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In this paper, the underestimated challenges of the burst-mode operation in the upstream path of time-and-wavelength-division-multiplexed passive optical networks are analyzed. Various challenges are disclosed, the influence of the associated physical effects on the signal quality is discussed and mitigation proposals are described: Intra-channel cross-talk can arise from optical network units induced background amplified-spontaneous emission noise and inter-channel cross-talk can be caused by non-ideal filter suppression between wavelength channels at the optical line termination receivers. Such cross-talk effects can be counteracted by reducing the laser burst-signal output power as well as its rival noise power simultaneously, i.e. by applying power leveling. A fast frequency drift in burst-mode operation is inherent to directly modulated lasers, but depends on the specific laser design, the laser output power and the burst length. Mitigation mechanisms are an optimized non-standard laser design or a specific mode of operation, e.g. an increase of the frequency drift during the preamble of the burst. The power dynamic range an optical pre-amplifier as part of the upstream signal receivers needs to handle can be in range of up to 40 dB, because of the multi-wavelength channel operation and of the optical distribution network differential path loss. This power dynamic faced at the receiver can not only cause challenges for the optical amplifier, but also for the burst-mode receivers. Additionally, the physical layer operation and maintenance of the upstream path is challenging too. Each optical network unit laser needs to be either wavelength pre-calibrated, which adds undesired costs or cross-channel synchronization of ranging windows has to be ensured. Otherwise rogue optical network unit behavior in wavelength and time domain will deteriorate system performance with every new optical network unit entering the network. Further, the operation of the optical network unit laser requires a wavelength fine tuning mechanism, which can depend on the receiver filter shape and on the received signal strength indicator implementation. [ABSTRACT FROM AUTHOR]