Metabolite Variation between Nematode and Bacterial Seed Galls in Comparison to Healthy Seeds of Ryegrass Using Direct Immersion Solid-Phase Microextraction (DI-SPME) Coupled with GC-MS.

Annual ryegrass toxicity (ARGT) is an often-fatal poisoning of livestock that consume annual ryegrass infected by the bacterium Rathayibacter toxicus. This bacterium is carried into the ryegrass by a nematode, Anguina funesta, and produces toxins within seed galls that develop during the flowering to seed maturity stages of the plant. The actual mechanism of biochemical transformation of healthy seeds to nematode and bacterial gall-infected seeds remains unclear and no clear-cut information is available on what type of volatile organic compounds accumulate in the respective galls. Therefore, to fill this research gap, the present study was designed to analyze the chemical differences among nematode galls (A. funesta), bacterial galls (R. toxicus) and healthy seeds of annual ryegrass (Lolium rigidum) by using direct immersion solid-phase microextraction (DI-SPME) coupled with gas chromatography−mass spectrometry (GC-MS). The method was optimized and validated by testing its linearity, sensitivity, and reproducibility. Fifty-seven compounds were identified from all three sources (nematode galls, bacterial galls and healthy seed), and 48 compounds were found to be present at significantly different (p < 0.05) levels in the three groups. Five volatile organic compounds (hexanedioic acid, bis(2-ethylhexyl) ester), (carbonic acid, but-2-yn-1-yl eicosyl ester), (fumaric acid, 2-ethylhexyl tridec-2-yn-1-yl ester), (oct-3-enoylamide, N-methyl-N-undecyl) and hexacosanoic acid are the most frequent indicators of R. toxicus bacterial infection in ryegrass, whereas the presence of 15-methylnonacosane, 13-methylheptacosane, ethyl hexacosyl ether, heptacosyl acetate and heptacosyl trifluoroacetate indicates A. funesta nematode infestation. Metabolites occurring in both bacterial and nematode galls included batilol (stearyl monoglyceride) and 9-octadecenoic acid (Z)-, tetradecyl ester. Among the chemical functional group, esters, fatty acids, and alcohols together contributed more than 70% in healthy seed, whereas this contribution was 61% and 58% in nematode and bacterial galls, respectively. This study demonstrated that DI-SPME is a valid technique to study differentially expressed metabolites in infected and healthy ryegrass seed and may help provide better understanding of the biochemical interactions between plant and pathogen to aid in management of ARGT.

Predicting potential global distribution and risk regions for potato cyst nematodes (Globodera rostochiensis and Globodera pallida).

Potato cyst nematodes (PCNs), golden (yellow) cyst nematode (Globodera rostochiensis, gPCN) and pale (white) cyst nematode (G. pallida, pPCN), are important invasive pests in many countries and regions where they can cause significant yield and economic loss for agriculture. Prediction and identification of habitats suitable for PCNs are critical for developing biosecurity strategies, both pre and post border, to maximise the potential for early elimination should an incursion occur. To date, the potential global distribution of PCNs has not been thoroughly studied. Therefore, this study conducted a species distribution model to illustrate the potential global distribution of PCNs and risk regions. In this study, the Maximum Entropy Model (Maxent) associated with the Geographic Information System (GIS) was employed to reveal the potential distribution of the gPCN and pPCN. In addition to bioclimate, soil quality was also included in the model. The global cultivated lands, whether the susceptible hosts were present or not, were used to assess the maximum potential risk regions. The limitation factors for PCNs distribution were also assessed. Results showed that 66% of the global land surface was suitable for gPCN or pPCN or both, and both species can colonise more than 75% of the global cultivated lands. The coldest quarter's mean temperature and precipitation were critical limitations in unsuitable regions. In summary, the global risk maps of PCNs contribute valuable additional information that complements previous national/regional distribution predictions. The results of this distribution research will contribute practical support for decision-makers and practitioners to implement biosecurity strategies from a global perspective, that incorporate prevention or promptly enforce control practices to limit the damage caused by future incursions.

Tea-Soybean Intercropping Improves Tea Quality and Nutrition Uptake by Inducing Changes of Rhizosphere Bacterial Communities.

