Agata Pijl

In this study, we adopt an interdisciplinary approach, integrating agronomic field experiments with soil chemistry, molecular biology techniques, and statistics to investigate the impact of organic residue amendments, such as vinasse (a by-product of sugarcane ethanol production), on soil microbiome and greenhouse gas (GHG) production. The research investigates the effects of distinct disturbances, including organic residue application alone or combined with inorganic N fertilizer on the environment. The methods assess soil microbiome dynamics (composition and function), GHG emissions, and plant productivity. Detailed steps for field experimental setup, soil sampling, soil chemical analyses, determination of bacterial and fungal community diversity, quantification of genes related to nitrification and denitrification pathways, measurement and analysis of gas fluxes (N₂O, CH₄, and CO₂), and determination of plant productivity are provided. The outcomes of the methods are detailed in our publications (Lourenço et al., 2018a; Lourenço et al., 2018b; Lourenço et al., 2019; Lourenço et al., 2020). Additionally, the statistical methods and scripts used for analyzing large datasets are outlined. The aim is to assist researchers by addressing common challenges in large-scale field experiments, offering practical recommendations to avoid common pitfalls, and proposing potential analyses, thereby encouraging collaboration among diverse research groups. • Interdisciplinary methods and scientific questions allow for exploring broader interconnected environmental problems. • The proposed method can serve as a model and protocol for evaluating the impact of soil amendments on soil microbiome, GHG emissions, and plant productivity, promoting more sustainable management practices. • Time-series data can offer detailed insights into specific ecosystems, particularly concerning soil microbiota (taxonomy and functions).

Plant Growth-Promoting Microbes (PGPM) have the potential to enhance sustainable agriculture, but there is still a limited understanding of how the complex interplay between plant genetic variability, the native soil community, and soil nutrients affects PGPM recruitment. To address this challenge, we investigated the impact of bacteria isolates and arbuscular mycorrhizal fungi (AMF) along with their accompany microbiome (AMFc) derived from a wild chrysanthemum on the growth of five different commercial chrysanthemum cultivars (Chic, Chic 45, Chic Cream, Haydar and Barolo), as well as their rhizosphere microbiomes, within a nutrient-rich complex substrate environment. We found 23 bacterial strains capable of producing siderophore, 14 strains capable of producing Indole-3-acetic acid, and 18 strains capable of solubilizing phosphate. The AMFc had six AMF species, and the bacterial and fungal communities associated with AMF belonged to different phyla. Using generalized joint models, we investigated the impact of the three most effective bacterial strains and the AMFc on plant growth (shoot and root dry mass) while integrating information on plant genotype, environment, and microbes. The impact of PGPM inoculation varied from positive to negative effects depending on the cultivar, with Chic Cream showing the highest increase in root biomass after inoculation with both bacterial strain SMF006 (57 %) and AMFc inoculation (79 %). Our study demonstrates that PGPM from wild relative can impact the growth and assembly of the chrysanthemum root microbiome, but this impact is cultivar-dependent. Furthermore, inoculation with a complex AMF containing community (AMFc) induced greater changes in the rhizosphere microbiome than with a single bacterial isolate. Our study shows that inoculation of a complex community of beneficial microbes results in more effective plant growth promotion.

Climate change will strongly affect the developmental timing of insects, as their development rate depends largely on ambient temperature. However, we know little about the genetic mechanisms underlying the temperature sensitivity of embryonic development in insects. We investigated embryonic development rate in the winter moth (Operophtera brumata), a species with egg dormancy which has been under selection due to climate change. We used RNA sequencing to investigate which genes are involved in the regulation of winter moth embryonic development rate in response to temperature. Over the course of development, we sampled eggs before and after an experimental change in ambient temperature, including two early development weeks when the temperature sensitivity of eggs is low and two late development weeks when temperature sensitivity is high. We found temperature-responsive genes that responded in a similar way across development, as well as genes with a temperature response specific to a particular development week. Moreover, we identified genes whose temperature effect size changed around the switch in temperature sensitivity of development rate. Interesting candidate genes for regulating the temperature sensitivity of egg development rate included genes involved in histone modification, hormonal signalling, nervous system development and circadian clock genes. The diverse sets of temperature-responsive genes we found here indicate that there are many potential targets of selection to change the temperature sensitivity of embryonic development rate. Identifying for which of these genes there is genetic variation in wild insect populations will give insight into their adaptive potential in the face of climate change.

Unfortunately, Table 1 was incorrectly published in the original article and the same is corrected by this Erratum. The original article was corrected.

