Systemic gene transfer of binding immunoglobulin protein (BiP) prevents disease progression in murine collagen-induced arthritis.

Summary Recombinant human binding immunoglobulin protein (BiP) has previously demonstrated anti-inflammatory properties in multiple models of inflammatory arthritis. We investigated whether these immunoregulatory properties could be exploited using gene therapy techniques. A single intraperitoneal injection of lentiviral vector containing the murine BiP (Lenti-mBiP) or green fluorescent protein (Lenti-GFP) transgene was administered in low- or high-dose studies during early arthritis. Disease activity was assessed by visual scoring, histology, serum cytokine and antibody production measured by cell enzyme-linked immunosorbent assay (ELISA) and ELISA, respectively. Lentiviral vector treatment caused significant induction of interferon (IFN)-γ responses regardless of the transgene; however, further specific effects were directly attributable to the BiP transgene. In both studies Lenti-mBiP suppressed clinical arthritis significantly. Histological examination showed that low-dose Lenti-mBiP suppressed inflammatory cell infiltration, cartilage destruction and significantly reduced pathogenic anti-type II collagen (CII) antibodies. Lenti-mBiP treatment caused significant up-regulation of soluble cytotoxic T lymphocyte antigen-4 (sCTLA-4) serum levels and down-regulation of interleukin (IL)-17A production in response to CII cell restimulation. In-vitro studies confirmed that Lenti-mBiP spleen cells could significantly suppress the release of IL-17A from CII primed responder cells following CII restimulation in vitro, and this suppression was associated with increased IL-10 production. Neutralization of CTLA-4 in further co-culture experiments demonstrated inverse regulation of IL-17A production. In conclusion, these data demonstrate proof of principle for the therapeutic potential of systemic lentiviral vector delivery of the BiP transgene leading to immunoregulation of arthritis by induction of soluble CTLA-4 and suppression of IL-17A production.

Monitoring of diamondback moth (Lepidoptera: Plutellidae) resistance to spinosad, indoxacarb, and emamectin benzoate.

Six to nine populations of the diamondback moth, Plutella xylostella (L.), were collected annually from fields of crucifer vegetables in the United States and Mexico from 2001 to 2004 for baseline susceptibility tests and resistance monitoring to spinosad, indoxacarb, and emamectin benzoate. A discriminating concentration for resistance monitoring to indoxacarb and emamectin benzoate was determined based on baseline data in 2001 and was used in the diagnostic assay for each population in 2002-2004 together with a discriminating concentration for spinosad determined previously. Most populations were susceptible to all three insecticides, but a population from Hawaii in 2003 showed high levels of resistance to indoxacarb. Instances of resistance to spinosad occurred in Hawaii (2000), Georgia (2001), and California (2002) as a consequence of a few years of extensive applications in each region. The collaborative monitoring program between university and industry scientists we discuss in this article has provided useful information to both parties as well as growers who use the products. These studies provide a baseline for developing a more effective resistance management program for diamondback moth.

Increased susceptibility to ventricular arrhythmias in hypertensive paraplegic rats.

Individuals with spinal cord injury (SCI) between thoracic vertebrae four (T4) and five (T5) have elevated levels of sympathetic activity to the heart. Notably, female spontaneously hypertensive rats (SHR) also have increased cardiac sympathetic nerve activity (SNA). Since elevated levels of cardiac SNA increase the risk for cardiac arrhythmias, we tested the hypothesis that hypertensive, paraplegic rats have an increased susceptibility to ventricular arrhythmias. To test this hypothesis, intact (n = 7) and paraplegic hypertensive rats (n = 6) were chronically instrumented with silver stimulating electrodes on the left ventricle, electrocardiogram (ECG) recording electrodes and an arterial catheter. After recovery, the effective refractory period, the electrical stimulation threshold to induce ventricular arrhythmias and cardiac sympathetic tonus (ST) were determined. Paraplegic rats had a lower effective refractory period (35%), lower electrical stimulation threshold to induce ventricular arrhythmias (62%), and higher cardiac ST (84%). These data document an increased susceptibility to ventricular arrhythmias in hypertensive, paraplegic rats.

Monitoring and characterization of diamondback moth (Lepidoptera: Plutellidae) resistance to spinosad.