The positive aspects of the tea plant/legume intercropping system draw attention to the Chinese tea industry for its benefit for soil fertility improvement with low fertilizer input. However, limited information exists as to the roles of intercropped legumes in the rhizosphere microbiome and tea quality. Hereby, soybean was selected as the intercropped plant to investigate its effect on bacterial communities, nutrient competition, tea plant development, and tea quality. Our data showed that intercropped soybean boosted the uptake of nitrogen in tea plants and enhanced the growth of young tea shoots. Nutrient competition for phosphorus and potassium in soil existed between soybeans and tea plants. Moreover, tea/soybean intercropping improved tea quality, manifested by a significantly increased content of non-ester type catechins (C, EGC, EC), total catechins and theanine, and decreased content of ester type catechins (EGCG). Significant differences in rhizobacterial composition were also observed under different systems. At the genus level, the relative abundance of beneficial bacteria, such as Bradyrhizobium, Saccharimonadales and Mycobacterium, was significantly increased with the intercropping system, while the relative abundance of denitrifying bacteria, Pseudogulbenkiania, was markedly decreased. Correlation analysis showed that Pseudogulbenkiania, SBR1031, and Burkholderiaceae clustered together showing a similar correlation with soil physicochemical and tea quality characteristics; however, other differential bacteria showed the opposite pattern. In conclusion, tea/soybean intercropping improves tea quality and nutrition uptake by increasing the relative abundance of beneficial rhizosphere bacteria and decreasing denitrifying bacteria. This study strengthens our understanding of how intercropping system regulate the soil bacterial community to maintain the health of soils in tea plantations and provides the basis for replacing chemical fertilizers and improving the ecosystem in tea plantations.

Gut-Testis Axis: Microbiota Prime Metabolome To Increase Sperm Quality in Young Type 2 Diabetes.

Young type 2 diabetes (T2D) affects 15% of the population, with a noted increase in cases, and T2D-related male infertility has become a serious issue in recent years. The current study aimed to explore the improvements of alginate oligosaccharide (AOS)-modified gut microbiota on semen quality in T2D. The T2D was established in young mice of 5 weeks of age with a blood glucose level of 21.2 ± 2.2 mmol/L, while blood glucose was 8.7 ± 1.1 mM in control animals. We discovered that fecal microbiota transplantation (FMT) of AOS-improved microbiota (A10-FMT) significantly decreased blood glucose, while FMT of gut microbiota from control animals (Con-FMT) did not. Sperm concentration and motility were decreased in T2D to 10% to 20% of those in the control group, while A10-FMT brought about a recovery of around 5- to 10-fold. A10-FMT significantly increased small intestinal Allobaculum, while it elevated small intestinal and cecal Lactobacillus in some extent, blood butyric acid and derivatives and eicosapentaenoic acid (EPA), and testicular docosahexaenoic acid (DHA), EPA, and testosterone and its derivatives. Furthermore, A10-FMT improved liver functions and systemic antioxidant environments. Most importantly, A10-FMT promoted spermatogenesis through the improvement in the expression of proteins important for spermatogenesis to increase sperm concentration and motility. The underlying mechanisms may be that A10-FMT increased gut-beneficial microbes Lactobacillus and Allobaculum to elevate blood and/or testicular butyric acid, DHA, EPA, and testosterone to promote spermatogenesis and thus to ameliorate sperm concentration and motility. AOS-improved gut microbes could emerge as attractive candidates to treat T2D-diminished semen quality. IMPORTANCE A10-FMT benefits gut microbiota, liver function, and systemic environment via improvement in blood metabolome, consequently to favor the testicular microenvironment to improve spermatogenesis process and to boost T2D-diminished semen quality. We established that AOS-improved gut microbiota may be used to boost T2D-decreased semen quality and metabolic disease-related male subfertility.

Role of Lipids in Phosphine Resistant Stored-Grain Insect Pests Tribolium castaneum and Rhyzopertha dominica.

