Larval Excretion Research Articles

Obstrucción Intestinal por infección por Strongyloides Stercolaris, a propósito de un caso

Strongyloidiasis is a parasitosis caused by the nematode Strongyloides stercoralis, which commonly causes gastrointestinal problems. We present the case of a 30-year-old man who developed Strongyloides hyperinfection, with a history of a presumptive diagnosis of polymyositis for which he received a short course of corticosteroids. Upon admission, he presented with hyporexia, generalized weakness, cachectic syndrome, self-limited intermittent diarrhea, oral intolerance, and mild abdominal distension. Our patient reached the dissemination stage, which resulted in severe damage at the intestinal level. The diagnosis was difficult due to the low excretion of larvae in the feces. Treatment was started with parenteral ivermectin at a dose of 1.2ml via sc every 48 hours for 03 doses, with good response to treatment and good tolerance of the oral route. The importance of presenting the case is to present the diagnostic and therapeutic approach to this disease that has been little studied in Peru.

Upper Gastrointestinal Bleeding Induced by Gastric Ulcer Secondary to Strongyloidiasis: A Case Report.

gastric ulcer; gastrointestinal haemorrhage; Strongyloides stercoralis; strongyloidiasis.

Extraction and quantification of 1-Deoxynojirimycin in Silkworm during metamorphosis and moulting stages

The contents of 1-deoxynojirimycin (1-DNJ) in the silkworm, Bombyx mori, were examined using reverse-phase high-performance liquid chromatography at various developmental stages. Mulberry leaves and silkworms naturally contain 1-deoxynojirimycin (DNJ). Although the contents of DNJ and its actions in mulberry leaves have been studied for decades, the content of DNJ in silkworms and its use have received less attention. DNJ was extracted and quantified in a variety of silkworms including Tasar silkworms, Muga silkworms, Eri silkworms and Mulberry silkworms. Group VI silkworms are more active than the others. Then, for male and female larvae of silkworm species, the variation in DNJ contents was examined. Silkworm larvae's 1-DNJ concentration varies greatly depending on their developmental stage. Male larvae (133.25 ± 11.45 mg/100g) and female larvae (129.65 ± 10.25 mg/100g) are substantially higher in both male and female mulberry silkworms. The larvae of the 3rd and the 5th day were used to calculate the DNJ. Male larvae had a higher 1-DNJ accumulation efficiency than females and there was also a significant difference between silkworm strains. The current findings demonstrate that 1-DNJ tissue distribution is substantially increased in the blood, intestinal juice and the alimentary canal while none was found in the silk gland. Furthermore, 1-DNJ was not detected in silkworms fed with an artificial diet devoid of mulberry leaf powder. This shows that silkworms get 1-DNJ from mulberry leaves; they could not make it on their own. During the larval stage, the storage and excretion of 1-DNJ fluctuate. 1-DNJ was not found in newly hatched larvae and it was primarily accumulated during the early and middle stages of each instar, before being expelled at later stages. Furthermore, 1-DNJ can be extracted from larval excrement, making it ideal for the development of 1-DNJ-related goods.

Adult parasite burden and excretion of first-stage larvae of Angiostrongylus vasorum in dogs: Methodologically relevant diagnostic aspects and associations with serological detection of parasite antigen and specific antibodies

