Citrus reticulata

HLB‐affected citrus shoots that used for dodder’s parasitizing were collected from two citrus growing provinces in China, i.e. Guangdong and Yunnan provinces, between March 2018 and December 2018. A total of four citrus cultivars were collected, including Shatangju (Citrus reticulata Blanco cv. Shatangju), Lemon (Citrus limon), Gongkan (Citrus reticulata Blanco cv. Gongkan) and Nianju (Citrus reticulata Blanco cv. Nianju) (Table 1, Table S1). The symptomatic citrus shoots were kept moisturized before bring back to the laboratory from citrus orchards. The bottom of citrus shoot, about 1 cm in length, was further cut out. The remaining stem of young shoot was maintained with the distilled water in a 15 ml tube immediately and fixed with cotton to keep the leaves in the air (Fig. 1A). Three citrus leaves from each citrus shoot were sampled for DNA extraction and used for PCR detection of CLas. Citrus shoots, which were detected as CLas‐positive and can be maintained in distilled water over three days with most leaves still attached, were further used for the CLas enrichment experiment.

In total, 128 consumers participated in the test corresponding to Harvest 1 and 139 in that corresponding to Harvest 2. For both tests, cultivars were assessed by evaluating acceptance and the CATA questions. Consumers were recruited from the IVIA, the Universidad Politécnica de Valencia (UPV) and the Instituto de Agroquímica y Tecnología de Alimentos (IATA-CSIC), and were selected according to their interest in participating. The whole group of participants was made up with administration staff and students; hence, they were not linked to food research field and can be considered naïve consumers. They were aged between 18 and 65, and the male/female ratio (%) was 43/57 and 46/54 for Harvests 1 and 2, respectively.
The evaluations were carried out in a standardized test room (ISO 8589; ISO 2007) [26 ]. Mandarins were served whole in cups coded with three-digit random numbers and were presented monadically following a Williams’ Latin square design [27 (link)], and consumers were provided with water to cleanse palate between samples. The consumers were instructed to first peel the mandarin, then taste it and score their overall acceptance on a nine-point hedonic scale ranging from 1 (“dislike extremely”) to 9 (“like extremely”). Afterwards, they were asked to answer the Check All That Apply questions, comprising of 23 descriptors. The descriptors were related to the peeling characteristics, flavor, taste and texture of mandarins. The descriptors were initially selected based on previous research [1 (link),2 (link)] and were adapted according to the specific characteristics of samples. To this end, a group of eight semi-trained panelists (people used to generate attributes) evaluated the samples in order to add attributes to the list. Finally, a group of eight consumers participated in a session to generate the final list of attributes that would be included in the CATA questions. They were given the samples, a list of the potential attributes and an outline of the procedure. The assessors were asked to taste the samples, and write down the most appropriate attributes to describe each one; they could use the terms in the list, but they were encouraged to suggest new terms that were relevant for describing the mandarins they tasted. All this information was used to gather a final list of attributes, in which, based on consumers descriptions, different intensities were included for some of the attributes. This design was previously described by Lado et al. [20 (link)], who reported that CATA questions may provide information about consumers’ perception of the intensity of sensory attributes of a food product, particularly fruit cultivars. In the final list, the attributes were ordered as they are likely to be perceived. The selected descriptors were: intense odor when starting to peel, hard starting to peeling, easy to peel, difficult to peel, stain hands when peeling, tasteless/dull, not very sweet, sweet, very sweet, overripe taste, not traditional/novel taste, refreshing taste, traditional mandarin taste, not very sour, sour, very sour, not very aromatic, very aromatic, very intense taste, soft, fibrous, juicy and juiceless.
Finally, when consumers had finished the mandarin’s assessment, they answered a few demographic questions such as gender, age and frequency of mandarin/orange consumption during the season. The response options for fruit consumption were as follows: “twice a week or more”, “once a week” or “2 or 3 times a month”.

To assess the effects of Meloidogyne spp. on root-associated microbiota, rhizosphere soil and root samples of healthy and parasitized plants were collected in Shunchang County of Fujian province, China (26° 38′–27° 121′ N, 117° 29′–118° 14′ E) in June 2016 (Additional Table S1). Samples of three vegetables, tomato (Solanum lycopersicum), lettuce (Lactuca sativa L. var. ramosa Hort.), and celery (Apium graveolens L.), were collected from a vegetable farm, monitored for RKN parasitism for at least 5 years before sample collection [67 ]. The field prevalence of RKN for the three crops were approximately 30–50%. Two perennial plants, Snakegourd fruit (Trichosanthes kirilowii Maxim.) and citrus (Citrus reticulata Blanco), attacked by RKN for at least 2 years (severe parasitism, with > 75% roots with galls, and swollen by > 75%), were separately collected from orchards with RKN. The collected lettuce and celery roots showed a low RKN parasitism symptom (less than one third roots with galls), and tomato root with a moderate RKN parasitism symptom (more than half of roots with galls) (Additional Table S1). At least three replicated healthy or nematode-parasitized plants were sampled for each plant species. The collected plants were used to separate rhizosphere soil and root samples for the 16S rRNA gene-based high-throughput sequencing and bacterial community analysis (Additional Table S1).

The 206 progeny individuals from five different F₁ mandarin populations growing at the University of Florida-IFAS Citrus Research and Education Center (Lake Alfred, FL) were used in this study (Figure 1). Mukaku Kishu (Citrus reticulata Blanco) (‘MK’), a completely seedless mandarin cultivar, was the common male parent in all the populations. All five maternal mandarin parents, ‘SB’ (Clementine mandarin × Minneola tangelo), ‘D’ [(Clementine mandarin × Orlando tangelo) × (Clementine mandarin × Ponkan mandarin)], Temple (‘T’) (a natural mandarin × sweet orange hybrid), Lee (‘L’) (Clementine mandarin × Orlando tangelo), and Clementine × Valencia orange (‘CVO’) produce fruit containing monoembryonic seeds. All of these, except ‘CVO’ are released commercial cultivars. To preclude any inadvertent inclusion of off types/nucellars in the mapping population, the hybridity of the population individuals (for ‘MK’ paternity) was verified through few homozygous SNPs polymorphic between the maternal parents and ‘MK’. The individuals with doubtful identity were not used in this study. All the populations were fruiting in the 2017-18 season.