Diversity of mites in blackberry genotypes in Pelotas , Rio Grande do Sul , Brazil

Blackberry is part of the small-fruit group and the blackberry cultivation area is expanding as an important option for small farmers using the agroecological production system. Mites are prominent among the world’s major crop pests, as they can affect plants from growth to fruiting. The objective of this study was to evaluate the occurrence of phytophagous and predatory mites in different blackberry genotypes in the municipality of Pelotas, RS, Brazil. This study was carried out in the period Nov. 2015 Oct. 2016, totaling 14 samples. A total of 11,158 mites were collected [phytophagous (5940) and predatory (323) mites, and those with varied feeding habits (4895)], which were distributed in 8 families. The species Neotetranychus asper (Feres & Flechtmann, 2000), Diptacus rubuscolum (Trinidad, Duarte & Navia, 2018), and Acalitus orthomerus (Keifer, 1951) had highest occurrence. According to the faunistic classification, the N. asper phytophagous species is predominant in genotypes 128 and 178; whereas, D. rubuscolum is predominant in genotype 132. Monitoring of these genotypes is recommended to detect possible damage to the crop.


INTRODUCTION
Cultivation of the blackberry tree (Rubus spp., Rosaceae) is expanding in Brazil, and it has been used in an agroecological production system (ANTUNES et al., 2010) due to its great adaptability, low installation cost, and management.In Brazil, its cultivation began in the 1970s, with development in the breeding program (RASEIRA & FRANZON, 2012).The cultivars currently used in Brazil are the result of crosses of the native and North-American genetic material (ANTUNES et al., 2014).
Blackberry is subject to a considerable number of pests, such as with other crops, and they can affect the blackberry tree by affecting its production (PAGOT et al., 2007).Currently, pest management in Brazil is restricted and based only on working knowledge, since research-generated information is lacking (NAVA et al., 2007).Blackberry weevil (Eulechriops rubi) and fruit fly (Anastrepha spp.) were cited as the main pests in Brazil (NAVA et Ciência Rural, v.49, n.2, 2019.Trinidad et al. al., 2007;ANTUNES et al., 2014).In addition, the presence of mites has already been reported in Brazil, specifically in Rio Grande do Sul State (MARCHETTI & FERLA, 2011).However, specific studies are necessary to obtain information about the predominant mite species, their potential for harm, and how they could become a threat to commercial systems for blackberry production, as it has occurred with Acalitus essigi Hassan, 1928(KEIFER et al., 1982;BAKER et al., 1996;DAVIES et al., 2001a, PINA et al., 2012) or Acalitus orthomerus (Keifer, 1951) (BAKER et al., 1996) in other countries.
The study conducted by MARCHETTI & FERLA (2011) on blackberry cultivars (Brazo, Caingangue, and Tupy) in the municipality of Ilópolis (RS) showed that the occurrence of phytophagous mites of the families Diptilomiopidae and Tetranychidae was predominant and those of the Tenuipalpidae family was smaller in number.Among predatory mites, the families Stigmaeidae (Agistemus brasiliensis Matioli, Ueckermann & Oliveira, 2002) was the most abundant species) and Phytoseiidae (Typhlodromalus aripo (DeLeon, 1967) was the most prevalent species in the Caingangue cultivar) were outstanding.
Given this scenario, knowledge about mite diversity in various blackberry genotypes in the southern region of RS will provide additional information to further studies in other regions of this state.This knowledge, along with subsequent studies on Biology and Ecology of mites, will contribute to the management of potential pests in the crop.Therefore, the objective of this study was to determine the fauna of mites in blackberry genotypes in the southern region of Rio Grande do Sul.

