CYP6G4 is a possible ortholog of the CYP6G1 gene in D. melanogaster and constitutive overexpression of CYP6G1 is causing DDT and neonicotinoid resistance in the fruit fly. A similar role for CYP6G4 in houseflies could be suggested. Recently, CYP6G4 has shown to be over-expressed in a pyrethroid resistant housefly strain from China, but no causal link was established. In this study, CYP6G4 expression was higher in the INCB28060 1029712-80-8 791spin strain compared to the susceptible strain, 2-fold in males and almost 15-fold in females. However, CYP6G4 gene expression in 791spin was lower than that of the spinosad susceptible strains. CYP6G4 gene expression was not significantly different between sexes, regardless of treatment. Spinosad treatment caused a gene expression increase in males of the 791spin strain; whereas spinosad treatment decreased CYP6G4 expression in females, but neither of the effects were significant. This is contradicted by the other spinosad susceptible strains having a higher expression level of CYP6G4 than 791spin, so a possible role of CYP6G4 seems to be minor. Further investigations, especially description of the CYP6G4 alleles, are needed to elucidate the role of CYP6G4 in this strain as well as its potential role in xenobiotic metabolism and its importance as a housefly insecticide resistance gene. Increased expression has been linked to elevated resistance by increased enzymatic degradation of insecticide e.g. degradation of deltamethrin by CYP6D1. But, what does down-regulation signify? It could be hypothesized that down-regulation is due to reduced energy costs. Perhaps down-regulation is a part of a coupled system, so that groups of P450 genes are up-regulated and others are down-regulated. An example of this is a transcription factor pathway in Drosophila where 20% of differential expressed genes are genes targeted by the transcription factor CncC. Alternatively the action is more direct and a given P450 enzyme activates the insecticide, making it more toxic. This is e.g. shown by bioassay experiments with neonicotinoids. The addition of the synergist PBO to feeding test with imidacloprid increased toxicity 9-fold, whereas toxicity of thiamethoxam decreased in male houseflies. Thiamethoxam has also been shown as a proinsecticide in plants. Down-regulation will thus have a direct survival effect. The overall high expression level of CYP4G2 throughout the strains also indicates importance of this gene. However, the data on 791spin are not conclusive concerning spinosad resistance and e.g. pointing to a single spinosad resistance gene. Small contributions from multiple P450s with different enzymatic capabilities could be speculated to do the job in 791spin. Analyzing the expression of metabolic detoxification genes rarely gives a clear and unambiguous answer to which enzymes are involved in resistance. Furthermore, the P450s ability to attack the highly complicated spinosad molecule is still unresolved. There is an indication of CYP6G4 as an insecticide-resistance gene, where involvement in spinosad resistance cannot be rejected. The high expression levels of P450 genes in flies from a field population compared to established laboratory strains presented in this study questions.