First Report of Alternaria alternata Causing Leaf Spot on Kidney Bean in China

Kidney bean (Phaseolus vulgaris), also being known as common bean, dry bean or french bean, is one of the most precious and highly nutritious legume crop cultivated and consumed worldwide(Blair et al.,2012; Choudhary et al.,2018) , which is an important edible foods or one of the most economically important vegetable crops in China. It is widely grown in the Heilongjiang Province in China. In July of 2020, leaf spot symptoms were found on the old or new leaves of Kidney bean plants (Phaseolus vulgaris L.) in our experimental fields located in Zhaozhou County(N45°42 '20.16 ", E125°15' 58.63" ), Daqing City, Heilongjiang Province, China. This field had disease incidences of approximately 20%. The leaf spot is conducive to the onset at high temperature and humidity environment, and this disease spreads very quickly after rainy days, therefore it is potentially a large risk for the development of Kidney bean industry. In its early occurrence phase, the infected leaves showed yellowish halo on the leaves, in which the middle mesophyll lost green. Thereafter, the yellow halo turned brown, and the middle leaf tissue of the halo appeared brown, ultimately the whole leaves had many brown spots (Supplementary Figure S1). To isolate the pathogen, diseased tissue (5×5 mm) was excised from the margins of individual lesions from the leaves of diseased plants with typical symptoms, and was disinfected with 75% ethanol for 10s followed by 2% NaClO for 3 min and then washed five to eight times with sterile water. Afterwards, the samples were transferred to potato dextrose agar (PDA) plates and incubated. After 5 to 7 days of incubation at 25°C (Wei et al.,2018), the mycelia were dark green with white margins in obverse and dark in reverse. Conidiophores were light brown with 2 to 4 septa and obclavate, 17.5to 44.0 × 6.5 to 14.5μm, with a short beak, and with 1 to 5 transverse septa and 0 to 2 longitudinal septa, light brown to olive-brown (Supplementary Figure S2). Based on morphological features and sporulation pattern, the pathogen was similar to characteristics described Alternaria alternate (Zhou et al,2014), being identified as A. alternata. To confirm pathogenicity, the isolates were cultured on PCA for 7days to prepare conidial suspensions, then being produced a final concentration of 1×108 spores/ml. Five potted Kidney bean plants were sprayed with conidial suspensions, and five control potted plants were sprayed with sterile distilled water, in which these potted Kidney bean plants were treated after wiping each leaf surface with 75% ethanol and washing each leaf with sterilized distilled water five times. These plants were incubated in an artificial growth chamber at 26 to 28°C with a 12 h light/dark photoperiod, with 85% relative humidity. After 3 days, yellowish halo lesions appeared on the inoculated plants, and pale lesions with distinct dark brownish red borders on Kidney bean leaves were observed after eight days, but no lesions were observed on the control leaves. Pathogenicity tests were repeated three times. The internal transcribed spacer (ITS) region of rDNA was amplified and sequenced with primers ITS1/ITS4. BLAST analysis of the sequences showed 100% sequence identity with a pathogenic A. alternata (Fr.) Keissl (Supplementary Figure S3), and the nucleotide sequence of the ITS region was submitted to GenBank under accession MZ951052. In China, there are no detailed records about the causal agent of this disease on Kidney bean in a paper in Chinese. To our knowledge, this is the first confirmed report of leaf spot causing by A. alternata on Kidney bean in China.

Characterstic And Inhibitory Activity Of Green And Red Beans Yoghurt Enriched With Honey Against Methicillin-Resistant Staphylococcus Aureus