Fourteen populations of the diamondback moth, Plutella xylostella (L.), were collected from fields of crucifer vegetables in the United States, Mexico, and Thailand in 1999 and 2000 for susceptibility tests with spinosad. Most populations were susceptible to spinosad and similar to earlier baseline values, but populations from Thailand and Hawaii showed high levels of tolerance. A statewide survey in Hawaii in 2000 and 2001 indicated resistance problems on several islands. One colony collected in October 2000 from Pearl City, HI, was subjected to further selection pressure, using spinosad in the laboratory, and then was used as the resistant strain (Pearl-Sel) for other tests. Spray tests using the recommended field rates of spinosad on potted broccoli plants in the greenhouse confirmed that field control failures due to resistance were possible in the areas of these collections. Analysis of probit lines from F1 reciprocal crosses between the Pearl-Sel and S strain indicated that resistance to spinosad was inherited autosomally and was incompletely recessive. A direct test of monogenic inheritance based on the F1 x Pearl-Sel backcrosses suggested that resistance to spinosad was probably controlled by one locus. The synergists S,S,S-tributyl phosphorotrithioate and piperonyl butoxide did not enhance the toxicity of spinosad to the resistant colony, indicating metabolic mediated detoxification was probably not responsible for the spinosad resistance. Two field colonies in Hawaii that were resistant to spinosad were not cross-resistant to emamectin benzoate or indoxacarb. Resistance developed in Hawaii due to the continuous cultivation of crucifers in which as many as 50 applications of spinosad per year may have been made to a common population of P. xylostella in sequential plantings, although each grower might have used the labeled restrictions for resistance management. Resistance management strategies will need to address such cropping and pest management practices.

Daily exercise reduces measures of heart rate and blood pressure variability in hypertensive rats.

This study was designed to test the hypothesis that daily spontaneous running (DSR) reduces measures of heart rate and blood pressure variability in spontaneously hypertensive rats (SHR). After 8 weeks of DSR or sedentary control, rats were chronically instrumented with arterial catheters. Daily exercise reduced most measures of heart rate (HR) and blood pressure variability. Specifically DSR decreased heart rate, Low Frequency Power (LF: 0.19-0.61 Hz), and Low Frequency/High Frequency (HF: 1.2-2.5 Hz) ratio of HR. Furthermore, Total Power (TP), LF power, and LF/HF ratio of systolic blood pressure were reduced by daily spontaneous running. Finally, TP, LF and HF powers and LF/HF ratios of diastolic blood pressure were reduced by daily spontaneous running. These data demonstrate that daily exercise reduces sympathetic activity and possibly increases cardiac reserve in hypertensive animals.

Sino-aortic denervation alters the hemodynamic response to exercise in hypertensive rats.

The effect of sino-aortic denervation (SAD) on the heart rate (HR), arterial pressure (AP) and regional blood flow responses during dynamic exercise was examined in spontaneously hypertensive rats (SHR). Intact (n= 14) and SAD (n= 17) rats were instrumented with arterial catheters and mesenteric and iliac Doppler ultrasonic flow probes. After recovery, all rats underwent a graded exercise test. Heart rate increased significantly during exercise in intact and SAD rats. There was no significant difference in the steady state heart rate response to exercise in the intact and SAD rats. Arterial pressure increased during exercise in the intact rats. In sharp contrast, arterial pressure decreased during exercise in the SAD rats. Iliac vascular conductance increased during exercise in the intact and SAD rats. The increase in iliac vascular conductance during exercise was significantly greater in the SAD rats. Mesenteric vascular conductance decreased during exercise in the intact and SAD rats. The decrease in mesenteric vascular conductance during exercise was significantly attenuated in the SAD rats. Results suggest that functioning arterial baroreceptors are required for the typical hemodynamic responses during dynamic treadmill running in hypertensive rats.

Mechanisms mediating NTS P2x receptor-evoked hypotension: cardiac output vs. total peripheral resistance.

We have previously shown that P2x purinoceptor activation in the subpostremal nucleus tractus solitarius (NTS) produces dose-dependent decreases in mean arterial pressure (MAP), heart rate, efferent sympathetic nerve activity, and significant peripheral vasodilation. However, the relative roles of cardiac output (CO) and total peripheral resistance (TPR) in mediating this depressor response are unknown. Bradycardia does not necessarily result in decreased CO, because, with the greater filling time, stroke volume may increase such that CO may be unchanged. We measured changes in CO (via a chronically implanted flow probe on the ascending aorta) and MAP in alpha-chloralose- and urethane-anesthetized male Sprague-Dawley rats in response to microinjection of the selective P2x purinoceptor agonist alpha,beta-methylene ATP (25 and 100 pmol/50 nl) into the subpostremal NTS. TPR was calculated as MAP/CO. At the low dose of NTS P2x purinoceptor agonist, the reduction in MAP was primarily mediated by reductions in TPR (-31.3 +/- 3.3%), not CO (-8.7 +/- 1.7%). At the high dose, both CO (-34.4 +/- 6.6%) and TPR (-40.2 +/- 2.5%) contribute to the reduction in MAP. We conclude that the relative contribution of CO and TPR to the reduction in MAP evoked by NTS P2x purinoceptor activation is dependent on the extent of P2x purinoceptor activation.

