First Report of the Root-Knot Nematode (Meloidogyne javanica) Infecting Hops (Humulus lupulus) in Florida, USA

Humulus lupulus (Cannabaceae), commonly referred to as hops, are perennial, herbaceous climbing plants, native to temperate northern climates. Hops are cultivated for their strobiles or cones, which are often used for flavoring and aroma in food, tea, and beer (Almaguer et al. 2014; Burgess 1964). Because of the high demand for hops from the micro-brewing industry in the Tampa-St. Petersburg area, it has recently been introduced in Florida. The crop grows rapidly in the early spring to late summer. Plants reach a mature height of 18-25 feet in one year and produce cones from mid-summer to early fall (Pearson, 2016). Hop rhizomes were planted in April 2016 at the Gulf Coast Research Station, Wimauma, Hillsborough Co., Florida, USA. In October 2016, several hop plants that exhibited yellowing leaves and stunted growth were uprooted and showed severe root galling (Figs.1;2). Rhizosphere soil samples were collected for nematode extraction and showed high numbers of root-knot nematode second-stage juveniles (J2) (up to 1500 J2/ 200cm3 soil). Heavily galled root samples were sent to the Florida Department of Agriculture and Consumer Services, Division of Plant Industry Nematology Laboratory in Gainesville, FL. Species identification was performed using morphological analyses of females perenal patterns (n=22), selected characters of second-stage juveniles (n=17), and isozyme phenotypes (esterase and malate dehydrogenase) of egg- laying females (n=26) extracted from the roots. Configuration of the perineal patterns, morphometrics of body, stylet and tail length of J2 and the esterase phenotype (EST= J3), which is species-specific and malate phenotype (MDH=N1), are consistente with those reported in the original descrition of M. javanica and many other populations of this nematode species collected in Florida and other countries (Brito et al., 2008; Cofcewick et al., 2005; Carneiro et al., 2004; Jepson, 1987; Esbenshade and Triantaphyllou, 1985). For molecular analyses, DNA was extracted from individual females and mitochondrial DNA was amplified with MORF (5?- ATC GGG GTT TAA TAA TGG G - 3?) and MTHIS (5? - AAA TTC AAT TGA AAT TAA TAG C -3?) primer set (Pagan et al., 2015; Stanton et al., 1997). A fragment of approximately 740 bp was produced, which has been reported for M. incognita and M. javanica found in Florida (Baidoo et al., 2016). To further confirm the species identification we use the speices-specific SCAR primer set Fjav (5 ?- GGT GCG CGA TTG AAC TGA GC - 3?) and Rjav (5? - CAG GCC CTT CAG TGG AAC TAT AC - 3?) (Zijlstra et al., 2000). This primer set yield a fragment of of approximately 670bp, which is identical to that previously reported for M. javanica (Humphreys-Pereira et al., 2017; Baidoo et al., 2016; Zijlstra et al., 2000). Additionally, NADH dehydrogenase subunit 5 gene was amplified using NAD5F2 (5?- TAT TTT TTG TTT GAG ATA TAT TAG - 3?) and NAD5R1 (5?- CGTGAATCTTGATTTTCCATTTTT-3?) primers as described by Janssen et al. (2016). The GenBank accession number of the nad5 gene sequence is MH230176. The obtained the nad5 gene sequence was identical to the reference sequence of M. javanica provided by Janssen et al. (2016). To our knowledge, this is the first report of H. lupulus as a host of the Javanese root-knot nematode (M. javanica) in Florida.Fil: Brito, J. A.. University of Florida; Estados UnidosFil: Subotini, S.A.. University of California; Estados UnidosFil: Desaeger, J.. University of Florida; Estados UnidosFil: Achinelly, Maria Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Centro de Estudios Parasitológicos y de Vectores. Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Centro de Estudios Parasitológicos y de Vectores; ArgentinaFil: Qiu, Sai. University of Florida; Estados Unido

First Report of the Root-Knot Nematode (Meloidogyne javanica) Infecting Hops (Humulus lupulus) in Florida, USA