Angiostrongylus vasorum is a widely distributed cardiopulmonary parasite of canids in Europe. Clinical signs in dogs can be highly variable and diagnostically challenging. A correct and early diagnosis is hence indispensable to adequately manage affected patients. First-stage larvae (L1) are excreted in the faeces of definitive hosts and conventionally identified using the Baermann technique. Moreover, ELISAs for the detection of circulating antigen and specific antibodies have been presented. The current study aimed at i) quantitatively assessing larval migration in the Baermann funnel after 12 h and 24 h; ii) investigating the influence of sample storage at 4 °C over the course of three days on the number of detected L1; iii) evaluating potential associations of adult worm burdens with larval shedding in dogs and ELISA optical density (OD) values for circulating parasite antigen and specific antibodies. Faecal samples were obtained from naturally infected dogs (n = 21) and Baermann funnels were set up in duplicate over the course of four consecutive days (days 0–3) starting with the day of sample collection. Funnels were harvested on days 1–4 after 12 and 24 h, respectively, and the number of L1 per gram faeces (LPG) was determined. The LPG did not differ between larval harvest after 12 h from harvest after 24 h. Storage of faecal samples at 4 °C for two and three days entailed a considerable decrease in LPG. Adult worm burdens and larval excretion data from previous experiments demonstrated a correlation between worm burden and LPG. In contrast, no correlations between worm burden and the level of parasite antigen and specific antibody OD values, respectively, were identified. Thus, OD values of both antigen and antibody ELISA did not allow for conclusions on infection intensity reflected by the number of adult parasites. For the detection of L1 in faeces, 12 or 24 h of larval migration time was not discriminating for A. vasorum positivity. Thus, early processing of faecal samples is essential, since larval detection and hence sensitivity of the approach considerably decreased over the course of three days of storage. Therefore, the common recommendation to collect faecal samples for three consecutive days and to subsequently analyse them needs to be reconsidered. The results of this study can be readily translated into precise recommendations for daily practice to adequately assess A. vasorum infected dogs.

A note on the relationship between fecal larval excretion and Protostrongylus rufescens lungworm burden in sheep.

The examination of feces for stages of parasitic helminths is the most widely used methodology for the intravital diagnosis in domestic animals of patent endoparasitism including pulmonary nematode infections. Although there is only little information on the relationship of lungworm larval excretion and corresponding parasite burdens, fecal larval counts are used as indirect measure ("biomarker") for the intensity of infection, for instance in anthelmintic efficacy studies. To assess the relationship between fecal larval and Protostrongylus rufescens parasite counts in sheep, log-transformed data of 14 naturally infected animals were analyzed. The larval excretion of the sheep was monitored in approximately weekly intervals over 6weeks before lungworm recovery. Analyses were performed on the larval counts (at a single time point or counts averaged over several consecutive time points) relating to parasite counts. Fecal larval counts and the P. rufescens nematode burden (range, 17 to 406) were significantly and strongly correlated (p < 0.05 for all analyses; Spearman's r > 0.6) with the number of larvae excreted increasing with increasing lungworm burden. Subsequently performed regression analysis demonstrated a statistically significant strong linear relationship between P. rufescens worm and fecal larval counts (p < 0.01 for all analyses; R2, range 0.5094 to 0.8150). Analyses based on larval counts averaged over two or more consecutive time points resulted in higher Spearman's r and R2 compared with analyses based on single time point larval counts. Despite of some variability, the analyses indicate that fecal larval counts can be regarded as a useful measure of the P. rufescens burden in sheep.

Treatment of natural Protostrongylus rufescens lungworm infection in sheep with eprinomectin 5 mg/mL topical solution

Parasitic respiratory infections in domestic sheep and goats are caused by Dictyocaulus filaria and various species belonging to the Protostrongylidae family of nematodes which frequently occur in mixed infections. Although the parasitism with protostrongylid lungworms is generally considered to be of low pathogenicity, there are reports of clinical disease including cases associated with Protostrongylus rufescens infection.The efficacy against P. rufescens of eprinomectin 5 mg/mL topical solution (EPRINEX® Multi, Boehringer Ingelheim) was thus evaluated in a clinical study compliant with GCP and VICH anthelmintic efficacy testing guidelines in adult sheep with naturally acquired infection. Following ranking on pre-treatment Protostrongylus fecal larval counts and forming into blocks of two animals, the sheep were randomly allocated to either remain untreated (control) or to be administered eprinomectin 5 mg/mL topical solution at 1 mL/5 kg body weight (equivalent to 1 mg eprinomectin per kg body weight) once as a pour-on. Fecal samples of the sheep were examined to monitor the larval excretion weekly for five weeks after treatment; then the animals were necropsied for lungworm recovery and count to determine the efficacy of the treatment.After treatment, Protostrongylus larval excretion decreased to zero within three weeks. Nematode counts demonstrated that the efficacy of the treatment with eprinomectin 5 mg/mL topical solution was 100 % against P. rufescens: no lungworms were recovered from any treated sheep while all controls harbored P. rufescens (range, 17–406) (p < 0.001).The treatment was well accepted; no treatment-related health problems were observed.