Insects rely on lipids as an energy source to perform various activities, such as growth, flight, diapause, and metamorphosis. This study evaluated the role of lipids in phosphine resistance by stored-grain insects. Phosphine resistant and susceptible strains of the two main stored-grain insects, Tribolium castaneum and Rhyzopertha dominica, were analyzed using liquid chromatography-mass spectroscopy (LC-MS) to determine their lipid contents. Phosphine resistant strains of both species had a higher amount of lipids than susceptible stains. Significant variance ratios between the resistant and susceptible strains of T. castaneum were observed for glycerolipids (1.13- to 53.10-fold) and phospholipids (1.05- to 20.00-fold). Significant variance ratios between the resistant and susceptible strains of R. dominica for glycerolipids were 1.04- to 31.50-fold and for phospholipids were 1.04- to 10.10-fold. Glycerolipids are reservoirs to face the long-term energy shortage. Phospholipids act as a barrier to isolate the cells from the surrounding environment and allow each cell to perform its specific function. Thus, lipids offer a consistent energy source for the resistant insect to survive under the stress of phosphine fumigation and provide a suitable environment to protect the mitochondria from phosphine. Hence, it was proposed through this study that the lipid content of phosphine-resistant and phosphine-susceptible strains of T. castaneum and R. dominica could play an important role in the resistance of phosphine.

Proanthocyanidins Modulate Rumen Enzyme Activities and Protein Utilization In Vitro.

This study investigated the principal leaf protein (rubisco) solubilization and in vitro ruminal enzyme activity in relation to the molecular structure of proanthocyanidins extracted from leaves of Anogeissus pendula and Eugenia jambolana. Six proanthocyanidin fractions were extracted by 50% (v/v) methanol−water followed by 70% (v/v) acetone−water and then distilled water from leaves of A. pendula (AP) and E. jambolana (EJ) to yield EJ−70, EJ−50, EJ−DW, AP−70, AP−50 and AP−DW. Fractions were examined for their molecular structure and their effects on sheep ruminal enzymes and solubilization of rubisco in vitro. All fractions significantly (p < 0.05) inhibited the activity of ruminal glutamic oxaloacetic transaminase and glutamic pyruvic transaminase. The fractions AP−50 and EJ−50 significantly inhibited the activity of the R-cellulase enzyme. Most of the fractions inhibited R-glutamate dehydrogenase activity (p < 0.05) by increasing its concentration, while protease activity decreased by up to 58% with increasing incubation time and concentration. The solubilization of rubisco was observed to be comparatively higher in A. pendula (16.60 ± 1.97%) and E. jambolana (15.03 ± 1.06%) than that of wheat straw (8.95 ± 0.95%) and berseem hay (3.04 ± 0.08%). A significant (p < 0.05) increase in protein solubilization was observed when wheat straw and berseem hay were supplemented with A. pendula and E. jambolana leaves at different proportions. The efficiency of microbial protein was significantly (p < 0.05) greater with the supplementation of leaves of A. pendula in comparison to E. jambolana. The overall conclusion is that the proanthocyanidins obtained from E. jambolana exhibited greater inhibitory activities on rumen enzymes, whereas A. pendula recorded higher protein solubilization. Thus, PAs from A. pendula and E. jambolana appear to have the potential to manipulate rumen enzyme activities for efficient utilization of protein and fiber in ruminants.

Sequential Extraction of Proanthocyanidin Fractions from Ficus Species and Their Effects on Rumen Enzyme Activities In Vitro.

Three proanthocyanidin fractions per species were sequentially extracted by 50% (v/v) methanol−water, 70% (v/v) acetone−water, and distilled water from leaves of Ficus racemosa (fractions FR) and F. religiosa (fractions FRL) to yield fractions FR-50, FR-70, FR-DW, FRL-50, FRL-70, and FRL-DW. Fractions were examined for their molecular structure, effect on ruminal enzyme activities, and principal leaf protein (Rubisco) solubilization in vitro. All fractions except FRL-70 contained flavonoids including (+) catechin, (−) epicatechin, (+) gallocatechin, (−) epigallocatechin, and their -4-phloroglucinol adducts. The fractions FRL-50 and FRL-DW significantly (p < 0.05) inhibited the activity of ruminal glutamic oxaloacetic transaminase and glutamic pyruvic transaminase. All fractions inhibited glutamate dehydrogenase activity (p < 0.05) with increasing concentration, while protease activity decreased 15−18% with increasing concentrations. Fractions FRL-50 and FRL-DW completely inhibited the activity of cellulase enzymes. Solubilization of Rubisco was higher in F. religiosa (22.36 ± 1.24%) and F. racemosa (17.26 ± 0.61%) than that of wheat straw (WS) (8.95 ± 0.95%) and berseem hay (BH) (3.04 ± 0.08%). A significant (p < 0.05) increase in protein solubilization was observed when WS and BH were supplemented with FR and FRL leaves at different proportions. The efficiency of microbial protein was significantly (p < 0.05) greater in diets consisting of WS and BH with supplementation of F. racemosa leaves in comparison to those supplemented with F. religiosa leaves. The overall conclusion is that the fractions extracted from F. religiosa showed greater inhibitory effects on rumen enzymes and recorded higher protein solubilization in comparison to the F. racemosa. Thus, PAs from F. religiosa are potential candidates to manipulate rumen enzymes activities for efficient utilization of protein and fiber in ruminants.

