Antibacterial and bacteriostatic potential of coelomic fluid and body paste of Pheretima posthuma (Vaillant, 1868) (Clitellata, Megascolecidae) against ampicillin resistant clinical bacterial isolates

Pheretima posthuma (Vaillant, 1868), a native earthworm of Pakistan and Southeast Asia, has wide utilization in vermicomposting and bioremediation process. In this study, P. posthuma coelomic fluid (PCF) and body paste (PBP) was evaluated as antibacterial agent against ampicillin (AMP) resistant five Gram positive and four Gram negative clinical isolates. The antibacterial effect of different doses (i.e. 25-100 µg/ml) of PCF and PBP along with AMP and azithromycin (AZM) (negative and positive controls, respectively) were observed through disc diffusion and micro-dilution methods. All nine clinical isolates were noticed as AMP resistant and AZM sensitive. Antibacterial effects of PCF and PBP were dose dependent and zone of inhibitions (ZI) against all clinical isolates were between 23.4 ± 0.92 to 0 ± 00 mm. The sensitivity profile of PCF and PBP against clinical isolates was noticed as 44.44 and 55.56%, respectively. Both PCF and PBP showed bacteriostatic (BTS) action against S. aureus, S. pyogenes, K. pneumonia, N. gonorrhoeae. Moreover, the cumulative BTS potential of PCF and PBP against all isolates was 66.67 and 55.56%, respectively. The MICs of PCF and PBP were ranged from 50-200 µg/ml against selected isolates. The bacterial growth curves indicated that PCF and PBP inhibited the growth of all isolates at their specific MIC concentrations. However, PBP has better antibacterial potential compared to PCF against selected isolates. Therefore, it is concluded that both PCF and PBP of P. posthuma possess antibacterial and BTS potential against ampicillin resistant clinical isolates. This organism might be considered as a second choice of antibacterial agents and can further be utilized in pharmaceutical industries for novel drug manufacturing by prospecting bioactive potential agents.

Preferential Begging Responses of Shiny Cowbirds to the Conspecific Chatter Call

Avian brood parasites lay their eggs into the nests of other species, which incubate them and raise the chicks until their independence. Despite living their early weeks of life surrounded by heterospecifics, young brood parasites have the ability to recognize and associate to conspecifics after independence. It has been proposed that the initial conspecific recognition develops when a young parasite encounters a unique species-specific signal that triggers the learning of other aspects of the producer of the signal. For cowbirds (Molothrus spp.), this species-specific signal is hypothesized to be the chatter call. Young birds also could express auditory biases, which in some cases lead to discrimination in favor of conspecific songs. Therefore, the perceptual selectivity for chatters might be also present in nestlings. Our aim was to assess if nestlings of the shiny cowbird (M. bonariensis) present a preferential begging response to conspecific chatter calls. We evaluated if they respond more to the parasitic vocalization than host chicks and if they respond more to the chatter than to heterospecific nonhost calls. We tested shiny cowbird chicks reared by chalk-browed mockingbirds (Mimus saturninus) or house wrens (Troglodytes aedon) and host chicks, as control species. We randomly presented to 6-day-old chicks the following playback treatments: (1) conspecific chatter calls, (2) host calls, used as positive controls, and (3) nonhost calls, used as negative control. We measured if chicks begged during the playback treatments and the begging intensity. When responding to the playback of chatter calls, shiny cowbird chicks begged at a higher frequency and more intensively than host chicks. Shiny cowbird chicks reared by mockingbirds begged more intensively to playbacks of conspecific chatter calls than to host calls, while those reared by wrens begged with a similar intensity to playbacks of conspecific chatter and host calls. On the contrary, wren nestlings begged more intensively to playbacks of the wren call than to chatter calls. Mockingbird nestlings did not beg during any treatment. None of the three species begged during the playback of nonhost calls. Our results show that the chatter call produced a preferential begging response in cowbird nestlings, which may be the beginning of a process of conspecific recognition.

Evaluating the Fertilising Potential of Blended Recovered Nutrients in Horticultural Growing Medium on Viola x wittrockiana L.