Genome and transcriptome sequencing of the green bottle fly, Lucilia sericata, reveals underlying factors of sheep flystrike and maggot debridement therapy

The common green bottle blow fly Lucilia sericata (family, Calliphoridae) is widely used for maggot debridement therapy, which involves the application of sterile maggots to wounds. The larval excretions and secretions are important for consuming necrotic tissue and inhibiting bacterial growth in wounds of patients. Lucilia sericata is also of importance as a pest of sheep and in forensic studies to estimate a postmortem interval. Here we report the assembly of a 565.3 Mb genome from long read PacBio DNA sequencing of genomic DNA. The genome contains 14,704 predicted protein coding genes and 1709 non-coding genes. Targeted annotation and transcriptional analyses identified genes that are highly expressed in the larval salivary glands (secretions) and Malpighian tubules (excretions) under normal growth conditions and following heat stress. The genomic resources will underpin future genetic studies and in development of engineered strains for genetic control of L. sericata and for biotechnology-enhanced maggot therapy.

Do All Roads Lead to Rome? The Potential of Different Approaches to Diagnose Aelurostrongylus abstrusus Infection in Cats

An infection with the cat lungworm, Aelurostrongylus abstrusus, can be subclinical, but it can also cause severe respiratory clinical signs. Larvae excretion, antibody levels, clinical assessment findings of the respiratory system and diagnostic imaging findings were recorded and compared for six cats with experimental aelurostrongylosis. In five cats, patency started 33–47 days post infection (pi), but two cats excreted larvae only in long intervals and low numbers. Positive ELISA results were observed in four cats with patent aelurostrongylosis, starting between five days before and 85 days after onset of patency. One seropositive cat remained copromicroscopically negative. Mild respiratory signs were observed in all cats examined. A computed tomographic (CT) examination of the lungs displayed distinct alterations, even in absence of evident clinical signs or when larvae excretion was low or negative. The thoracic radiograph evaluation correlated with the CT results, but CT was more distinctive. After anthelmintic treatment in the 25th week post infection, pulmonary imaging findings improved back to normal within 6–24 weeks. This study shows that a multifaceted approach, including diagnostic imaging, can provide a clearer diagnosis and monitoring of disease progression. Furthermore, a CT examination provides an alternative to post mortem examination and worm counts in anthelmintic efficacy studies.

Bronchopulmonary Nematodes in Alpine Ibex: Shedding of First Stage Larvae Analyzed at the Individual Host Level.

Pneumonia is the most frequent cause of death for Alpine ibex (Capra ibex) in Gran Paradiso National Park, (Italy). The etiology of this form of pneumonia is currently unknown and the identification of the primary etiological agent remains difficult due to biological and logistic constraints. Uncovering individual differences in Protostrongylid prevalence and intensity is important to further investigate the epidemiology of respiratory diseases and their relationship to heterozygosity and inbreeding in a once almost extinct population like C. ibex. In a group of 21 individually recognizable adult male we monitored monthly prevalence and intensity of Protostrongylid first-stage larvae using Baerman's technique from June to September 2019. First-stage larvae of 5 genera were detected. Muellerius (P = 100%, CI95% = 84–100) and Protostrongylus (P = 86%, CI95%:71–100) were two dominant genera according to Bush's importance index. Neostrongylus (P = 38%,CI95%: 17–59), Cystocaulus (P = 33%,CI95% = 13–53) were classified as co-dominant genera while Dictyocaulus filaria (P = 0.05%, CI95% = 0.04–0.13) was detected, for the first time in Alpine ibex, in one subject. Protostrongylidae larval excretion varied significantly over time, with minimum L1 excretion in July. Individual median larval intensity ranged from 4.4 lpg to 82.2 lpg with Poulin's discrepancy index showing highly aggregated distribution patterns for Muellerius spp. (D = 0.283, CI95% = 0.760–0.895) and Protostrongylus spp. (D = 0.635, CI95% = 0.580–0.705). Presented data provide the necessary base point to further investigate how lungworm infection account for the different rates of progression of pneumonia in C. ibex. Individual aggregation of larval intensity must be further evaluated to determine whether these differences mirror different levels of parasitic infection related to individual differences in immune response, hormonal-states or genetic fitness.

