Evaluation of Ice Nucleation Activity (INA) and INA Gene Detection in the Bacteria Isolated from Pistachio Trees in Kerman Province, Iran

Abstract

IIce nucleation active (INA) bacteria are common epiphytic inhabitants that cause frost damage in many plants in the near-zero temperatures. Yet, no studies were found in ice nucleation bacteria associated with pistachio trees. In our earlier study some INA strains were identified and reported. These were assigned as <em>Pseudomonas fragi</em>,<em> P. putida</em>,<em> P. moraviensis </em>and<em> Pantoea agglomerans</em>. In current work, two new strains namely<em> P. viridiflava </em>and<em> Entrobacter cloacea</em> were identified. Their ice nucleation frequency were evaluated and compared with above-mentioned ice positive strains isolated from pistachio trees.<em> Pseudomonas fragi</em> raf₃ was considered as the most ice nucleation active bacteria. This was followed by <em>P. putida </em>raf₆,<em> P. moraviensis </em>raf₁, <em>P. moraviensis </em>raf₅,<em> Pantoea agglomeranse </em>raf₇,<em> P. viridiflava </em>raf_2,<em>Entrobacter cloacea </em>raf₈ and<em> Pseudomonas </em>sp. raf₄, respectively. To detect INA genes, two sets of degenerate primers were used and partial INA gene sequences were amplified. INA gene sequence) 425bp) for <em>Pseudomonas</em><em> putida </em>raf_6, <em>Pantoea agglomerans </em>raf₇ and<em> P. fragi </em>raf₃ were amplified with primer pair of 3308/3463. Whereas, a fragment of 194bp was detected in <em>Pseudomonas </em>sp. raf₄, <em>P. moraviensis </em>raf₅ and<em> P. moraviensis </em>raf₁using forward and reverse primer pair of 3076/3463. <em>Entrobacter cloacea </em>raf₈has reported for the first time as epiphytic ice plus strain. The capability of the latter as a bacterial <em>biocontrol</em> agent against insect pests was reported.