Viola x wittrockiana L. is an ornamental plant in high demand in horticulture. It is becoming more critical for greenhouse growers to focus on sustainable production to enhance plant quality while reducing negative environmental impacts. Therefore, assessing the effect of recycled phosphorous (P) and nitrogen (N) sources on the growth of viola could become very useful for producers in terms of sustainability. This experiment analysed the optimal fertiliser composition to grow viola using recovered fertilisers in a greenhouse trial under controlled conditions. Well-rooted viola plugs were grown in a standard peat-based growing medium. Using recycled sources of P and N as struvite and potassium struvite, ammonium sulphate, and ammonium nitrate, 14 fertiliser blends were prepared, tested, and compared with the slow-release commercial fertiliser Osmocote. Plants treated with ammonium nitrate showed healthy growth and optimal plant N concentrations. In contrast, most blends using the recovered ammonium sulphate resulted in an unacceptable increase of ammonium concentrations in the growing medium. The combination of ammonium sulphate and potassium sulphate caused an increase in the electrical conductivity in the growing medium, negatively affecting plant growth. However, blend 13 containing struvite, ammonium sulphate and potassium struvite expressed the best chemical composition with non-significant differences in the biomass from the positive controls, as it reduced the amount of potassium sulphate needed. Our results indicate that fertiliser blends containing P as struvite, N as ammonium nitrate or reduced amount of ammonium sulphate, and K as potassium struvite can substitute the use of mineral fertiliser blends to grow ornamental plant species as viola.

Contamination of personal protective equipment during COVID-19 autopsies

Confronted with an emerging infectious disease at the beginning of the COVID-19 pandemic, the medical community faced concerns regarding the safety of autopsies on those who died of the disease. This attitude has changed, and autopsies are now recognized as indispensable tools for understanding COVID-19, but the true risk of infection to autopsy staff is nevertheless still debated. To clarify the rate of SARS-CoV-2 contamination in personal protective equipment (PPE), swabs were taken at nine points in the PPE of one physician and one assistant after each of 11 full autopsies performed at four centers. Swabs were also obtained from three minimally invasive autopsies (MIAs) conducted at a fifth center. Lung/bronchus swabs of the deceased served as positive controls, and SARS-CoV-2 RNA was detected by real-time RT-PCR. In 9 of 11 full autopsies, PPE samples tested RNA positive through PCR, accounting for 41 of the 198 PPE samples taken (21%). The main contaminated items of the PPE were gloves (64% positive), aprons (50% positive), and the tops of shoes (36% positive) while the fronts of safety goggles, for example, were positive in only 4.5% of the samples, and all the face masks were negative. In MIAs, viral RNA was observed in one sample from a glove but not in other swabs. Infectious virus isolation in cell culture was performed on RNA-positive swabs from the full autopsies. Of all the RNA-positive PPE samples, 21% of the glove samples, taken in 3 of 11 full autopsies, tested positive for infectious virus. In conclusion, PPE was contaminated with viral RNA in 82% of autopsies. In 27% of autopsies, PPE was found to be contaminated even with infectious virus, representing a potential risk of infection to autopsy staff. Adequate PPE and hygiene measures, including appropriate waste deposition, are therefore essential to ensure a safe work environment.

Nested PCR for the Diagnosis of Feline Sporotrichosis From Formalin-Fixed and Paraffin-Embedded Samples Using Different DNA Extraction Protocols