Efficacy of Bravecto\xae Plus spot-on solution for cats (280 mg/ml fluralaner and 14 mg/ml moxidectin) for the prevention of aelurostrongylosis in experimentally infected cats

BackgroundThe feline lungworm Aelurostrongylus abstrusus affects the lower respiratory tract in cats worldwide. As infections may lead to chronic respiratory changes or even death, preventive treatment in cats with outdoor access is warranted.MethodsThe preventive efficacy of a spot-on solution (Bravecto® Plus spot-on solution for cats, MSD) against cat aelurostrongylosis was evaluated using three different preventive treatment regimes in a negative controlled, randomized and partially blinded laboratory efficacy study with 31 purposed-bred cats. The minimum recommended dose of 2.0 mg moxidectin + 40 mg fluralaner/kg bodyweight was applied once 12 (Group [G]1), 8 (G2) or 4 (G3) weeks before experimental infection with 300 third-stage larvae (L3) of A. abstrusus. Another group served as untreated control (G4). Individual faecal samples were analysed as of day 30 post infection (pi) to monitor larvae excretion. Necropsy was performed at days 47–50 pi. The lungs were examined macroscopically for pathological findings and (pre-)adult worms were counted to assess preventive efficacy.ResultsBeginning at day 32–40 pi, all cats of the control group were constantly shedding larvae of A. abstrusus, whereas only one animal of G1 excreted larvae at several consecutive days. In addition, two cats of G1 and G3 and three of G2 were positive on a single occasion. The geometric mean (GM) of the maximum number of excreted larvae was 7574.29 in the control group compared to 1.10 (G1), 1.19 (G2) and 0.53 (G3), resulting in a GM reduction of > 99.9% in all treatment groups. All lungs of the control animals showed severe or very severe alterations at necropsy, while in 94.44% of the treated cats lung pathology was rated as absent or mild. The GM number of (pre-)adult A. abstrusus retrieved from the lungs was 26.57 in the control group, 0.09 in G1 and 0.00 in G2 and G3. Thus, GM worm count reduction was 99.66% in G1 and 100% in G2 and G3.ConclusionsA single application of Bravecto® Plus spot-on solution at a dose of 2.0 mg moxidectin + 40 mg fluralaner/kg bodyweight reliably prevents cat aelurostrongylosis for at least 12 weeks.Graphical

The Role of Chrysoperla carnea (Steph.) (Neuroptera: Chrysopidae) as a Potential Dispersive Agent of Noctuid Baculoviruses.

Simple SummaryBaculoviruses (BV) infect several lepidopteran pests of economic importance, such as the beet armyworm Spodoptera exigua. The joint use of microbiological and macrobiological strategies may improve the efficacy of control. Laboratory bioassays were developed to evaluate the interactions between two BVs: the multiple nucleopolyhedroviruses of S. exigua (SeMNPV) and Autographa californica (AcMNPV), and the predator Chrysoperla carnea. The excretion products of the predator’s larvae (drops) and adults (meconia) were microscopically examined after the ingestion of BV-infected S. exigua larvae. For both types of excreta and BVs, viral occlusion bodies (OBs) (resistance forms) were observed. These OBs were infective to healthy S. exigua larvae when applied in water suspension and in direct deposition. The virulence of meconia was higher in suspensions (higher viral load), while larval drops were more virulent in direct application due to their liquid nature and their easiness of consumption. The fitness of C. carnea was slightly affected by the consumption of both BV-infected prey. No preference was shown between healthy and BV-infected S. exigua, and both were preferred vs. the aphid Macrosiphum euphorbiae. Our findings present C. carnea, and particularly its larvae, as a promissory candidate for BV dispersion in the field.Baculoviruses (BV) are highly effective against lepidopteran pests of economic importance such as Spodoptera exigua. The combined use of entomopathogens and macrobiological control agents requires the study of their relationships. Laboratory bioassays were developed to evaluate the interactions between the multiple nucleopolyhedroviruses of S. exigua (SeMNPV) and Autographa californica (AcMNPV), and the predator Chrysoperla carnea. The microscopic examination of predator’s excreta (larval drops and meconia) after the ingestion of BV-infected S. exigua revealed the presence of viral occlusion bodies (OBs). The reinfection of S. exigua larvae with BVs-contaminated excreta by using OBs water suspensions or by direct application both yielded high mortality values but different speed-of-kill results. Meconia killed before in suspensions due to their higher viral load and larval excretion drops did so in direct application due to their liquid nature and their easiness of consumption. The prey-mediated ingestion of SeMNPV and AcMNPV triggered slight effects in C. carnea, which were probably derived from the food nutritional quality. Chrysoperla carnea larvae did not discriminate between healthy and BV-infected S. exigua, while a preference was shown for S. exigua (healthy or infected) vs. Macrosiphum euphorbiae. Our findings present C. carnea, and particularly its larvae, as a promissory candidate for BV dispersion in the field.

