Spices      *For correspondence
ing bacterial antagonists against soil-                 Aromat. Crops, 1995, 4, 17–23.                 e-mail: knanith@yahoo.com

Characterization of polyhydroxy alkanoates – Biodegradable
plastics from marine bacteria
A wide variety of bacteria accumulate               duct with desirable properties for com-            produced by these organisms are pre-
polyhydroxy alkanoates (PHAs) as intra-             mercial applications1. Polymers may fail           sented here.
cellular storage material1–4. Because of            in specific applications, simply because              The selected isolates were grown
their physical and structural properties            they do not possess the necessary strength         routinely in 50 ml of E2 mineral medium12
and amenability to biodegradation, PHAs             to carry the designed load or occasional           consisting of NaNH4HPO4 ⋅ 4H2O, 3.5 g;
are considered potential substitutes for            overload10. Hence, it is important to              K2HPO4 ⋅ 3H2O, 7.5 g; KH2PO4, 3.7 g per
petrochemical plastics. PHAs vary in their          study the mechanical and physical pro-             litre; MgSO4 ⋅ 7H2O, 0.17 g, and micro-
mechanical properties depending on the              perties of such commercially important             elements stock solution, 1 ml contain-
composition of the monomeric units5.                polymers before their use in the industry.         ing FeSO4 ⋅ 7H2O, 2.78 mg; MnCl4 ⋅ 4H2O,
The medium chain-length PHAs are semi-                 Tropical mangrove and marine ecosys-            1.98 mg; CoSO4⋅7H2O, 2.81 mg; CaCl2⋅
crystalline elastomers with a low melting           tems from the mid-west coast of India              2H2O, 1.47 mg; CuCl2 ⋅ 2H2O, 0.17 mg,
point (Tm), low tensile strength and high           were screened for promising bacteria,              and ZnSO4 ⋅ 7H2O, 0.29 mg supplemented
elongation to break6,7, and can be used             with capability of accumulating high               with yeast extract 0.04% (w/v), glucose
as biodegradable rubber. Polyhydroxy                amounts of PHA11. The isolates desig-              2% (w/v), for 48 h on an Orbitek shaker
butyrate (PHB), smallest known PHA                  nated as 61/4, 64/4, 87/4, 182/5, 12/BL,           at 28°C and 150 rpm. The cells were
displays a similar degree of crystallinity          85/6 and 86/6, which accumulated more              washed with saline by centrifugation.
and Tm as polystyrene8, and is stiffer and          than one gram PHA per litre culture broth          The PHA extracted from the cell pellet
more brittle than polypropylene8,9; but its         were studied for physico-chemical fac-             by the hypochlorite method13, was washed
copolymerization with hydroxy valerate              tors influencing quantitative yield of             with methanol and acetone consecutively
(HV) monomer units reduces its stiffness            PHA (unpublished results). The physical            and centrifuged at 8000 rpm for 20 min.
and increases its toughness, giving a pro-          and mechanical properties of the PHA               The polymers were then dissolved in hot

562                                                                                       CURRENT SCIENCE, VOL. 83, NO. 5, 10 SEPTEMBER 2002

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