Research On Biological Response Of Different Phosphine Resistant Strains Of Cryptolestes Ferrugineus(Stephens) To Hypoxia Stress

Phosphine is the main fumigant for grain storage pest control,but the increasing phosphine resistance of stored grain pests has seriously affected its effect and use in insect pest control.The use of controlled atmosphere（CA）for the insect control is an important alternative to phosphine.The adaptability and mechanisms of phosphine resistant pests to CA,especially in low concentration of oxygen enriched with nitrogen,can affect the application of insect control by CA and even the development of tolerance of insect to CA.Research on the biological response mechanisms of pests under low oxygen atmosphere will be conducive to to the effective use of insect control by CA and avoid the development of tolerance of insect to CA.The problem of resistance to phosphine is particularly prominent in Cryptolestes ferrugineus（Stephens）among various stored grain insects.Research on the biological response mechanisms of C.ferrugineus to low oxygen atmosphere will also be beneficial for its scientific management.In this study,the strains of C.ferrugineus from the grain storage and processing center in Nansha Lingnan Suiliang,Guangdong（labeled in LNCf strain name）,a grain warehouse in Zhongshan City,Guangdong（labeled in ZSCf strain）,and Manitoba,Canada（labeled in MCf strain）were used for resistance test.The mortality of eggs,larvae,pupae,and adults of C.ferrugineusr for all three strains under 2%O₂at different exposure times were studied at temperatures of 18,23,and 28℃,respectively.Population life tables were constructed for C.ferrugineus that survived form 2%O₂after exposure for half lethal time.The generation survival rates,oviposition,developmental period,adult lifespan,and population trend index of different strains of C.ferrugineus were also examined at the same conditions.Furthermore,the activities of Catalase（CAT）,Peroxidase（POD）,and Superoxide dismutase（SOD）as well as lactate content in the bodies of different strains of C.ferrugineuss adults were measured after exposure in 2%O₂for 24,48,and 72 h.The main results were as followed:（1）The resistance of different strains of C.ferrugineus was determined using the FAO recommended method.The toxic regression equation for the LNCf strain was y=7.981x-9.624,with an LC₅₀value of 16.074 mg/L,the toxic regression equation for the ZSCf strain was y=3.630x-1.699,with an LC₅₀value of 2.938 mg/L,and the toxic regression equation for the MCf strain was y=4.498x-6.323,with an LC₅₀value of 0.035 mg/L.Compared with the reported LC₅₀value of the reference strain of C.ferrugineus（0.011 mg/L）,the resistance factors（Rf）of the LNCf strain used in this study reached 1461,the Rf of the ZSCf strain was 267,and the Rf of the MCf strain was 3.The resistance of the C.ferrugineus strains used in this study included three levels:extremely high resistance,high resistance,and no resistance.The control for the strains of C.ferrugineus collected from two locations in Guangdong Province by fumigation was more difficult compared with the relatively sensitive MCf strain.（2）At 18℃,the LT₉₉（exposure time to reach 99%mortality）values of eggs,larvae,pupae,and adults for the extremely resistant LNCf strain of C.ferrugineus in 2%O₂were 32,22,32,and 18 d,respectively.The LT₉₉values of eggs,larvae,pupae,and adults for the high resistant ZSCf strain were 24,18,24,and 16 d,respectively.The LT₉₉values of eggs,larvae,pupae,and adults of the non-resistant MCf strain were 20,16,20,and 14 d,respectively.At23℃,the LT₉₉values of eggs,larvae,pupae,and adults for the extremely resistant LNCf strain of C.ferrugineus were 24,16,24,and 14 d,respectively.The LT₉₉values of eggs,larvae,pupae,and adults for the high resistant ZSCf strain were 18,14,18,and 12 d,respectively.The LT₉₉values of eggs,larvae,pupae,and adults of the non-resistant MCf strain were 16,12,16,and 10 d,respectively.At 28℃,the LT₉₉values of eggs,larvae,pupae,and adults for the extremely resistant LNCf strain of C.ferrugineus were 18,12,18,and 9 d,respectively.The LT₉₉values of eggs,larvae,pupae,and adults for the high resistant ZSCf strain were 13,10,14,and 8 d,respectively.The LT₉₉values of eggs,larvae,pupae,and adults of the non-resistant MCf strain were 12,10,12,and 7 d,respectively.At every test temperature,the order of tolerance to 2%O₂for different stages of the