Human toxocariasis, a silent helminthic disease revealed in Savannakhet, Lao PDR

Toxocariasis is a zoonotic helminthiasis caused by the migrating larvae of Toxocara canis and T. cati, common roundworms of dogs and cats. Our previous study in Savannakhet Province of Lao PDR showed an infection rate of 44.1% of Toxocara spp. in dogs. Thus, we investigate if this previous high prevalence in the definitive hosts influenced the occurrence of human toxocariasis. For that we used a 38 kDa recombinant protein derived from T. canis larvae excretion secretion products in ELISA. Human serum samples were collected in the Lahanam area of Savannakhet province. The population attending the study in Lahanam village were aged from 7 to 59 years old (y/o) 65.9% (54/82) were male and 34.1% (28/82) were female. The total percentage of seropositivity to Toxocara sp. was 30.4% (25/82). Males were more likely to test positive for toxocariasis with a risk ratio of 2.70 (CI95 0.87–4.93). No significant differences between ages were seen. However, it was possible to observe an increase of optical density (OD) values in ELISA according to age. The awareness of the health system on the high prevalence of seropositivity to Toxocara sp. in Savannakhet can prevent irreversible consequences as permanent vision loss and seizures caused by this silent chronic disease revealed in the Lahanam area.

Efficacy of a topical combination of eprinomectin, praziquantel, fipronil and (S)-methoprene against developing and adult Troglostrongylus brevior lungworms (Nematoda, Crenosomatidae) in cats

The efficacy of the eprinomectin, praziquantel, fipronil and (S)-methoprene combination parasiticide Broadline® (Boehringer Ingelheim Animal Health) was evaluated against developing larval and adult stages of Troglostrongylus brevior, a metastrongyloid pulmonary nematode which is reported to parasitize domestic cats in southern Europe with increasing frequency.Twenty four purpose-bred cats were experimentally infected with 100 third-stage T. brevior larvae each and randomly allocated to either remain untreated (control) or to be treated with the combination product when T. brevior were developing larval (6 days post inoculation, dpi) or adult nematodes (28 dpi) (eight cats per group). Treatments were administered topically at the minimum label dose of 0.12mL/kg. Fecal samples of the cats were examined to confirm the presence of patent (adult) nematode infections prior to treatment at 28 dpi and to monitor the larval excretion. At necropsy (49 dpi), the weight of the pulmonary lymph nodes and lungs were determined, and T. brevior lungworms were recovered and counted.All control animals and cats to be treated at 28 dpi excreted T. brevior larvae 24 dpi and 26 dpi while no larvae were excreted by the cats treated at 6 dpi. Following treatment at 28 dpi, T. brevior larval excretion decreased immediatetly and ceased prior to necropsy. Nematode counts demonstrated that treatment with the combination product was 100 % efficacious against both developing larval and adult T. brevior: no lungworms were recovered from any treated cat while all control animals harbored T. brevior (range, 6–52) (p<0.001).No treatment-related health problems or any other clinical signs were observed in the cats. However, significantly higher absolute and relative (organ weight to body weight ratio) pulmonary lymph node weights of the control animals compared with the treated cats at 6 dpi (p<0.001 and p<0.001, respectively) and at 28 dpi (p=0.003 and p=0.019, respectively) indicated the pathology of the T. brevior infection.In conclusion, the combination product was demonstrated to be 100 % efficacious against developing larval and adult T. brevior. Furthermore, it was demonstrated that indicators of impaired respiratory and immune systems resultant from T. brevior infection can be prevented with an efficacious treatment when administered during the pre-patent period of infection or are improving substantially within three weeks of treatment of cats harboring adult lungworms.