The Effect of Ozone Treatment on Metabolite Profile of Germinating Barley.

Ozone is widely used to control pests in grain and impacts seed germination, a crucial stage in crop establishment which involves metabolic alterations. In this study, dormancy was overcome through after-ripening (AR) in dry barley seed storage of more than 4 weeks; alternatively, a 15-min ozone treatment could break the dormancy of barley immediately after harvest, with accelerated germination efficiency remaining around 96% until 4 weeks. Headspace solid-phase microextraction (HS-SPME) and liquid absorption coupled with gas chromatography mass spectrometry (GC-MS) were utilized for metabolite profiling of 2-, 4- and 7-day germinating seeds. Metabolic changes during barley germination are reflected by time-dependent characteristics. Alcohols, fatty acids, and ketones were major contributors to time-driven changes during germination. In addition, greater fatty acids were released at the early germination stage when subjected to ozone treatment.

Ozone Efficiency on Two Coleopteran Insect Pests and Its Effect on Quality and Germination of Barley.

Ozone (O3) is a potential fumigant to control pests in stored grain since it can safely and rapidly auto-decompose without leaving residues. In this study, the efficacy of O3 on all life stages of Rhyzopertha dominica (Fabricius) and Tribolium castaneum (Herbst) in barley and the physiological effects on barley and its quality were investigated. Complete control of all life stages of pests was obtained at 700 ppm for 1440 min of ozone exposure without negatively impacting the contents of soluble protein, moisture content, seed colour, hardness, and the weight of thousand barley seeds. The eggs and pupae of these two insects were the more tolerant stages than their larvae and adults. Prolonged exposure times (40 to 1440 min) and mortality assessment intervals (1, 2, and 7 days) increased O3 efficacy due to the reaction characteristics and delayed toxicity. Aging barley seeds appeared to be more sensitive to prolonged ozone duration than new seeds. A total of 20 and 40 min could promote germination rate, and longer O3 exposure (1440 min) was unfavourable for germination and seedling growth. Thus, it is imperative to select an optimal O3 exposure time to transfer ozone into quality contributors of final products and achieve the desired functional outcomes.

Gut Microbiota-Testis Axis: FMT Mitigates High-Fat Diet-Diminished Male Fertility via Improving Systemic and Testicular Metabolome.

High-fat diet (HFD)-induced obesity is known to be associated with reduced male fertility and decreased semen quality in humans. HFD-related male infertility is a growing issue worldwide, and it is crucial to overcome this problem to ameliorate the distress of infertile couples. For the first time, we discovered that fecal microbiota transplantation (FMT) of alginate oligosaccharide (AOS)-improved gut microbiota (A10-FMT) ameliorated HFD-decreased semen quality (sperm concentration: 286.1 ± 14.1 versus 217.9 ± 17.4 million/mL; sperm motility: 40.1 ± 0.7% versus 29.0 ± 0.9%), and male fertility (pregnancy rate: 87.4 ± 1.1% versus 70.2 ± 6.1%) by benefiting blood and testicular metabolome. A10-FMT improved HFD-disturbed gut microbiota by increasing gut Bacteroides (colon: 24.9 ± 1.1% versus 8.3 ± 0.6%; cecum: 10.2 ± 0.7% versus 3.6 ± 0.7%) and decreasing Mucispirillum (colon: 0.3 ± 0.1% versus 2.8 ± 0.4%; cecum: 2.3 ± 0.5% versus 6.6 ± 0.7%). A10-FMT benefited gut microbiota to improve liver function by adjusting lipid metabolism to produce n-3 polyunsaturated fatty acids, such as eicosapentaenoic acid (blood: 55.5 ± 18.7 versus 20.3 ± 2.4) and docosahexaenoic acid (testis: 121.2 ± 6.2 versus 89.4 ± 6.7), thus ameliorating HFD-impaired testicular microenvironment to rescue spermatogenesis and increase semen quality and fertility. The findings indicated that AOS-improved gut microbiota may be a promising strategy to treat obesity or metabolic issues-related male infertility in the future. IMPORTANCE HFD decreases male fertility via upsetting gut microbiota and transplantation of AOS-benefited gut microbiota (A10-FMT) improves gut microbiota to ameliorate HFD-reduced male fertility. Moreover, A10-FMT improved liver function to benefit the blood metabolome and simultaneously ameliorated the testicular microenvironment to turn the spermatogenesis process on. We demonstrated that AOS-benefited gut microbiota could be applied to treat infertile males with obesity and metabolic issues induced by HFD.

