The overall organisation and branching pattern of the embryonic tracheal system is unaffected in cold^f05607 mutants. However, both the shape and the length of the tracheal dorsal trunk segments are abnormal in mutant flies. Differing from the wild-type, this structure appears in stage 15 embryos as a succession of bulging cysts and, by stage 16, as an abnormally convoluted tube. The fibrous structure of the tracheal chitin cable is disorganised in mutant embryos and chitin is deposited as an amorphous material in the tracheal lumen.

In contrast to wild-type controls, soluble dye rapidly diffuses into the lumen of stage 16 cold^f05607 mutant trachea, indicating a role for cold in organisation of the paracellular barrier.

cold^f05607 mutants do not exhibit defects in cell polarity or in the assembly of other adhesion structures.

cold^f05607 embryos lacking the cold maternal contribution (but rescued paternally with a wild-type allele, using the FLP-DFS technique) do not show any phenotype and survive into adulthood. In contrast, mutants embryos lacking both maternal and zygotic contributions die during embryogenesis but do not display obvious morphological defects and are indistinguishable from embryos lacking only the cold zygotic contribution.

A 10kDa dextran dye readily diffuses into the ventral cord of cold^f05607 mutant embryos, whereas it is efficiently excluded from this structure in wild-type embryos of the same age. Similarly, injected dye diffuses into the lumen of the chordotonal organs in stage 17 cold^f05607 embryos whereas septate junctions established between the cap, scolopal and ligament cells prevent dye intake in wild-type controls.

cold^f05607 mutants display a dye penetration phenotype where fluorescent dye diffuses across the blood-brain barrier into the CNS. The overall organization of the nervous system as well as the distribution of all glial cells, however, appears normal.

Homozygous cold^f05607 mutant stage 17 embryos lack septae between subneurial glial cells, explaining the disrupted blood-brain barrier.

cold^f05607/Df(2L)ED49 embryos develop elongated tracheal dorsal trunks and have disrupted tracheal paracellular barrier function (as assessed by dextran-dye injection).