MATERIALS AND METHODS
This study was carried out in the period Nov. 2015 -Oct.2016 in an experimental orchard of Embrapa Clima Temperado (latitude 31° 42'S, longitude 52° 24'O, and altitude de 57m), in the municipality of Pelotas, RS, Brazil.Blackberry genotypes 128, 132, 139, 164, 178, 181, and 198 and "Tupy" were used.All plants were arranged in parallel lines (50cm between plants and 3m between rows), without support system, except those of Tupy cultivar, which were supported by two wire strands and driven approximately in V. Five plants (replicates) per genotype were evaluated.In each crop and in the period of experimentation, the plants were kept without agrotoxic application and the cultural treatments were carried out according to the technical recommendations for the crop.
Samples were collected monthly from November to October, except in November and December (fruiting months), when they were collected biweekly (total: 14 samplings).Six leaflets (two in each of the upper, middle and lower thirds) were collected from each plant (total: 240 leaflets per sample).In the flowering and fruiting period, two flowers and four fruits (both from the upper and middle third) were collected per repetition, and 10 samples were collected per genotype (total: 80 flowers and 160 fruits) per collection season.Samples were placed in paper bags and packed in styrofoam boxes to be transported to the Acarology Laboratory (Federal University of Pelotas), where they were processed (under a Nikon SMZ 745 stereomicroscope, with the help of brushes and tweezers).Mites were identified in the same laboratory using a phase-contrast (Nikon E200) microscope.
In the laboratory, the leaves were observed (on the adaxial and abaxial surface) under a stereomicroscope, when the mites were collected and quantified.Regarding Eriophyoidea, mites were quantified in 1cm 2 area of the leaf (on the abaxial surface) by which it is united with the petiole.Fruits were evaluated in only half the sample, examining the regions between drupes and counting the number of specimens.Specimens collected from leaves, flowers, and fruits were transferred to Eppendorf flasks containing 70% alcohol solution.Then, they were mounted on slides containing Hoyer's medium, remaining in an oven (50 o C; 10-15 days), and adult mites were then identified and quantified.
The morphological characters observed on both surfaces of the leaves were described from samples of different parts of the plants.The presence or absence of acule and domatia (GONÇALVES & LORENZI, 2007) and trichomes and types of tricoma and indument (BEENTJE, 2016) were evaluated according to the literature.Leaf trichome densities were evaluated by comparing genotypes.This morphological description was performed at the Botanical Laboratory (Embrapa Clima Temperado).
Results were analyzed using the AnaFau software (MORAES et al., 2003).The faunistic indexes for dominance, abundance, frequency, constancy, and Shannon-Wiener diversity (H') were calculated to characterize the mite fauna in different blackberry genotypes.

RESULTS AND DISCUSSION
The occurrence of 12 species of mites distributed in eight families was recorded (Table 1), totaling 11,158 mites classified as phytophagous (5940), predatory (323), and miscellaneous-feeding ( 4895) mites.In analyzing mite distribution per family, the Tydeidae, Diptilomiopidae, Tetranychidae, and Eriophyidae families showed a higher representability.The predominance of certain mite families may be associated with their preference for the host plant, favoring their establishment and development.In addition, the presence of the tetranychid Neotetranychus asper (Feres & Flechtmann, 2000) and tydeid Tydeus californicus (Banks, 1904) was observed in all plant genotypes.Conversely, the eriophyoid Diptacus rubuscolum (Trinidad, Duarte & Navia, 2018) showed a relatively-high population in all plant genotypes in the months of November, December, and January of 2016; although, this species has only been described recently by TRINIDAD et al. (2018).However, the eriophid Acalitus orthomerus (Keifer, 1951) occurred in drupes in greater quantity in Dec. 2015 and Jan. 2016, when fruits reach maturation and are harvested; although, it has not been sampled in leaves, floral buds, or open flowers.This has led to a drastic reduction in mite population, which is also related to the significant seasonal changes in host plant morphology (DAVIES et al., 2001b) and it is good if the decrease in the number of mites in the plant is considered.Families/Species -------------Individuals------------  Significant differences were observed between the number of mites in each genotype evaluated (Table 2).The N. asper had a similar frequency in the plant genotypes 139, 164, 181, and 198 and "Tupy".However, genotype 164 and "Tupy" were the most favorable for the tetranychid mite Oligonychus yothersi (McGregor, 1915).Diptacus rubuscolum, Polyphagotarsonemus latus (Banks, 1904), and Brevipalpus spp.(Donnadieu, 1875) showed no significant difference in the evaluated genotypes and "Tupy".Acalitus orthomerus had a similar occurrence in all genotypes (except genotype 128) and "Tupy".Tydeus californicus occurred similarly in genotypes 132, 139, 164, 178, and 198 and "Tupy".Few mites Euseius mesembrinus (Dean, 1957) were found mostly in genotype 128 and "Tupy".Phytoseiid mites Amblyseius operculatus (De Leon, 1967), Galendromus (Galendromus) annectens (De Leon, 1958), and Typhlodromalus aripo (De Leon, 1967) did not differ in the number of individuals in each genotype and "Tupy".
Regarding the number of occurrences, N. asper, D. rubuscolum, and T. californicus showed similar amounts in genotypes 128 and 164 (Table 2).In genotype 132, the observed number of D. rubuscolum and T. californicus was similar.Tydeus californicus showed difference in relation to other mite species reported in genotypes 139 and 178.Genotype 181 showed no difference in the number of N. asper, D. rubuscolum, A. orthomerus, P. latus, Agistemus spp., and T. californicus.Occurrence of N. asper, D. rubuscolum, A. orthomerus, and T. californicus did not differ in the number of individuals in genotype 198 and "Tupy".
According to the faunal analysis (Tables 3 and 4), variation in mite distribution and behavior was observed in various genotypes.This may be related to the differences between genotypes regarding morphological characters on the adaxial surface of the leaf (Table 5).In genotypes 128 and 178, N. asper, D. rubuscolum, and T. californicus were considered prevailing, as they reached the maximum rates in faunal classification (Tables 3  and 4).For genotype 128, only the E. mesembrinus predatory mite showed to be dominant, very abundant, very frequent, and constant.This may suggest that the complex of phytoseiid mites found in the present study was not sufficient to regulate the number of these species of pest mites.
The same species was also reported by MARCHETTI & FERLA (2011) and occurred in Brazos and Tupy cultivars.Mites of the genus Euseius are classified as generalist predators, for which pollen is an important part of the diet, increasing their reproductive potential after flowering (MCMURTRY et al., 2013).

