Sex reversal in nature
It is well known that sex determination can be influenced by environmental or social parameters throughout evolution. For example, in Reptilia, fish and amphibia, temperature alone can push for sex reversal.
Social Sex Determination: fish can undergo sex reversal which depends on mating system for example: dominant female in sea bass harem changes into male when territorial male disappears. [Douglas Y. Shapiro, BULLETIN OF MARINE SCIENCE,31(2):383-398,1981]
Some reptiles use incubation temperatures to determine sex. In some species, this follows the pattern that eggs in extremely high or low temperatures become male and eggs in medium temperatures become female [“Temperature dependent sex determination”. Turtles of the World]
At the turn of the 20th century, biologists believed amphibian sex determination to be the result of interactions between genetic forces and environmental conditions (King, 1909, 1919; Witschi, 1929). Emerging theory suggests that sex reversal may be an important process for amphibian evolution (Perrin, 2009; Grossen, Neuenschwander & Perrin, 2012).
Sex reversal in birds
In chickens, sex determination takes place only after day 6 of embryonic development in which time gonads start to differentiate to ovaries or testes according to the chromosomal blueprint of the gametes. Until then, the gonads are “bi-potential”, meaning, they can differentiate into either testes or ovaries regardless of genome status29. “Soos treatment” exploits the “bi-potentiality” window of the gonads and by introducing sound vibration energy to the regular incubation profile induces up-regulation of feminine gene expression over masculine gene expression. As a result, genomic male chicks present female phenotype (ovaries) and function (laying eggs). [Okuno et. al, 2020, Scientific Reports 10:20073]
Acoustic impact on cells
Cell culture research was previously used to demonstrate the extensive effect of sound waves on cell growth, viability, differentiation, proliferation and functionality. For example, sound waves were used to promote the development of neural stem cells from the cochlear nuclei [Tao Xue et al, 2015] and viability and proliferation of human bone marrow-derived mesenchymal stem cells [Xi Chen et al, 2015]. Considering in-vitro results, raising a possible strong effect of acoustic vibrations may have on embryo and gonad development.
Immunofluorescence staining with antibodies against clusterin (CLU) in neural stem cells (NSCs) from the cochlear nuclei (protein expression of CLU). (A) Exposure to normal environment [about 40dB sound pressure level (SPL)]; (B) exposure to weakened acoustic environment (20dB SPL) for 7 days; (C) exposure to augmented acoustic environment (70dB SPL) for 7 days; (D) exposure to augmented acoustic environment (70dB SPL) for 12 days. Scale bar represents 50μm. [Tao Xue et al, 2015, Int. J. Mol. Med. 35, 637–644].
Effects of acoustic-frequency vibratory stimulation (AFVS) on cell proliferation of human bone marrow-derived mesenchymal stem cells (BM-MSCs) was improved by AFVS at 800Hz on day 7 when stimulated with AFVS at the frequencies of 0, 30, 400, and 800Hz. Values are mean ± standard error of six independent experiments (n= 6). *P < 0.05, **P < 0.01 in the indicated groups from unpaired t-test. #P < 0.05, ##P < 0.01 compared with the 0Hz control group at the same time point from unpaired t-test. [Xi Chen et al, 2015, BioMed Research International, https://doi.org/10.1155/2015/540731]
Discrimination of genomic males or females is done by amplification of a unique sequence that resides in a specific gene on W chromosome that has minimal homology of this gene on Z chromosome.
Amplification of this sequence is preformed using Real Time PCR machine with specific primers designed in-house. Detection achieved using SYBR Green Supermix (Bio-Rad).