C.ferrugineus strains was:pupae>eggs>larvae>adults.The LT₉₉values of the pupal stage of the extremely resistant LNCf strain of C.ferrugineus were four times bigger than those of the adult stage,while the LT₉₉values of the pupal stage of the high resistant ZSCf strain were three times bigger than those of the adult stage.The LT₉₉values of the pupal stage of the non-resistant MCf strain were also three times bigger than those of the adult stage.At the same temperature,the LT₉₉values of different developmental stages of the extremely resistant LNCf strain of C.ferrugineus were also bigger than those of the high resistant ZSCf strain and the non-resistant MCf strain.For the same resistance strain of C.ferrugineus,the LT₉₉values of different developmental stages at 18℃were bigger than those at 23℃and28℃,and decreasing the temperature significantly increased the time required to cause mortality in the pests.（3）At 30℃and 2%hypoxia stress,the total developmental period,generation survival rate,male lifespan,female lifespan,inhibition rate of oviposition,and population trend index of the extremely resistant LNCf strain of C.ferrugineus after exposure for the half lethal time were 29 d,36.33%,93 d,108 d,28.75%,and 15.52,respectively.The corresponding values for the high resistant ZSCf strain were 31 d,29.67%,80 d,84 d,36.59%,and 13.41,respectively.The corresponding values for the non-resistant MCf strain were 33 d,27.33%,72 d,79 d,41.05%,and 12.25,respectively.The generation survival rate,male lifespan,female lifespan,and population trend index of the extremely resistant LNCf strain of C.ferrugineus were all bigger than those of the high resistant ZSCf strain and the non-resistant MCf strain.The total developmental period and oviposition inhibition rate of the non-resistant MCf strain were bigger than those of the high resistant ZSCf strain and the extremely resistant LNCf strain.The order of adaptability to hypoxia stress in C.ferrugineus was LNCf>ZSCf>MCf strain.（4）The activities of CAT,POD,and SOD and the lactic acid content in the extremely resistant LNCf strain of C.ferrugineus under 2%hypoxia stress for 24 h were 38.44,14.71,22.32 U/mg prot,and 19.27 mmol/（L·mg）,respectively.Under 2%hypoxia stress for 48 h,the activities of CAT,POD,and SOD and the lactate content were 60.89,17.47,24.53 U/mg prot,and 45.75 mmol/（L·mg）,respectively.Under 2%hypoxia stress for 72 h,the activities of CAT,POD,and SOD and the lactate content were 54.19,12.48,17.42 U/mg prot,and53.62 mmol/（L·mg）,respectively.The activities of CAT,POD,and SOD and the lactic acid content in the high resistant ZSCf strain of C.ferrugineus under 2%hypoxia stress for 24 h were 27.67,11.87,27.18U/mg prot,and 18.37 mmol/（L·mg）,respectively.Under 2%hypoxia stress for 48 h,the activities of CAT,POD,and SOD and the lactic acid content were 37.53,14.12,29.29 U/mg prot,and 45.92 mmol/（L·mg）,respectively.Under 2%hypoxia stress for 72 h,the activities of CAT,POD,and SOD and the lactic acid content were 31.16,10.39,24.15 U/mg prot,and51.83 mmol/（L·mg）,respectively.The activities of CAT,POD,and SOD and the lactic acid content in the non-resistant MCf strain of C.ferrugineus under 2%hypoxia stress for 24 h were 10.59,6.43,32.48 U/mg prot,and 19.46 mmol/（L·mg）,respectively.Under 2%hypoxia stress for 48 h,the activities of CAT,POD,and SOD and the lactic acid content were 17.13,8.23,35.41 U/mg prot,and46.21 mmol/（L·mg）,respectively.Under 2%hypoxia stress for 72 h,the activities of CAT,POD,and SOD and the lactic acid content were 16.17,5.51,29.33 U/mg prot,and 52.72mmol/（L·mg）,respectively.Under hypoxia stress conditions,the activities of CAT and POD in the extremely resistant LNCf strain of C.ferrugineus with different levels of phosphine resistance were significantly higher than those in the high resistant ZSCf strain and non-resistant MCf strain.The activity of SOD in the extremely resistant LNCf strain of C.ferrugineus was significantly lower than that in the high resistant ZSCf strain and non-resistant MCf strain.There was no significant difference in lactate content among the extremely resistant LNCf strain,high resistant ZSCf strain,and non-resistant MCf strain.The activities of CAT,POD,and SOD in C.ferrugineus increased first and then decreased with the inscreasing of exposure time,and reached the maximum value at 48 h.The lactic acid content in C.ferrugineus increased with the inscreasing of exposure time.