Currently probiotics are widely developed because of their health benefits such as helping to maintain a healthy digestive tract. One of the probiotic fermented products is yogurt. Yogurt on the market is made from cow's milk, so it cannot be consumed by people who are allergic to cow's milk or lactose intolerant. The purpose of this study was to develop a yogurt formulation based on vegetable juice, namely mung bean and kidney bean juice. The characterization of mung bean and kidney bean juice yoghurt was carried out by of organoleptic observation, fat content test, protein content test, ash content test, acidity test, total plate number (ALT) test and antibacterial activity test against Methicilli-resistant Staphylococcus Aureus (MRSA) by the agar diffusion method using wells. The results of the product characterization based on mung bean juice and kidney bean enriched with honey meet the quality characteristics of SNI yogurt (2981:2009), including the appearance of thick-solid liquid, normal/typical odor, sour/typical taste, homogeneous consistency. Fat content test, the average F1 was 1.063% (SD = 0.0026), F2 1.025% (SD = 0.0031), F3 1.035 (SD = 0.0033); the average protein content of F1 12.51% (SD=0.01244), F2 11.90% (SD=0.0120), F3 12.08% (SD=0.02); mean ash content of F1 0.36% (SD= 0.0410), F2 0.52% (0.0821), F3 0.39% (SD 0.0294); the average acid content of F1 0.858% (SD= 0.0084), F2 0.873% (0.0199), F3 0.936 (SD= 0.0336); the number of starter bacteria on average was 5.4 x 109 CFU/mL (SD= 0.927), F2 4.3 x 109 CFU/mL, F3 5.6 x 109 CFU/mL. The antibacterial activity of the yogurt was included the strong category, the inhibition zone produced more than 20 mm. The average inhibition zone by F1 was 21.667 mm (SD= 0.489), F2 22.667 mm (SD= 0.489), F3 22 mm (0.4).

First Report of Leaf Spot on Kidney Bean Caused by Nigrospora oryzae in China

Kidney bean (Phaseolus vulgaris L.) is a legume with high nutritional and economic value. This vegetable crop is widely cultivated in China, providing a year-round supply of young edible pods. In July 2020, a leaf spot disease on kidney bean cultivar ‘Dabailong’ was observed on a two-hectare field in Longli County (26°16′15.66″ N, 106°48′12″ E), Guizhou Province, China. Disease incidence was estimated to be nearly 50%. Foliar symptoms manifested as black circular spots, surrounded by a yellow halo and accompanied by white mycelium. To identify the pathogen, small portions of tissue (5×5 mm) from margins of leaf spots were cut from 20 symptomatic leaves, surface disinfected with 75% ethanol for 30 s, rinsed two times with sterile distilled water, dried on a sterile filter paper, and incubated on potato dextrose agar (PDA) at 28°C for 3 days. A total of 39 single-spore isolates were obtained. The colonies on PDA were fluffy, changing from white to gray or black with age, and reaching 7-cm diameter in 5 days at 28°C. Conidia were black, globose to subglobose, smooth, solitary, measuring 13.0 to 16.0 × 10.5 to 16.0 µm (n=30). Morphological characteristics were consistent with Nigrospora oryzae. In addition, the rDNA internal transcribed spacer (ITS), large subunit (LSU), β-tubulin (TUB) and translation elongation factor 1-alpha (TEF1) loci were amplified by PCR and sequenced (White et al. 1990, Glass and Donaldson 1995, O'Donnell et al. 1998; Carbone and Kohn 1999). The ITS, LSU, TUB and TEF1 sequences of two isolates, GUCC19-5105 and GUCC19-5192, were submitted to GenBank. BLASTn analysis of these sequences showed >98% homology with those of N. oryzae strain LC 7293 in GenBank (ITS, 99.80% (MZ145361 vs KX985931 – 498/499 bp) and 99.62% (MZ148445 vs KX985931 – 525/527 bp); LSU, 100% (MZ146317 vs KY806236 – 837/837 bp) and 99.76% (MZ148446 vs KY806236 – 847/849 bp); TUB, 98.72% (MZ329335 vs KY019601 – 386/391 bp) and 98.67% (MZ329337 vs KY019601 – 373/378 bp) and TEF1, 98.91% (MZ329336 vs KY019396 – 452/457 bp) and 98.89% (MZ329334 vs KY019396 – 444/449 bp) respectively). The phylogenetic tree of the combined 4 sequences showed that both isolates clustered with N. oryzae. Based on morphological characteristics and the multigene phylogenetic analysis, GUCC19-5105 and GUCC19-5192 isolates were identified as N. oryzae. Pathogenicity tests were performed twice by spraying conidial suspension (1×105 conidia/mL) of the two isolates (GUCC19-5105 and GUCC19-5192) on leaves of ten (five per isolate) healthy 5-week-old kidney bean cultivar ‘Dabailong’ plants. Two plants sprayed with sterile water served as controls. After inoculation, all the plants were kept moist in plastic bags for 24 hours and incubated in a greenhouse at 25°C for 20 days. Leaf spots similar to those observed in the field were observed 20 days post inoculation, but no lesions were observed on control plants. N. oryzae was reisolated from the infected tissues of inoculated kidney bean plants and the identity of the reisolated pathogen was confirmed as N. oryzae through morphology and sequencing ITS, LSU, TUB and TEF1 loci. In recent years, N. oryzae has been reported to infect a variety of plants such as Aloe vera, Citrullus lanatus and Costus speciosus (Begum et al. 2018; Chen et al. 2019; Sun et al. 2020). To our knowledge, this is the first report of leaf spot disease on kidney bean caused by N. oryzae in the world and provides a basis for diagnosticians and researchers to identify the disease and develop disease management strategies.