Humulus lupulus (Cannabaceae), commonly referred to as hops, are perennial, herbaceous climbing plants, native to temperate northern climates. Hops are cultivated for their strobiles or cones, which are often used for flavoring and aroma in food, tea, and beer (Almaguer et al. 2014; Burgess 1964). Because of the high demand for hops from the micro-brewing industry in the Tampa-St. Petersburg area, it has recently been introduced in Florida. The crop grows rapidly in the early spring to late summer. Plants reach a mature height of 18-25 feet in one year and produce cones from mid-summer to early fall (Pearson, 2016). Hop rhizomes were planted in April 2016 at the Gulf Coast Research Station, Wimauma, Hillsborough Co., Florida, USA. In October 2016, several hop plants that exhibited yellowing leaves and stunted growth were uprooted and showed severe root galling (Figs.1;2). Rhizosphere soil samples were collected for nematode extraction and showed high numbers of root-knot nematode second-stage juveniles (J2) (up to 1500 J2/ 200cm3 soil). Heavily galled root samples were sent to the Florida Department of Agriculture and Consumer Services, Division of Plant Industry Nematology Laboratory in Gainesville, FL. Species identification was performed using morphological analyses of females perenal patterns (n=22), selected characters of second-stage juveniles (n=17), and isozyme phenotypes (esterase and malate dehydrogenase) of egg- laying females (n=26) extracted from the roots. Configuration of the perineal patterns, morphometrics of body, stylet and tail length of J2 and the esterase phenotype (EST= J3), which is species-specific and malate phenotype (MDH=N1), are consistente with those reported in the original descrition of M. javanica and many other populations of this nematode species collected in Florida and other countries (Brito et al., 2008; Cofcewick et al., 2005; Carneiro et al., 2004; Jepson, 1987; Esbenshade and Triantaphyllou, 1985). For molecular analyses, DNA was extracted from individual females and mitochondrial DNA was amplified with MORF (5?- ATC GGG GTT TAA TAA TGG G - 3?) and MTHIS (5? - AAA TTC AAT TGA AAT TAA TAG C -3?) primer set (Pagan et al., 2015; Stanton et al., 1997). A fragment of approximately 740 bp was produced, which has been reported for M. incognita and M. javanica found in Florida (Baidoo et al., 2016). To further confirm the species identification we use the speices-specific SCAR primer set Fjav (5 ?- GGT GCG CGA TTG AAC TGA GC - 3?) and Rjav (5? - CAG GCC CTT CAG TGG AAC TAT AC - 3?) (Zijlstra et al., 2000). This primer set yield a fragment of of approximately 670bp, which is identical to that previously reported for M. javanica (Humphreys-Pereira et al., 2017; Baidoo et al., 2016; Zijlstra et al., 2000). Additionally, NADH dehydrogenase subunit 5 gene was amplified using NAD5F2 (5?- TAT TTT TTG TTT GAG ATA TAT TAG - 3?) and NAD5R1 (5?- CGTGAATCTTGATTTTCCATTTTT-3?) primers as described by Janssen et al. (2016). The GenBank accession number of the nad5 gene sequence is MH230176. The obtained the nad5 gene sequence was identical to the reference sequence of M. javanica provided by Janssen et al. (2016). To our knowledge, this is the first report of H. lupulus as a host of the Javanese root-knot nematode (M. javanica) in Florida.Centro de Estudios Parasitológicos y de Vectore

First Report of the Root-Knot Nematode (Meloidogyne javanica) Infecting Hops (Humulus lupulus) in Florida, USA