Alcohol consumption alters anti-Strongyloides stercoralis antibodies production.

Individuals infected with Strongyloides stercoralis have been reported to produce different immunoglobulins isotypes, yet few studies have evaluated their use in strongyloidiasis diagnosis. The aim of this work was to evaluate the immunoreactivity of different classes and subclasses of anti-S. stercoralis circulating antibodies in alcoholic patients by ELISA and to perform immunoblotting in samples with discordant results between parasitological and immunological methods. 345 male patients with a clinical diagnosis of alcoholism hospitalized at a reference center for alcoholics in Salvador, Bahia, Brazil, were included in this study. The fecal samples were examined by three different parasitological methods (spontaneous sedimentation, Baermann-Moraes and Agar Plate Culture methods). The ELISA was performed for the detection of IgG, IgG1, IgG4, IgE and IgA1 anti-S. stercoralis. Immunoblotting, for the detection of specific IgA1, was used to elucidate discordant results between parasitological and immunological methods. S. stercoralis infection frequency in alcoholic patients by parasitological methods was 21.4% (74/345). Although IgE-ELISA demonstrated a high sensitivity and specificity in non-alcoholic patients, about 30% (22/74) of alcoholics with larvae in feces were negative. IgG1-ELISA detected the lowest frequency of antibodies in alcoholic patients with larvae in feces, only 57% (42/74). IgG4-ELISA was the best assay for S. stercoralis infection immunodiagnosis. Immunoreactivity in the immunoblotting for IgA1 at 90, 75, 26 and/or 17 kDa bands was observed in 92% (33/36) of alcoholics with larvae excretion and negative ELISA for one or more antibody isotypes. In conclusion, IgG4-ELISA showed the highest sensitivity and specificity, thus demonstrating its superiority for strongyloidiasis immunodiagnosis in alcoholic and non-alcoholic individuals. Both, IgE and IgG1-ELISA presented high sensitivities and specificities for S. stercoralis infection diagnosis in non-alcoholics, however there was low reactivity in alcoholic individuals. This can be associated with an increased susceptibility to severe strongyloidiasis in these patients. IgA1-immunoblotting can be used to confirm S. stercoralis infection when there are discordant results between parasitological methods and ELISA.

Bactericidal Activity of Chrysomya rufifacies and Cochliomyia macellaria (Diptera: Calliphoridae) Larval Excretions-Secretions Against Staphylococcus aureus (Bacillales: Staphylococcaceae).

The inhibitory effect of Chrysomya rufifacies (Macquart) and Cochliomyia macellaria (Fabricius) larval excretions-secretions (ES) on Staphylococcus aureus was determined using a portable colorimetric method without the need for any dedicated spectral instrument. Polystyrene 96 well microplates were used and 100 μl of the bacterial inoculum (5 × 105 CFU/ml) plus 100 μl of the dipteran exosecretions at different concentrations were added to each well. Subsequently, 50 μl of a 1% solution of the triphenyl tetrazolium chloride stain was added to each well to determine the bacterial viability. The color development in each well was measured with the ImageJ software S. aureus was exposed to different concentrations of the ES of both species individually. At a concentration of 800 ppm ES of C. rufifacies or Co. macellaria, bacterial growth was inhibited 97.45 ± 1.70% and 82.21 ± 1.88%, respectively. As expected, exposure to a lower concentration (i.e., 50 ppm) was less inhibitory (C. rufifacies ES, 77.65 ± 4.25% and Co. macellaria ES, 43.54 ± 4.63%). This study demonstrates for the first time the bactericidal activity of C. rufifacies and Co. macellaria ES against S. aureus. This finding is promising as it could result in the identification and synthesis of proteins capable of suppressing pathogen development in wounds. Additionally, the proposed method can simplify the use of expensive laboratory instruments for antimicrobial activity determination.

Sarconesin II, a New Antimicrobial Peptide Isolated from Sarconesiopsis magellanica Excretions and Secretions.