Response of Aphid Parasitoids to Volatile Organic Compounds from Undamaged and Infested Brassica oleracea with Myzus persicae.

Headspace solid microextraction (HS-SPME) and GC-MS were used to investigate volatile organic compounds (VOCs) from cabbage plants infested and uninfested with green peach aphid Myzus persicae. The HS-SPME combined with GC-MS analysis of the volatiles described the differences between the infested and uninfested cabbage. Overall, 28 compounds were detected in infested and uninfested cabbage. Some VOCs released from infested cabbage were greater than uninfested plants and increased the quantity of the composition from infested plants. According to the peak area from the GC-MS analysis, the VOCs from infested cabbage consisted of propane, 2-methoxy, alpha- and beta pinene, myrcene, 1-hexanone, 5-methyl-1-phenyl-, limonene, decane, gamma-terpinen and heptane, 2,4,4-trimethyl. All these volatiles were higher in the infested cabbage compared with their peak area in the uninfested cabbage. The results of the study using a Y-shape olfactometer revealed that the VOCs produced by infested cabbage attracted Myzus persicae substantially more than uninfested plants or clean air. The percentage of aphid choice was 80% in favor of infested cabbage; 7% were attracted to the clean air choice and uninfested plants. A total of aphids 7% were attracted to clean air. Comparing between infested and uninfested cabbage plants, the aphid was attracted to 63% of the infested cabbage, versus 57% of the uninfested cabbage. The preferences of Aphidus colemani and Aphelinus abdominalis to the infested or uninfested plants with M. persicae and compared with clean air indicated that parasitoids could discriminate the infested cabbage. Both parasitoids significantly responded to the plant odor and were attracted to 86.6% of the infested cabbage plants.

Profiling Real-Time Aroma from Green Tea Infusion during Brewing.

Aroma substances are the most crucial criteria for the sensory evaluation of tea quality, and also key attractors influencing consumers to make the decision for purchasing tea. Understanding the aromatic properties of tea infusion during different brewing time is crucial to control the tea aromatic quality. Here, headspace and direct immersion solid-phase microextraction (HS-SPME and DI-SPME), coupled with GC-MS, were employed to investigate the impact of brewing time on the changes of the volatile features of green tea infusion. Esters, aldehydes, alcohols, fatty acids, and alkaloids were the predominant volatile groups from tea infusions. Two to three minutes was identified as the best duration for the tea brewing that can maximize the abundance of aromatic chemicals in the headspace emitted from the tea infusions. The variation of the key aromatic contributors between the tea infusion and the headspace over the infusion tended to equilibrate during the tea brewing process. This study provides a theory-based reference method by analyzing the real-time aromatic characteristics in green tea. The optimal time was determined for aromatic quality control, and the complementary relationship between the volatiles in the headspace and its counterpart, tea infusion, was primarily elucidated.

Evaluation of Aphicidal Effect of Essential Oils and Their Synergistic Effect against Myzus persicae (Sulzer) (Hemiptera: Aphididae).

