To improve the efficiency of papaya anther culture, we investigated (1) hormonal medium conditions for inducing haploids or dihaploids; (2) identified the sex of established plantlets using a sex-specific DNA molecular marker and (3) estimated their ploidy by flow cytometry analysis of DNA content. Anthers with a mixture of uninucleate, mitotic, and binucleate microspores were collected from a male plant, and cultured on MS agar medium with different concentrations of CPPU and NAA. An embryo induction rate of 13.8% was attained on MS agar medium with 0.01 mg l −1 CPPU and 0.1 mg l −1 NAA. The induced embryos were subcultured on medium with 0.0025 mg l −1 CPPU. Rooting of the developed shoots was promoted by treating their basal parts with 1500 mg l −1 IBA in a 50% ethanol solution for about 10 seconds. All the embryo-derived plantlets (27 plants) were identified as female, implying that they were derived from microspores. In addition, 26 plants were determined to be triploids and one to be tetraploids. We also observed a wide range of morphological variation (e.g., in tree height and fruit size) among the established plants. Based on the results, we discussed a potential value of anther culture techniques for the breeding of papaya.

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Carica papaya L. is an important fruit crop grown by small-and large-scale farmers in Kenya for local and export markets. However, its production is constrained by papaya ringspot disease (PRSD). The disease is believed to be caused by papaya ringspot virus (PRSV). Previous attempts to detect PRSV in papaya plants showing PRSD symptoms, using enzyme-linked immunosorbent assay (ELISA) and reverse transcriptase-polymerase chain reaction (RT-PCR) procedures with primers specific to PRSV, have not yielded conclusive results. Therefore, the nature of viruses responsible for PRSD was elucidated in papaya leaves collected from 22 counties through Illumina MiSeq next-generation sequencing (NGS) and validated by RT-PCR and Sanger sequencing. Viruses were detected in 38 out of the 48 leaf samples sequenced. Sequence analysis revealed the presence of four viruses: a Potyvirus named Moroccan watermelon mosaic virus (MWMV) and three viruses belonging to the genus Carlavirus. The Carlaviruses include cowpea mild mottle virus (CpMMV) and two putative Carlaviruses-closely related but distinct from cucumber vein-clearing virus (CuVCV) with amino acid and nucleotide sequence identities of 75.7-78.1 and 63.6-67.6%, respectively, in the coat protein genes. In reference to typical symptoms observed in the infected plants, the two putative Carlaviruses were named papaya mottle-associated virus (PaMV) and papaya mild mottle-associated virus (PaMMV). Surprisingly, and in contrast to previous studies conducted in other parts of world, PRSV was not detected. The majority of the viruses were detected as single viral infections, while a few were found to be infecting alongside another virus (for example, MWMV and PaMV). Furthermore, the NGS and RT-PCR analysis identified MWMV as being strongly associated with ringspot symptoms in infected papaya fruits. This study has provided the first complete genome sequences of these viruses isolated from papaya in Kenya, together with primers for their detection-thus proving to be an important step towards the design of long-term, sustainable disease management strategies.

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