MHC class Ia-restricted T cells partially account for beta2-microglobulin-dependent resistance to Mycobacterium tuberculosis.

Recent studies have highlighted the heterogeneous nature of the CD8(+) T cell response during human Mycobacterium tuberculosis infection; MHC class Ia, MHC class Ib and CD1 have all been identified as significant restriction elements. Here we have attempted to define the role of MHC class Ia in resistance to M. tuberculosis infection in mice. The course of M. tuberculosis infection in mice deficient in a single MHC class Ia molecule, either H2-K(b) or H2-D(b), was essentially identical to that observed in wild-type mice. In contrast, mice fully deficient in MHC class Ia molecules (H2-K(b) / H2-D(b) double knockout mice) were substantially more susceptible to M. tuberculosis infection. However, the double knockout mice were not as susceptible as beta 2-microglobulin-deficient mice, which have a broader phenotypic deficit. Thus, antigen presentation via MHC class Ia is an important component in resistance to M. tuberculosis, but its absence only partially accounts for the increased susceptibility of beta 2-microglobulin-deficient mice.

Recovery from carotid artery catheterization performed under various anesthetics in male, Sprague-Dawley rats.

This study was designed to determine the time to recovery from carotid artery catheterization using multiple criteria and to compare recovery times between three common anesthetics. Male Sprague-Dawley rats, chronically instrumented with radio-telemetry transmitters, were anesthetized with sodium pentobarbital, halothane or a mixture of ketamine, xylazine and acepromazine before an indwelling catheter was placed in the carotid artery. The procedure was completed in less than 15 min. Changes in body weight, food and water consumption, blood pressure, heart rate and activity were used to determine recovery. As judged by recovery of body weight, animals anesthetized with each of the anesthetics recovered by the 4th day after catheterization. Food and water consumption normalized by 1-2 days after surgery. Heart rates and blood pressures during the light phase of the photoperiod were significantly increased for 2 days by all anesthetics. During the dark phase of the photoperiod, heart rates and blood pressures were not significantly affected by pentobarbital or halothane anesthesia, but were significantly decreased and increased respectively on the night immediately following surgery in the ketamine / xylazine / acepromazine-anesthetized rats. Delayed elevations of heart rate were observed in pentobarbital and halothane anesthetized rats on days and/or nights 5 and 6 post surgery. Animal activity patterns during the light phase of the photoperiod were not affected by pentobarbital or halothane, but were increased by ketamine 2 days after surgery. During the dark phase, halothane transiently reduced activity whereas ketamine-anesthetized rats showed reduced activity for 4 nights post surgery. These studies show that recovery depends on the criteria selected and the anesthetic used, but, in general, 2-4 days were required for recovery from this relatively simple procedure.

Central blockade of vasopressin V(1) receptors attenuates postexercise hypotension.

We tested the hypothesis that central arginine vasopressin (AVP) mediates postexercise reductions in arterial pressure (AP) and heart rate (HR). To test this hypothesis, nine spontaneously hypertensive rats (SHR) were instrumented with a 22-gauge stainless steel guide cannula in the right lateral cerebral ventricle and with a carotid arterial catheter. After the rats recovered, AP and HR were assessed before and after a single bout of dynamic exercise with the central administration of vehicle or the selective AVP V(1)-receptor antagonist d(CH(3))(5) Tyr(Me)-AVP (AVP-X). AP and HR were significantly decreased below preexercise values with central administration of vehicle [P < 0.05, change (Delta)-21 +/- 4 mmHg and Delta-20 +/- 6 beats/min, respectively]. In sharp contrast, after exercise with central administration of AVP-X, both AP (Delta+8 +/- 5 mmHg) and HR (Delta+24 +/- 9 beats/min) were not significantly different from preexercise values (P > 0.05). Furthermore, AVP-X at rest did not significantly alter AP (181 +/- 11 vs. 178 +/- 11 mmHg, P > 0.05) or HR (328 +/- 24 vs. 331 +/- 22 beats/min, P > 0.05). Thus central blockade of AVP V(1) receptors prevented postexercise reductions in AP and HR. These data suggest that AVP, acting within the central nervous system, mediates postexercise reductions in AP and HR in the SHR.