The insecticidal activities of essential oils obtained from black pepper, eucalyptus, rosemary, and tea tree and their binary combinations were investigated against the green peach aphid, Myzus persicae (Aphididae: Hemiptera), under laboratory and glasshouse conditions. All the tested essential oils significantly reduced and controlled the green peach aphid population and caused higher mortality. In this study, black pepper and tea tree pure essential oils were found to be an effective insecticide, with 80% mortality when used through contact application. However, for combinations of essential oils from black pepper + tea tree (BT) and rosemary + tea tree (RT) tested as contact treatment, the mortality was 98.33%. The essential oil combinations exhibited synergistic and additive interactions for insecticidal activities. The combination of black pepper + tea tree, eucalyptus + tea tree (ET), and tea tree + rosemary showed enhanced activity, with synergy rates of 3.24, 2.65, and 2.74, respectively. Essential oils formulation was effective on the mortality of aphids. Fourier Transform Infrared Spectroscopy (FTIR) analysis showed that stability of a mixture of essential oils was not affected by store temperature (15, 25, and 35 °C) and the functional groups were not changed during storage. Based on our results, the essential oils can be used as a commercial insecticide against M. persicae.

Study on the Correlation between the Protein Profile of Lupin Milk and Its Cheese Production Compared with Cow's Milk.

Australian sweet lupin, the largest legume crop grown in Western Australia, is receiving global attention from the producers of new foods. To understand the effect of protein on cheese yield, lupin milk proteins were separated from the first, second, and third filtrations by cheesecloths. However, proteins from the first and second were analyzed using two-dimensional polyacrylamide gel electrophoresis; then, the isolated proteins associated with cheese production were identified. The research also focused on identifying the optimal method of cheese production based on the coagulation process, temperature, yield, and sensory evaluation. Lupin curds from the two cultivars, Mandelup and PBA Jurien, were produced using vinegar, lemon juice, starter culture, vegetable rennet enzyme as coagulant, as well as curd generated using starter culture and vegetable rennet enzyme. Cow's milk was used as a control. The results indicated that first-time filtration produced better extraction and higher yield of lupin proteins and cheese than the second filtration. A sensory analysis indicated that lupin cheese produced from PBA Jurien lupin milk using vinegar, 7.80% expressed as acetic acid, and ground in 45 °C water, was the most acceptable. The cheeses were examined for their protein, carbohydrates, fat, ash, and moisture contents. The concentration of protein was approximately 27.3% and 20.6%, respectively, in the cheese from PBA Jurien and Mandelup. These results suggest that lupin milk can adequately supply the proteins needed in human diets and, thus, could be used in the production of many existing products that require animal milk as an input.

Mediterranean Fruit Fly Ceratitis capitata (Diptera: Tephritidae) Eggs and Larvae Responses to a Low-Oxygen/High-Nitrogen Atmosphere.

The Mediterranean fruit fly, Ceratitis capitata (Wiedemann) (Diptera: Tephritidae), is one of the most damaging horticultural insect pests. This study used a low-oxygen/high-nitrogen bioassay to control C. capitata. Two low-oxygen treatments were applied (0.5% O2 + 99.5 N2 and 5% O2 + 95% N2) to C. capitata eggs and 1st, 2nd and 3rd instar larvae from 0 to nine days on a carrot diet at 25 °C; 70-75% RH. The pupariation, adult emergence, and sex ratios of survived flies were examined. The results demonstrate that increased mortality of all tested life stages correlated with increased exposure times at both levels of low-oxygen treatments. Complete control of eggs was achieved after eight days and nine days for larvae using 0.5% O2 at 25 °C; 70-75% RH. The 3rd instar was the most tolerant stage, while the egg was the most susceptible stage to the low-oxygen environment. There were no significant differences in sex ratios between emerged adults after low-oxygen and control treatments. The present work demonstrates and confirms the mortalities of C. capitata caused by low-oxygen treatment, which may help develop new postharvest strategies to control this destructive fruit fly pest.

Study on Effect of Extraction Techniques and Seed Coat on Proteomic Distribution and Cheese Production from Soybean Milk.

Soybean-based food products are a major source of protein. In the present study, proteins in soybean milk from seeds of the cultivar Bunya (Glycine max) were extracted using the cheesecloth and the centrifuge methods. The milk was produced through mechanical crushing of both whole and split seeds in water. Following separation by either the cheesecloth or centrifuge, proteins were isolated from the soybean milk by using thiourea/urea solubilisation and then separated them using two-dimensional polyacrylamide gel electrophoresis. The isolated proteins were identified by mass spectrometry. A total of 97 spots were identified including 49 that displayed different abundances. Of the two separation techniques, centrifuge separation gave higher protein extraction and more intense protein spots than cheesecloth separation. Eleven of the ß-subunits of ß-conglycinin, three of the α-subunits of ß-conglycinin, and four of the mutant glycinin showed different levels of abundances between separation techniques, which might be related to subsequent cheese quality. Notably, split-seed soybean milk has less allergenic proteins with four α-subunits of ß-conglycinin compared to whole-seed milk with eight of those proteins. The sensory evaluation showed that the cheese produced from split-soybean milk received higher consumer preferences compared to that of whole seed, which could be explained by their proteomic differences. The demonstrated reference map for whole and split-seed soybean milk could be further utilized in the research related to soybean cheesemaking.