Life Table and Preference Choice of Frankliniella occidentalis (Thysanoptera: Thripidae) for Kidney Bean Plants Treated by Exogenous Calcium

Exogenous calcium (Ca) has been used to induce host plant resistance in response to abiotic and biotic stresses, including from thrips attack. The aim of this study was to determine whether exogenously applied Ca affects the performance of Frankliniella occidentalis. We assessed the development time, total longevity, reproduction, and population parameters of F. occidentalis, and its preference choice on Ca-treated or untreated control kidney bean plants under laboratory conditions. The results showed that F. occidentalis fed on Ca-treated leaves had a longer developmental time but lower longevity (female and male) and fecundity than F. occidentalis fed on control leaves. Population parameters, including the intrinsic rate of increase (r), finite rate of increase (λ), and net reproductive rate (R0), were all found higher in control leaves than in Ca-treated leaves, and the mean generation time (T) was shorter. In preference choices, the number of thrips on control plants was higher than the number of thrips on Ca-treated kidney bean plants. Overall, our results indicated that exogenous Ca pretreatment on kidney bean plants affected the life history and preference choice of F. occidentalis, suggesting Ca might be used as a promising elicitor of inducible plant defense against thrips.

Age-stage, two-sex life table analysis of Spodoptera frugiperda (JE Smith) (Lepidoptera: Noctuidae) reared on maize and kidney bean

Background Spodoptera frugiperda (JE Smith), or fall armyworm, is one of major migratory agricultural pests with a wide range of hosts. The effect of different hosts (maize and kidney bean) on the growth and reproduction of S. frugiperda were investigated using the age-stage, two-sex life table method. Results The results showed that S. frugiperda could complete its entire life cycle on both hosts, albeit with significantly different development and reproduction. The durations of larval and pupal development were significantly prolonged whereas adult lifespan was shortened on kidney bean compared to maize. The differences of survival rates at each instar, prepupal and pupal stages between the two hosts were not statistically significant. The total pre-oviposition period was longer on kidney bean than that on maize (42.05 vs 39.04 days), but there was no difference in the oviposition rate (64.77 on kidney bean vs 62.48 on maize). The differences of net reproductive rate, intrinsic rate of increase and finite rate of increase were nonsignificant, while the mean generation time on kidney bean (42.05 days) was significantly longer than that on maize (40.92 days). Conclusions The results indicate that S. frugiperda can grow and reproduce normally on both hosts, although maize is more suitable. Nonetheless, the data show that this pest is harmful when the population density is high or when the preferred host is scarce. Graphic abstract