Sporotrichosis is a chronic, cosmopolitan granulomatous mycosis that affects humans and animals. The infection is caused by the dimorphic fungi Sporothrix sp. The aims of the present study were to evaluate, standardize and validate a nested PCR technique using two DNA purification kits for the extraction of DNA from formalin fixed and paraffin-embedded tissues (FFPE) for Sporothrix sp. detection. FFPE mycological culture pellet samples of different Sporothrix species (S. chilensis, S. mexicana, S. pallida, S. globosa, S. brasiliensis and S. schenckii) were used as positive controls and clinical FFPE tissue samples of animals positive for Cryptococcus sp., Leishmania infantum and Histoplasma sp. were used as negative controls. Ten clinical FFPE skin samples from cats with sporotrichosis were used to validate the nested PCR. These samples were cut into two distinct paraffin sectioning protocols (5 and 16 μm thick). The paraffin sections were subjected to two different DNA extraction kits (chemical and thermal extractions). A nested PCR was performed on the extracted DNA to identify the genus Sporothrix. The chemical extraction protocol with the 5 μm thick paraffin section was more effective in extracting DNA from Sporothrix sp. from FFPE samples and the nested PCR technique showed the highest sensitivities (100% in the positive controls and of 50% in the skin samples of cats) and specificity (100%). Therefore, the nested PCR using this protocol has great potential to be applied in Sporothrix sp. diagnosis in FFPE samples of cats.

Influenza Vaccine Effectiveness Estimates among US Department of Defense Adult Beneficiaries over Four Consecutive Influenza Seasons: A Test-Negative Design Study with Different Control Groups

A test-negative design study with different control groups (influenza test-negative controls, non-influenza virus positive controls, and pan-negative controls) was conducted to assess inactivated influenza vaccine effectiveness (VE) in adults aged ≥18 years, 2016–2017 through 2019–2020 influenza seasons. A database was developed from the US Department of Defense Global Respiratory Pathogen Surveillance Program. VE was estimated using a generalized linear mixed model with logit link and binomial distribution, adjusted for confounding effects. A total of 7114 adults including 2543 medically attended, laboratory-confirmed influenza-positive cases were identified. Using influenza test-negative controls, the adjusted VE in adults was 40% [95% confidence interval (CI): 33–46%] overall, including 46% (95% CI: 36–55%) for influenza A(H1N1)pdm09, 32% (95% CI: 19–42%) for influenza A(H3N2), and 54% (95% CI: 44–62%) for influenza B. The age-stratified analysis showed that VE estimates against influenza A(H1N1)pdm09 (34%; 95% CI: −29–66%) and influenza A(H3N2) (6%; 95% CI: −60–45%) were low and non-significant for elderly adults ≥65 years of age. Overall VE estimates against any influenza or by influenza (sub)types in adults were consistent when using influenza test-negative controls, non-influenza virus positive controls, and pan-negative controls. Inactivated influenza vaccination provided moderate protection against influenza virus infection, based on the analysis from a large number of adults aged ≥18 years over multiple influenza seasons.

Design of primers in the molecular detection of Feline Panleukopenia Virus

Feline Panleukopenia is a disease characterized by a reduction in the number of circulating leukocytes and enteritis with degeneration of the intestinal villi. The etiologic agent, called Feline Panleukopenia virus (FPV), belongs to the Parvoviridae family, is highly contagious and has high mortality and morbidity. Although vaccination of healthy cats is the most effective way to prevent the disease, once the symptoms appear, the treatment is supportive, presenting high mortality in the first days of the disease. FPV positive cats should be hospitalized and isolated for at least 2 weeks to avoid viral transmission. Early detection is usually done with the enzyme-linked immunoadsorption assay (ELISA) test that detects viral antigens in stool samples. As a complementary diagnostic method, in this work it was proposed to implement the Polymerase Chain Reaction (PCR) aimed at the diagnosis of FPV initially in positive samples from two feline vaccines and one canine vaccine. As an approximation to real samples, the commercial vaccines were mixed with feces and blood from a healthy cat. The results showed that the In Silico design was successful strategy based on the VP2 gene sequences of FPV available in GenBank® in conjunction with the sharp visualization of positive controls of the expected size and without amplification. nonspecific or negative controls. The fragments obtained has a nucleotide identity percentage greater than 97% with respect to the information available in Genbank® and corroborates the detection of a VP2 fragment. Thus, the future option of applying the same protocol in a diagnostic way is opened, using samples obtained from suspected patients who are infected with FPV.