Retraction: Al-Behadili, F.J.M., et al. Cold Responses of the Mediterranean Fruit Fly Ceratitis capitata Wiedemann (Diptera: Tephritidae) in Blueberry. Insects 2020, 11, 276.

The published article [...].

Cold Responses of the Mediterranean Fruit Fly Ceratitis capitata Wiedemann (Diptera: Tephritidae) in Blueberry.

In this study, the effects of cold treatment at 1.0 ± 0.2 °C were investigated on the eggs of the 1st, 2nd, and 3rd instar larvae of Ceratitis capitata in two Australian blueberry cultivars C99-42 and C00-09. Pupariation, emerged adults, and sex ratios were examined on C. capitata after the treatment. The results showed that exposure time at low temperatures was a key factor to affect pupariation and adult emergence. Eleven days of exposure to cold treatment at 1.0 ± 0.2 °C were enough to eradicate all four immature stages in both cultivars. Cold tolerance of the four C. capitata stages was not affected when reared on two different blueberry cultivars. The third instar larva is the most tolerant stage for cold treatment in two blueberry cultivars. There were no significant differences in sex ratios from surviving C. capitata among different stages after treatment. This study improves understanding of C. capitata responses to cold treatment in blueberries, which may contribute to phytosanitary required quarantine treatment of this destructive horticulture pest species.

Proteomic Characterisation of Lupin (Lupinus angustifolius) Milk as Influenced by Extraction Techniques, Seed Coat and Cultivars.

Lupin seeds are rich in proteins and other essential ingredients that can help to improve human health. The protein contents in both whole and split seeds of two lupin cultivars (Mandleup and PBA Jurien) were used to produce the lupin milk using the cheesecloth and centrifuge method. Proteins were extracted from the lupin milk using thiourea/urea solubilization. The proteins were separated by a two-dimensional polyacrylamide gel electrophoresis and then identified with mass spectrometry. A total of 230 protein spots were identified, 60 of which showed differential abundances. The cheesecloth separation showed protein extractability much better than that of the centrifuge method for both the cultivars. The results from this study could offer guidance for future comparative analysis and identification of lupin milk protein and provide effective separation technique to determine specific proteins in the cheese-making process.

Preliminary Study on the Differences in Hydrocarbons Between Phosphine-Susceptible and -Resistant Strains of Rhyzopertha dominica (Fabricius) and Tribolium castaneum (Herbst) Using Direct Immersion Solid-Phase Microextraction Coupled with GC-MS.

Phosphine resistance is a worldwide issue threatening the grain industry. The cuticles of insects are covered with a layer of lipids, which protect insect bodies from the harmful effects of pesticides. The main components of the cuticular lipids are hydrocarbon compounds. In this research, phosphine-resistant and -susceptible strains of two main stored-grain insects, T. castaneum and R. dominica, were tested to determine the possible role of their cuticular hydrocarbons in phosphine resistance. Direct immersion solid-phase microextraction followed by gas chromatography-mass spectrometry (GC-MS) was applied to extract and analyze the cuticular hydrocarbons. The results showed significant differences between the resistant and susceptible strains regarding the cuticular hydrocarbons that were investigated. The resistant insects of both species contained higher amounts than the susceptible insects for the majority of the hydrocarbons, sixteen from cuticular extraction and nineteen from the homogenized body extraction for T. castaneum and eighteen from cuticular extraction and twenty-one from the homogenized body extraction for R. dominica. 3-methylnonacosane and 2-methylheptacosane had the highest significant difference between the susceptible and resistant strains of T. castaneum from the cuticle and the homogenized body, respectively. Unknown5 from the cuticle and 3-methylhentriacontane from the homogenized body recorded the highest significant differences in R. dominica. The higher hydrocarbon content is a key factor in eliminating phosphine from entering resistant insect bodies, acting as a barrier between insects and the surrounding phosphine environment.