Antibiotic resistance is at dangerous levels and increasing worldwide. The search for new antimicrobial drugs to counteract this problem is a priority for health institutions and organizations, both globally and in individual countries. Sarconesiopsis magellanica blowfly larval excretions and secretions (ES) are an important source for isolating antimicrobial peptides (AMPs). This study aims to identify and characterize a new S. magellanica AMP. RP-HPLC was used to fractionate ES, using C18 columns, and their antimicrobial activity was evaluated. The peptide sequence of the fraction collected at 43.7 min was determined by mass spectrometry (MS). Fluorescence and electronic microscopy were used to evaluate the mechanism of action. Toxicity was tested on HeLa cells and human erythrocytes; physicochemical properties were evaluated. The molecule in the ES was characterized as sarconesin II and it showed activity against Gram-negative (Escherichia coli MG1655, Pseudomonas aeruginosa ATCC 27853, P. aeruginosa PA14) and Gram-positive (Staphylococcus aureus ATCC 29213, Micrococcus luteus A270) bacteria. The lowest minimum inhibitory concentration obtained was 1.9 μM for M. luteus A270; the AMP had no toxicity in any cells tested here and its action in bacterial membrane and DNA was confirmed. Sarconesin II was documented as a conserved domain of the ATP synthase protein belonging to the Fli-1 superfamily. The data reported here indicated that peptides could be alternative therapeutic candidates for use in infections against Gram-negative and Gram-positive bacteria and eventually as a new resource of compounds for combating multidrug-resistant bacteria.

Sarconesin: Sarconesiopsis magellanica Blowfly Larval Excretions and Secretions With Antibacterial Properties.

Larval therapy (LT) is an alternative treatment for healing chronic wounds; its action is based on debridement, the removal of bacteria, and stimulating granulation tissue. The most important mechanism when using LT for combating infection depends on larval excretions and secretions (ES). Larvae are protected against infection by a spectrum of antimicrobial peptides (AMPs); special interest in AMPs has also risen regarding understanding their role in wound healing since they degrade necrotic tissue and kill different bacteria during LT. Sarconesiopsis magellanica (Diptera: Calliphoridae) is a promising medically-important necrophagous fly. This article reports a small AMP being isolated from S. magellanica ES products for the first time; these products were obtained from third-instar larvae taken from a previously-established colony. ES were fractionated by RP-HPLC using C18 columns for the first analysis; the products were then lyophilised and their antimicrobial activity was characterized by incubation with different bacterial strains. These fractions’ primary sequences were determined by mass spectrometry and de novo sequencing; five AMPs were obtained, the Sarconesin fraction was characterized and antibacterial activity was tested in different concentrations with minimum inhibitory concentrations starting at 1.2 μM. Potent inhibitory activity was shown against Gram-negative (Escherichia coli D31, E. coli DH5α, Salmonella enterica ATCC 13314, Pseudomonas aeruginosa 27853) and Gram-positive (Staphylococcus aureus ATCC 29213, S. epidermidis ATCC 12228, Micrococcus luteus A270) bacteria. Sarconesin has a significant similarity with Rho-family GTPases which are important in organelle development, cytoskeletal dynamics, cell movement, and wound repair. The data reported here indicated that Sarconesin could be an alternative candidate for use in therapeutics against Gram-negative and Gram-positive bacterial infections. Our study describes one peptide responsible for antibacterial activity when LT is being used. The results shown here support carrying out further experiments aimed at validating S. magellanica AMPs as novel resources for combating antibacterial resistance.

The metabolic fate of dietary nicotine in the cabbage looper, Trichoplusia ni (Hübner)