Mite families and species
Trinidad et al. (MCMURTRY et al., 2013), is probably linked to the presence of O. yothersi, which was also reported in the present study.Agistemus spp.(Sumers, 1960) was classified as common and frequent in almost all (128, 132, 139, 164, 178, and 181) genotypes and "Tupy".Agistemus brasiliensis was reported in the Caingangue blackberry cultivar (MARCHETTI & FERLA, 2011) and cited as a potential predator of the mite vector of citrus leprosis (MATIOLI & OLIVEIRA, 2007).
Diptacus rubuscolum and T. californicus reached maximum classification values in genotype 132, as superdominant, superabundant, super frequent, and constant, without significant presence of predators.
Acalitus orthomerus reached high faunal rates in fruits of genotypes 128, 132, and 139, as well as of genotype 178, and this was its first record in Brazil.This deserves more attention due to the potential damage of blackberry because it was reported to cause blisters and warts as a result of its large-scale feeding from buds.In addition, high infestation by A. orthomerus resulted in galls that do not allow budding (BAKER et al., 1996); although, mite-associated symptom was not observed in plants during the study period.It should be noted that symptoms of mite attack were observed outside the sampling period, but such symptoms corresponded to those reported for A. essigi, i.e., those that cause the so-called "redberry" symptom of mulberry (KEIFER et al., 1982;BAKER et al., 1996;DAVIES et al., 2001a).In Portugal, this mite is known as the "redberry mite" (FERREIRA & PINA, 2012).
Thus, they are not enough to confirm and consider that it is a pest, compared to other mites in other cultures, despite a relatively high number of eriophid mites was found.
In "Tupy", T. californicus and A. asper were classified as dominant, very abundant, very frequent, and constant; whereas, A. orthomerus was present in less than 50% of the samples.However, this is important for the symptoms they could cause on fruits.
In the study of blackberry genotypes, the values for the diversity index (H) of mites varied in the range 0,83-1,79.The maximum value for H was reached in genotype 181 (Table 4).MARCHETTI & FERLA (2011) studied the diversity and fluctuation of mite population in blackberry cultivars and obtained a variable diversity index in the Caingangue (0.42) and Tupy (1.29) cultivars.

CONCLUSION
Twelve mite species are distributed in eight families in the blackberry orchard (Municipality of Pelotas, RS).The phytophagous N. asper and D. rubuscolum are predominant in blackberry genotypes 164 and 128, respectively.Therefore, future studies should focus on the characterization of damage caused by these mites.

Table 2 -
Average number of mites per plant collected in blackberry orchard in the period Nov 2015 -Oct.2016, in the municipality of Pelotas, RS.

Table 3 -
Faunal analysis of adult mites collected in blackberry orchard in the period Nov. 2015 -Oct 2016, in the municipality of Pelotas, RS.

Table 4 -
Faunal analysis of adult mites collected in blackberry orchard in the period Nov 2015 -Oct 2016, in the municipality of Pelotas, RS.
VINCENT et al. (2010)or the species.In blackberry genotype 164, only the N. asper mite species was predominant.In blackberry genotype 181, D. rubuscolum mite, in addition to the tetraniquid N. asper mite, was classified as dominant, very abundant, very frequent, and constant during the evaluation period.In blackberry genotypes 181 and 198, the tarsonemid P. latus mite was classified as dominant, very abundant, and very frequent.Although, this is the first report on its occurrence in blackberry in Brazil, it is considered a cosmopolitan species.VINCENT et al. (2010)reported the occurrence of the white mite in organic blackberry production causing the ripple symptom in the leaves.It was identified in vine crops (Bento Gonçalves, RS), occurring mainly in new vine leaves after green pruning (