Ateleia glazioveana and Ocimum basilicum: plants with potential larvicidal and repellent against Aedes aegypti (Diptera, Culicidae)

The Aedes aegypti is the main vector of the transmission of diseases such as dengue, yellow fever, Chikungunya, and Zika. The agents used for the control of mosquito involve the use of synthetic compounds, which may cause toxicity and contamination environmental. Aiming to test the effectiveness of natural products, the aim of this study was to evaluate the larvicidal and repellent activity of extracts of Ateleia glazioveana Baill. (timbó) and essential oil of Ocimum basilicum L. (manjericão), against A. aegypti. The activities of hydroalcoholic (HEA) and dichloromethane (DEA) extracts from A. glazioveana against A. aegypti larvae were assessed after exposure for 24, 48, and 72 h, using water and Bacillus thuringiensis israelensis as negative and positive controls, respectively. For repellent evaluation, we investigated the sting attempts of A. aegypti in response to the application of HEA- and DEA-based body creams (5%), as well as an alcoholic spray based on the essential oil from O. basilicum. The repellency of these preparations was compared with negative (vehicle) and positive (DEET) controls. Although HEA showed no apparent larvicidal activity, DEA at concentrations of 150 to 500 mg/ml showed high larvicidal potential after 24 h, compared to the negative control group (p < 0.001). Moreover, the DEA-based cream and the alcoholic spray of O. basilicum based essential oil showed higher repellency compared with the vehicle (84.5% and 70.5%, respectively). Accordingly, it is possible to infer that extracts and preparations of A. glazioveana and O. basilicum exhibit larvicidal and repellent activity against A. aegypti.

Metagenomic Next-Generation Sequencing of Cerebrospinal Fluid for the Diagnosis of Cerebral Aspergillosis

Purpose: Cerebral aspergillosis (CA) is a rare but often fatal, difficult-to-diagnose, opportunistic infection. The utility of metagenomic next-generation sequencing (mNGS) for diagnosis of CA is unclear. We evaluated the usefulness of mNGS of the cerebrospinal fluid (CSF) for the diagnosis of CA.Methods: This prospective study involved seven consecutive patients with confirmed CA in whom CSF mNGS was performed. Serum (1→3)-β-D-glucan and galactomannan levels were determined, and histopathological examination and mNGS of the CSF were conducted. CSF specimens from three non-infected patients were used as positive controls.Results: mNGS of the CSF was positive in six of the seven confirmed CA cases (85.71% sensitivity). In the cryptococcal meningitis group (control), mNGS of the CSF was positive for Aspergillus in two patients (84.62% specificity). The positive likelihood ratio, negative likelihood ratio, and Youden’s index of mNGS for CA in the CSF were 5.565, 0.169, and 0.7, respectively. Among the six mNGS-positive cases, more than two Aspergillus species were found in four (4/6, 66.67%). In the positive controls, the addition of one A. fumigatus spore yielded a standardised species-specific read number (SDSSRN) of 25.45 by mNGS; the detection rate would be 0.98 if SDSSRN was 2.Conclusion: mNGS facilitates the diagnosis of CA and may reduce the need for cerebral biopsy in patients with suspected CA.Trial Registration Number: Chinese Clinical Trial Registry, ChiCTR1800020442.

Characterization of Fibrous Wollastonite NYAD G in View of Its Use as Negative Standard for In Vitro Toxicity Tests

Today, despite considerable efforts undertaken by the scientific community, the mechanisms of carcinogenesis of mineral fibres remain poorly understood. A crucial role in disclosing the mechanisms of action of mineral fibres is played by in vitro and in vivo models. Such models require experimental design based on negative and positive controls. Commonly used positive controls are amosite and crocidolite UICC standards, while negative controls have not been identified so far. The extensive characterisation and assessment of toxicity/pathogenicity potential carried out in this work indicate that the commercial fibrous wollastonite NYAD G may be considered as a negative standard control for biological and biomedical tests involving mineral fibres. Preliminary in vitro tests suggest that wollastonite NYAD G is not genotoxic. This material is nearly pure and is characterized by very long (46.6 µm), thick (3.74 µm) and non-biodurable fibres with a low content of metals. According to the fibre potential toxicity index (FPTI) model, wollastonite NYAD G is an inert mineral fibre that is expected to exert a low biological response during in vitro/in vivo testing.