Cabbage looper (Trichoplusia ni) larvae are generalist herbivores that feed on numerous cultivated plants and weeds including crucifers, other vegetables, flowers, and field crops. Consuming plant material from a wide range of plant species exposes these larvae to a considerable variety of plant secondary metabolites involved in chemical defense against herbivory. The ability of the cabbage looper larvae to detoxify plant secondary metabolites, such as nicotine, has been attributed to the rapid induction of excretion via the Malpighian tubules. However, the role of metabolism in the detoxification of plant secondary metabolites in cabbage looper larvae is not well studied. We investigated nicotine metabolism in 4th larval instar cabbage looper using UPLC-MS/MS analysis to resolve the time course of nicotine metabolism, the kinetic distribution of nicotine, and the presence or absence of major metabolites of nicotine in larval tissue and excretions. The major metabolite found in our analysis was cotinine, with trace amounts of cotinine N-oxide and nicotine N-oxide. The nicotine metabolites detected are similar to those of the nicotine-tolerant Lepidopteran tobacco hornworm (Manduca sexta). The results of our study demonstrate that the 5′C-oxidation of nicotine to cotinine is the primary pathway for nicotine metabolism in cabbage looper larvae. This study showed that metabolism of nicotine and subsequent excretion of nicotine and its metabolites occurs in the larvae of the cabbage looper. Our results suggest that 5′C-oxidation in lepidopteran insects is a conserved metabolic pathway for the detoxification of plant secondary metabolites.

Identification and Preliminary Evaluation of a Novel Recombinant Protein for Serodiagnosis of Strongyloidiasis.

Strongyloides stercoralis is a human parasite that can cause a long-term infection. In immunosuppressed patients, strongyloidiasis may be fatal when there is overwhelming autoinfection resulting in the migration of large numbers of larvae through many organs. Definitive diagnosis is still a challenge, and a combination of symptoms, microscopic identification, and serology test results are often used to arrive at a clinical decision. However, intermittent larval excretion, low parasite burden, and occult infections are challenges with parasitological diagnosis of infection with S. stercoralis. Meanwhile, serologic tests using immunoglobulin G and parasite antigen extract have problems of cross-reactivity with other helminthic infections. Recombinant antigen-based serodiagnosis is a good alternative to overcome the laboratory diagnostic issues. Herein, we report on the isolation of cDNA clone encoding an antigen of potential diagnostic value identified from immunoscreening of a S. stercoralis cDNA library. The translated protein had highest similarity to Strongyloides ratti immunoglobulin-binding protein 1. The recombinant antigen produced, rSs1a, was assessed using western blot and enzyme-linked immunosorbent assay. The latter showed 96% diagnostic sensitivity and 93% specificity; thus, rSs1a has good potential for use in serodiagnosis of human strongyloidiasis.

The effect of patent Dictyocaulus viviparus (re)infections on individual milk yield and milk quality in pastured dairy cows and correlation with clinical signs

BackgroundInfections with the bovine lungworm Dictyocaulus viviparus might lead to reduced milk production and detrimental impacts on milk quality resulting in considerable economic losses in dairy farming.MethodsIn the presented field study, 1988 faecal samples were collected from 1166 Black and White dairy cows allocated in 17 small and medium-sized German grassland farms. Faecal samples were collected in summer and autumn 2015 to assess D. viviparus larvae excretion. Test-day records were used to estimate the association between patent D. viviparus infections in individual cows and the milk production parameters milk yield, milk protein and milk fat content by using linear mixed models. Bulk tank milk (BTM) samples from each farm and individual milk samples from those cows which were excreting larvae in summer were collected in autumn. In addition, occurrence of the clinical symptom “coughing” was noted in individual cows during autumn sampling to determine its association with patent lungworm infections.ResultsPatent D. viviparus infections were found on 23.5% (4/17) of farms with a prevalence at the individual cow level of 0.9% (9/960) in summer and 3.4% (35/1028) in autumn. No BTM sample exceeded the BTM ELISA cut-off value of 0.410 optical density ratio (ODR), the mean value was 0.168 ODR. Only one individual milk sample exceeded the individual milk ELISA cut-off value of 0.573 ODR (mean value of 0.302 ODR). A patent D. viviparus infection status was associated with a lower average daily milk yield of 1.62 kg/cow/day (P = 0.0406). No significant association was found with milk protein or fat content representing milk quality parameters. Coughing was observed in 5.9% (61/1028) of cows. Of the coughing cows, only 4.9% (3/61) had a patent lungworm infection. Fisher’s exact test showed no significant difference between infected and non-infected coughing cows.ConclusionsFarmers and veterinarians should be aware that patent lungworm (re)infections in dairy cows reduce milk yield, despite the absence of clinical signs. Furthermore, if dairy cows present with coughing, other differential diagnoses need to be considered in addition to dictyocaulosis.