INHIBITING ACTIVITY OF ANGIOTENSIN CONVERTING ENZYME-1 BY BEAN PROTEIN HYDROLYSATE GENUS PHASEOLUS

Kidney bean (Phaseolus.vulgaris L. Chitra) and lima bean (Phaseolus lunatus L. Sweet) hydrolysates were obtained by alcalase and flavourzyme hydrolysis of the bean seed protein. Peptide in the bean hydrolysates, with hydrophobic amino acids had been studied for their inhibitory ACE-1 activity preventing transformation into ACE-2 that prevention hypertension. This study aimed to measure inhibitory ACE-1 activity of protein hydrolyzates from the bean Phaseolus genus spp. grown in Jember, and its solubility. The bean protein (19.8-20,2%) was extracted using isoelectric precipitation at pH 4-4,6. The extract were hydrolyzed at pH 8-9 for alcalase and pH 7 for flavourzyme, followed with inactivation at 80-85 o C. ACE-1 inhibitory activity was measured based on the amount of hippuric acid (HA) formed by the hydrolysis of Hippuryl-His-Leu (HHL) in spectrophotometry detection method (228 nm). The ultra chromatography evaluation showed that the protein hydrolysates of kidney bean contained higher hydrophobic amino acids (455.5 mg/g protein) compare to those of lima bean (350 mg/g protein). Protein hydrolysates of both beans from alcalase hydrolysis have higher ACE-1 inhibitory rather than those from flavourzyme. Protein hydrolysate from Phaseolus spp bean protein hydrolysis by alcalase, contain small molecular weight peptides (3.9-22.6 kDa) high ACE-1 inhibition ability (83 -88%), and therefore suggested as antihypertensive nutraceuticals. Highest solubility of protein hydrolysate resulted from alcalase hydrolysis of both beans were observed at pH 8-9, while those resulted from flavorzyme hydrolysis were at pH 7.

Biochemical and Functional Characterization of Kidney Bean Protein Alcalase-Hydrolysates and Their Preservative Action on Stored Chicken Meat

A new preservation approach is presented in this article to prolong the lifetime of raw chicken meat and enhance its quality at 4 °C via coating with highly soluble kidney bean protein hydrolysate. The hydrolysates of the black, red, and white kidney protein (BKH, RKH, and WKH) were obtained after 30 min enzymatic hydrolysis with Alcalase (E/S ratio of 1:100, hydrolysis degree 25–29%). The different phaseolin subunits (8S) appeared in SDS-PAGE in 35–45 kD molecular weight range while vicilin appeared in the molecular weight range of 55–75 kD. The kidney bean protein hydrolysates have considerable antioxidant activity as evidenced by the DPPH-scavenging activity and β-carotine-linolenic assay, as well as antimicrobial activity evaluated by disc diffusion assay. BKH followed by RKH (800 µg/mL) significantly (p ≤ 0.05) scavenged 95, 91% of DPPH and inhibited 82–88% of linoleic oxidation. The three studied hydrolysates significantly inhibited the growth of bacteria, yeast, and fungi, where BKH was the most performing. Kidney bean protein hydrolysates could shield the chicken meat because of their amphoteric nature and many functional properties (water and oil-absorbing capacity and foaming stability). The quality of chicken meat was assessed by tracing the fluctuations in the chemical parameters (pH, met-myoglobin, lipid oxidation, and TVBN), bacterial load (total bacterial count, and psychrophilic count), color parameters and sensorial traits during cold preservation (4 °C). The hydrolysates (800 µg/g) significantly p ≤ 0.05 reduced the increment in meat pH and TVBN values, inhibited 59–70% of lipid oxidation as compared to control during 30 days of cold storage via eliminating 50% of bacterial load and maintained secured storage for 30 days. RKH and WKH significantly (p ≤ 0.05) enhanced L*, a* values, thus augmented the meat whiteness and redness, while, BKH increased b* values, declining all color parameters during meat storage. RKH and WKH (800 µg/g) (p ≤ 0.05) maintained 50–71% and 69–75% of meat color and odor, respectively, increased the meat juiciness after 30 days of cold storage. BKH, RKH and WKH can be safely incorporated into novel foods.