Greenhouse tests on resistance management of Bt transgenic plants using refuge strategies.

Experimental evaluation of the effectiveness of resistance management tactics is vital to help provide guidelines for the deployment of transgenic insecticidal crops. Transgenic broccoli expressing a Cry1Ac gene of Bacillus thuringiensis (Bt) and the diamondback moth, Plutella xylostella (L.), were used in greenhouse tests to evaluate the influence of size and placement of nontransgenic refuge plants on changes in resistance allele frequency and pest population growth. In the first test with an initial Cry1Ac-resistance (R) allele frequency of 0.007, P. xylostella were introduced into cages with the following treatments: 0, 3.3, 10, 20, and 100% refuge plants. Results after four generations showed that resistance could be delayed by increasing the proportion of refuge plants in the cage. Population growth was also influenced by refuge size with the highest populations occurring in treatments that had either no refuge plants or all refuge plants. In the second test, we evaluated the effect of refuge placement by comparing 20% separate and 20% mixed refuges. P. xylostella with an initial frequency of resistant alleles at 0.0125 were introduced into cages and allowed to cycle; later generations were evaluated for resistance and population growth. Separating the refuge had a pronounced effect on delaying resistance and slowing establishment of resistant larvae on Bt plants. Combining information from both trials, we found a strong negative correlation between the number of larvae on Bt plants and the mortality of the population in leaf dip bioassays. Results from larval movement studies showed that separate refuges delayed resistance better than mixed refuges because they conserved relatively more susceptible alleles than R alleles and did not increase the effective dominance of resistance.

Severe exercise alters the strength and mechanisms of the muscle metaboreflex.

Previous studies have shown that in dogs performing mild to moderate treadmill exercise, partial graded reductions in hindlimb blood flow cause active skeletal muscle to become ischemic and metabolites to accumulate thus evoking the muscle metaboreflex. This leads to a substantial reflex increase in mean arterial pressure (MAP) mediated almost solely via a rise in cardiac output (CO). However, during severe exercise CO is likely near maximal and thus metaboreflex-mediated increases in MAP may be attenuated. We therefore evoked the metaboreflex via partial graded reductions in hindlimb blood flow in seven dogs during mild, moderate, and severe treadmill exercise. During mild and moderate exercise there was a large rise in CO (1.5 +/- 0.2 and 2.2 +/- 0.3 l/min, respectively), whereas during severe exercise no significant increase in CO occurred. The rise in CO caused a marked pressor response that was significantly attenuated during severe exercise (26.3 +/- 7.0, 33.2 +/- 5.6, and 12.2 +/- 4.8 mmHg, respectively). We conclude that during severe exercise the metaboreflex pressor response mechanisms are altered such that the ability of this reflex to increase CO is abolished, and reduced pressor response occurs only via peripheral vasoconstriction. This shift in mechanisms likely limits the effectiveness of the metaboreflex to increase blood flow to ischemic active skeletal muscle. Furthermore, because the metaboreflex is a flow-raising reflex and not a pressure-raising reflex, it may be most appropriate to describe the metaboreflex magnitude based on its ability to evoke a rise in CO and not a rise in MAP.

Carotid baroreflex pressor responses at rest and during exercise: cardiac output vs. regional vasoconstriction.

The arterial baroreflex mediates changes in arterial pressure via reflex changes in cardiac output (CO) and regional vascular conductance, and the relative roles may change between rest and exercise and across workloads. Therefore, we quantified the contribution of CO and regional vascular conductances to carotid baroreflex-mediated increases in mean arterial pressure (MAP) at rest and during mild to heavy treadmill exercise (3.2 kph; 6.4 kph, 10% grade; and 8 kph, 15% grade). Dogs (n = 8) were chronically instrumented to measure changes in MAP, CO, hindlimb vascular conductance, and renal vascular conductance in response to bilateral carotid occlusion (BCO). At rest and at each workload, BCO caused similar increases in MAP (average 35 +/- 2 mmHg). In response to BCO, neither at rest nor at any workload were there significant increases in CO; therefore, the pressor response occurred via peripheral vasoconstriction. At rest, 10.7 +/- 1.4% of the rise in MAP was due to vasoconstriction in the hindlimb, whereas 4.0 +/- 0.7% was due to renal vasoconstriction. Linear regression analysis revealed that, with increasing workloads, relative contributions of the hindlimb increased and those of the kidney decreased. At the highest workload, the decrease in hindlimb vascular conductance contributed 24.3 +/- 3.4% to the pressor response, whereas the renal contribution decreased to only 1.6 +/- 0.3%. We conclude that the pressor response during BCO was mediated solely by peripheral vasoconstriction. As workload increases, a progressively larger fraction of the pressor response is mediated via vasoconstriction in active skeletal muscle and the contribution of vasoconstriction in inactive beds (e.g., renal) becomes progressively smaller.