Humulus lupulus (Cannabaceae), commonly referred to as hops, are perennial, herbaceous climbing plants, native to temperate northern climates. Hops are cultivated for their strobiles or cones, which are often used for flavoring and aroma in food, tea, and beer (Almaguer et al. 2014; Burgess 1964). Because of the high demand for hops from the micro-brewing industry in the Tampa-St. Petersburg area, it has recently been introduced in Florida. The crop grows rapidly in the early spring to late summer. Plants reach a mature height of 18-25 feet in one year and produce cones from mid-summer to early fall (Pearson, 2016). Hop rhizomes were planted in April 2016 at the Gulf Coast Research Station, Wimauma, Hillsborough Co., Florida, USA. In October 2016, several hop plants that exhibited yellowing leaves and stunted growth were uprooted and showed severe root galling (Figs.1;2). Rhizosphere soil samples were collected for nematode extraction and showed high numbers of root-knot nematode second-stage juveniles (J2) (up to 1500 J2/ 200cm3 soil). Heavily galled root samples were sent to the Florida Department of Agriculture and Consumer Services, Division of Plant Industry Nematology Laboratory in Gainesville, FL. Species identification was performed using morphological analyses of females perenal patterns (n=22), selected characters of second-stage juveniles (n=17), and isozyme phenotypes (esterase and malate dehydrogenase) of egg- laying females (n=26) extracted from the roots. Configuration of the perineal patterns, morphometrics of body, stylet and tail length of J2 and the esterase phenotype (EST= J3), which is species-specific and malate phenotype (MDH=N1), are consistente with those reported in the original descrition of M. javanica and many other populations of this nematode species collected in Florida and other countries (Brito et al., 2008; Cofcewick et al., 2005; Carneiro et al., 2004; Jepson, 1987; Esbenshade and Triantaphyllou, 1985). For molecular analyses, DNA was extracted from individual females and mitochondrial DNA was amplified with MORF (5?- ATC GGG GTT TAA TAA TGG G - 3?) and MTHIS (5? - AAA TTC AAT TGA AAT TAA TAG C -3?) primer set (Pagan et al., 2015; Stanton et al., 1997). A fragment of approximately 740 bp was produced, which has been reported for M. incognita and M. javanica found in Florida (Baidoo et al., 2016). To further confirm the species identification we use the speices-specific SCAR primer set Fjav (5 ?- GGT GCG CGA TTG AAC TGA GC - 3?) and Rjav (5? - CAG GCC CTT CAG TGG AAC TAT AC - 3?) (Zijlstra et al., 2000). This primer set yield a fragment of of approximately 670bp, which is identical to that previously reported for M. javanica (Humphreys-Pereira et al., 2017; Baidoo et al., 2016; Zijlstra et al., 2000). Additionally, NADH dehydrogenase subunit 5 gene was amplified using NAD5F2 (5?- TAT TTT TTG TTT GAG ATA TAT TAG - 3?) and NAD5R1 (5?- CGTGAATCTTGATTTTCCATTTTT-3?) primers as described by Janssen et al. (2016). The GenBank accession number of the nad5 gene sequence is MH230176. The obtained the nad5 gene sequence was identical to the reference sequence of M. javanica provided by Janssen et al. (2016). To our knowledge, this is the first report of H. lupulus as a host of the Javanese root-knot nematode (M. javanica) in Florida.Centro de Estudios Parasitológicos y de Vectore

Plant-Parasitic Nematodes Associated with Cannabis sativa in Florida

The subtropical climate of Florida allows for a wide range of crops to be grown. With the classification of hemp (Cannabis sativa L., <0.3% delta-9-tetrahydrocannabinol) as an agricultural commodity, hemp has become a potential alternative crop in Florida. Hemp cultivars of different geographies (Europe, China, and North America), and uses (fiber, oil and CBD), were evaluated in three field experiments. The field experiments evaluated a total of 26 cultivars and were conducted for two consecutive seasons at three different locations (soil types) in North (sandy loam), Central (fine sand), and South Florida (gravelly loam). Nematode soil populations were measured at the end of each season. A diverse population of plant-parasitic nematodes was found, with reniform nematodes (RN, Rotylenchulus reniformis) the dominant species in North and South Florida (up to 27.5 nematodes/cc soil), and RKN (Meloidogne javanica) the main species in central Florida (up to 4.7 nematodes/cc soil). Other nematodes that were commonly found in south Florida (and to a lesser extent north Florida) were spiral (Helicotylenchus spp.), stunt (Tylenchorhynchus spp.) and ring nematodes (Criconemoids), while in central Florida, stubby root (Nanidorus minor) and sting nematodes (Belonolaimus longicaduatus) were found. No significant difference among hemp cultivars was noted at any of the locations. RKN were found in all three regions and soils, while RN were only found in North and South Florida. This is the first report on plant-parasitic nematodes associated with hemp in Florida fields. Natural nematode populations varied greatly, depending on where in Florida hemp was grown. Growers who wish to include hemp in their crop rotation need to be aware of potential pest pressure from nematodes. More research is needed to determine to what extent nematodes, especially RKN and RN, can reduce hemp growth and yield