The most common etiologic agents of genital ulcer disease (GUD) are herpes simplex virus type 1 (HSV-1), HSV-2, Treponema pallidum, and Haemophilus ducreyi. In an outpatient clinic for sexually transmitted diseases in Amsterdam, The Netherlands, specimens from 372 patients with GUD were collected from February to November 1996. Sera were collected at the time of the symptoms and, for most patients, also during follow-up visits. Swabs in viral transport medium were used for HSV culture and for detection of DNA. The most prevalent pathogen found was HSV-2, which was detected by culture in 35% of the patients and by PCR in 48% of the patients. Also, HSV-1 infection was more often detected by PCR (7.8%) than by culture (5.6%). Evidence for an active infection with T. pallidum was found in 1.9% of the patients, using serological tests. A multiplex PCR for simultaneous T. pallidum and H. ducreyi DNA detection was positive for T. pallidum in 3.3% of the samples and for H. ducreyi in only 0.9% (3 out of 368) of the samples. The sensitivity of the PCR was superior to that of culture for HSV detection and to that of serology for T. pallidum detection. Specific H. ducreyi immunoglobulin G antibodies were detected in sera of 5.2% of the patients, with no concordance between serology and PCR. In 37% of the cases, none of the tested microorganisms was detected. Performance of PCR in addition to conventional techniques significantly improved the diagnosis of GUD.

The transmission of sporadic community-acquired hepatitis A virus (HAV) among different risk groups in Amsterdam was verified by applying molecular techniques on fecal samples. These were collected in 1997/1998 from 33 persons with HAV infection that was confirmed serologically. From 8 of these persons serial stool samples were collected. Nested RT-PCR targeting the VP3-VP1 and VP1-P2a regions followed by sequence analysis established the duration of fecal HAV RNA excretion in stool and the epidemiological molecular relationships between patients. The samples of 31 patients were RT-PCR positive, of which 24 were positive for both regions. Fecal HAV shedding was found to occur for at least 33 days after onset of disease, which was the longest time span tested. Sequencing showed that the hepatitis A virus subgenotype circulating among persons from Moroccan descent (type IB) was different from the subgenotype circulating among Dutch homosexual men (type IA). If the latter is endemic in the Netherlands, its presence is of importance to the national vaccination strategy.

Two assays for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae were compared: the LCx Probe system (the LCx system; Abbott Diagnostic Laboratories, North Chicago, Ill.) and the COBAS AMPLICOR C. trachomatis/N, gonorrhoeae system (the COBAS AMPLICOR system; Roche Diagnostic Systems, Branchburg, N.J.). Endocervical swab specimens, male urethral swab specimens, and female and male urine specimens were collected from 503 female and 498 male visitors attending a sexually transmitted diseases clinic in Amsterdam, The Netherlands. Prevalences for C. trachomatis were 12.5% (63 of 503) and 10.0% (50 of 498) in females and males, respectively. The prevalences for N. gonorrhoeae were 1.2% (6 of 503) and 4.2% (21 of 498) in females and males, respectively. Both assays showed high values for sensitivity and specificity with regard to the detection of C. trachomatis in endocervical swab specimens, male urethral swab specimens, and female and male urine specimens. The sensitivities for the LCx system were 92.1, 90.0, 88.9, and 94.0% for each type of specimen, respectively; and the sensitivies for the COBAS AMPLICOR system were 96.8, 98.0, 82.5, and 92.0% for each type of specimen, respectively. Specificities ranged between 98.4 and 100%. The sensitivity of the LCx system for the detection of N. gonorrhoeae was 100% for female cervical swab and urine specimens and male urethral swab specimens, while for male urine specimens the sensitivity was 95.2%; the specificity was 100% for all types of specimens. For the detection of N. gonorrhoeae by the COBAS AMPLICOR assay, the sensitivity for female cervical swab and male urethral swab specimens was 100%, that for female urine specimens was 66.7%, and that for male urine specimens was 95.2%. However, the predictive values of a positive test for female cervical swab specimens and urine specimens were 31.6 and 36.4%, respectively. Sequence analysis of the amplimers obtained by an in-house 16S rRNA PCR of the solely COBAS AMPLICOR system-positive swab specimens revealed neither N. gonorrhoeae nor other Neisseria spp. The COBAS AMPLICOR assay was considered not suitable for screening for infections with N. gonorrhoeae. If this assay is used for detection of N. gonorrhoeae, confirmation of positive results by a reliable test is mandatory.