Different cross-resistance patterns in the diamondback moth (Lepidoptera: Plutellidae) resistant to Bacillus thuringiensis toxin Cry1C.

Two strains of the diamondback moth, Plutella xylostella (L.), were selected using Cry1C protoxin and transgenic broccoli plants expressing a Cry1C toxin of Bacillus thuringiensis (Bt). Both strains were resistant to Cry1C but had different cross-resistance patterns. We used 12 Bt protoxins for cross-resistance tests, including Cry1Aa, Cry1Ab, Cry1Ac, Cry1Bb, Cry1C, Cry1D, Cry1E, Cry1F, Cry1J, Cry2Ab, Cry9Aa, and Cry9C. Compared with the unselected sister strain (BCS), the resistance ratio (BR) of one strain (BCS-Cry1C-1) to the Cry1C protoxin was 1,090-fold with high level of cross-resistance to Cry1Aa, Cry1Ab, Cry1Ac, Cry1F, and Cry1J (RR > 390-fold). The cross-resistance to Cry1A, Cry1F, and Cry1J in this strain was probably related to the Cry1A resistance gene(s) that came from the initial field population and was caused by intensive sprayings of Bt products containing Cry1A protoxins. The neonates of this strain can survive on transgenic broccoli plants expressing either Cry1Ac or Cry1C toxins. The other strain (BCS-Cry1C-2) was highly resistant to Cry1C but not cross-resistant to other Bt protoxins. The neonates of this strain can survive on transgenic broccoli expressing Cry1C toxin but not Cry1Ac toxin. The gene(s) conferring resistance to Cry1C segregates independently from Cry1Ac resistance in these strains. The toxicity of Cry1E and Cry2Ab protoxins was low to all of the three strains. The overall progress of all work has resulted in a unique model system to test the stacked genes strategy for resistance management of Bt transgenic crops.

Prospects for better tuberculosis vaccines.

Tuberculosis remains one of the top three infectious disease killers. Treatment is long and expensive and drug resistant strains of Mycobacterium tuberculosis are already on the rise. The current vaccine, BCG, is ineffective in parts of the world where the disease is most widespread and therefore the search for a novel, more effective vaccine is paramount. In this review we discuss the current state of vaccine research, including the identification of candidate antigens and the current methods used for their evaluation.

Daily exercise attenuated the sympathetic component of the spontaneous arterial baroreflex control of heart rate in hypertensive rats.

The influence of daily spontaneous running on the sympathetic and parasympathetic components of the spontaneous arterial baroreflex control of heart rate was examined in 22 female spontaneously hypertensive rats [12 sedentary and 10 daily spontaneous running]. Following 8 weeks of sedentary control or daily spontaneous running, animals were chronically instrumented with an arterial catheter. Daily spontaneous running resulted in an increased heart weight/body weight ratio (5.2 +/- 0.27 vs 4.3 +/- 0.01 g/kg) and a resting bradycardia (321+/- 8 bpm vs 360 +/- 6). The spontaneous changes in arterial pressure and the reflex responses of heart rate were examined under three experimental conditions: 1) pre-blockade, 2) following beta1-adrenergic receptor blockade, and 3) following muscarinic-cholinergic receptor blockade. Daily spontaneous running attenuated the spontaneous gain of the arterial baroreflex control of heart rate (56%). After muscarinic-cholinergic receptor blockade, the spontaneous gain remained reduced in daily spontaneous running rats (57%). In contrast, after beta1-adrenergic receptor blockade the spontaneous gain was not different between sedentary control and daily spontaneous running animals. Results demonstrate that daily spontaneous running decreased the sympathetic component resulting in an apparently greater influence of the parasympathetic component on the spontaneous arterial baroreflex control of heart rate.

Autonomic and endothelial dysfunction in experimental diabetes.