First Report of Direct Damage Caused by the Stubby-Root Nematode, Nanidorus minor, to Strawberry (Fragaria x ananassa), in Florida

In 2019–2022, declining symptoms were observed in two commercial strawberry farms in Hillsborough County, Florida. The fields in the two farms consisted of raised beds covered by plastic mulch. Both were fumigated with a mixture of 1,3-dichloropropene (40%) + chloropicrin (60%) before planting. Samples collected from large patches with declining plants were infested with stubby-root nematodes. No sting and root-knot nematode species were detected. The results of morphological and molecular analyses indicated that the stubby-root nematode populations were representative of the species Nanidorus minor. The two cultivars ‘Florida Brilliance’ and ‘Florida Sensation’ in the two fields included plants with stubby root symptoms showing a reduction in the size of the root system and arrested growth and elongation of the feeder roots on the first strawberry crop. The nematode population densities in the two fields increased at the end of strawberry season and averaged 66 and 96 specimens in 200 cm3 soil. In one of the fields, a second strawberry crop was established as in the previous year using the same practices (fumigation and raised beds covered with plastic). However, in this field the population of N. minor declined and did not reach damaging levels at the end of the season on the second strawberry crop. The factors causing the decline of the nematode population were not elucidated. This is the first report of a direct damaging effect of N. minor to strawberry

First Report of Meloidogyne javanica Infecting Strawberry (Fragaria × ananassa) in the United States

Strawberry (Fragaria × ananassa) is native to temperate regions. However, it has been produced in tropical areas, as a seasonal crop including in Florida, USA during the winter months. In March 2022, root galls resembling those induced by root-knot nematodes (Meloidogyne spp.) were observed in declining strawberry plants ‘WinterstarTM FL 05-107’ growing in an organic-certified research site in Hillsborough County, Florida, USA. To our knowledge, M. hapla is the only root-knot species reported to infect strawberry in Florida. Preliminary molecular analyses, including newly synthesized DNA sequences (TW81/AB28 = OQ469833 - OQ469836; D2A/D3B= OQ473043 - OQ473047) using extracted nematode females from the strawberry roots, initially identified the RKN as M. javanica. Nematode species confirmation was further performed using the morphology of the female perineal patterns and isozyme analysis, mainly esterase (EST) and malate dehydrogenase (MDH), DNA sequencing, (NAD5-F/NAD5-R) and the SCAR primer set (Fjav/Rjav), species-specific for M. javanica. Isozyme analyses, EST= J3, which is specific for M. javanica and MDH=N1, as well as the morphology of female perineal patterns, agreed with data previously reported for M. javanica. A pathogenicity test on strawberry ‘WinterstarTM FL 05-107’ transplants was performed using 10,000 eggs of the original M. javanica population, which induced galls on strawberry plants (Gall index, GI = 4.1) with egg masses clearly visible outside of the roots, producing an average of 1,344 eggs/gram of fresh root and 9,201 ± 4,206 eggs/root system. No galls or egg masses were observed on non-inoculated plants. Tomato ‘HM 1823’ was used as a control for the viability of the inoculum and showed numerous galls and egg masses (GI=5.0;)