The effect of streptozotocin induced diabetes on autonomic regulation of heart rate and endothelial function was examined in Sprague-Dawley rats. Weanling rats (3-4 weeks of age) of either sex were randomly assigned to a non-diabetic (male 5, female 6) or diabetic (male 4, female 5). Diabetes was induced with a single intraperitoneal (IP) injection of streptozotocin (STZ, 100 mg/kg). Nondiabetic rats received an IP injection of saline. Eight weeks after injection, rats were chronically instrumented with a left jugular venous catheter and a left carotid arterial catheter. After recovery (5 days) cardiac sympathetic tonus, parasympathetic tonus and intrinsic heart rate were determined. On an alternative day, the pressor response to nitric oxide synthase inhibition (NOS-X) was determined in areflexic rats. Cardiac sympathetic tonus (72 +/- 13 vs. 41 +/- 7), parasympathetic tonus (-51 +/- 10 vs. -22 +/- 7), and intrinsic heart rate (368 +/- 6 vs. 292 +/- 9), were reduced in diabetic rats. Furthermore, diabetic rats had a smaller pressor response (A33 +/- 7 vs. A66 +/- 5) to NOS-X. These results document impaired autonomic control of heart rate and endothelial dysfunction in 8-week streptozotocin induced diabetic rats.

Development and characterization of diamondback moth resistance to transgenic broccoli expressing high levels of Cry1C.

A field-collected colony of the diamondback moth, Plutella xylostella, had 31-fold resistance to Cry1C protoxin of Bacillus thuringiensis. After 24 generations of selection with Cry1C protoxin and transgenic broccoli expressing a Cry1C protein, the resistance that developed was high enough that neonates of the resistant strain could complete their entire life cycle on transgenic broccoli expressing high levels of Cry1C. After 26 generations of selection, the resistance ratios of this strain to Cry1C protoxin were 12,400- and 63,100-fold, respectively, for the neonates and second instars by a leaf dip assay. The resistance remained stable until generation 38 (G38) under continuous selection but decreased to 235-fold at G38 when selection ceased at G28. The Cry1C resistance in this strain was seen to be inherited as an autosomal and incompletely recessive factor or factors when evaluated using a leaf dip assay and recessive when evaluated using Cry1C transgenic broccoli. Saturable binding of (125)I-Cry1C was found with brush border membrane vesicles (BBMV) from both susceptible and Cry1C-resistant strains. Significant differences in Cry1C binding to BBMV from the two strains were detected. BBMV from the resistant strain had about sevenfold-lower affinity for Cry1C and threefold-higher binding site concentration than BBMV from the susceptible strain. The overall Cry1C binding affinity was just 2.5-fold higher for BBMV from the susceptible strain than it was for BBMV from the resistant strain. These results suggest that reduced binding is not the major mechanism of resistance to Cry1C.

Assessment of insecticide resistance after the outbreak of diamondback moth (Lepidoptera: Plutellidae) in California in 1997.

During an outbreak of the diamondback moth, Plutella xylostella (L.), in California in 1997, nine populations were collected from the major broccoli areas throughout the state. Populations were assayed for their susceptibility to currently used materials (Bacillus thuringiensis subsp. kurstaki, permethrin, and methomyl) and to newer materials that had not yet been commercially used in California (spinosad, emamectin benzoate, and chlorfenapyr). For the currently used insecticides, elevated levels of resistance were seen only with permethrin and seven of the nine populations had tolerance ratios (TR) of > 100. With the newer chemistries, TR values were all < 15. To compare potential cross-tolerance, TR values of the currently used insecticides were compared with TR values of the newer insecticides. There were significant relationships found between: methomyl and emamectin benzoate, methomyl and spinosad, and permethrin and spinosad. Further biochemical studies are needed to confirm the actual mechanisms that lead to these relationships and field tests are needed to determine what impact, if any, such TR levels would have on control in the field. These data indicate that resistance to at least one of the commonly used insecticides (permethrin) may have played a role in the outbreak during 1997. However, other factors may have been at least equally important. The winter of 1996-1997 was warmer than normal, and during the period from February through August of 1997 the amount of rainfall was < 50% of normal. Hot and dry conditions are known to be conducive to outbreaks of P. xylostella. These data add to an overall knowledge about the geographic variation of resistance in P. xylostella populations within the United States. They also serve as a baseline for monitoring changes in susceptibility to these newer insecticides and can also help explain the occurrence of outbreaks caused by factors other than insecticide resistance.