Biology subjects

Laue, M.

Publications and source records attributed to Laue, M..

11 recordsLinked to original sources

bioRxiv2026

Cryptosporidium sex employs a complex male fertilization apparatus that is required for continued infection

Most apicomplexan parasites replicate asexually to colonize their mammalian hosts, and sexually in the context of transmission. In contrast, for Cryptosporidium sex is essential to sustain intestinal infection in vivo and therefore is a promising opportunity to prevent and treat the disease. However, the mechanism of sex, which occurs within host enterocytes, is poorly understood, limiting the discovery of targets for such intervention. Here we identify an elaborate fertilization apparatus and numerous of its molecular components in male gametes. We discern two elements, one displayed on the apex, that makes intimate contact with host cells containing female parasites, and a second internal structure, that harbors the membrane fusogen HAP2. Ablation of HAP2 by conditional mutagenesis blocked fertilization and oocyst production, rapidly cured infected mice and left them with enhanced resistance to subsequent challenge. Our studies provide insights into a remarkable fertilization mechanism and highlight specific molecules as targets for translation.

microbiology↗

bioRxiv2026

Cryptosporidium sequentially remodels its single rhoptry into the host interface

The apicomplexan parasite Cryptosporidium is a leading cause of diarrheal disease in young children. Within the intestinal epithelium, Cryptosporidium establishes a unique intracellular niche in the apical brush border of enterocytes. A structurally complex interface between host and parasite acts as a holdfast and enables protein and metabolite transport. How the parasite invades its host cell and builds the interface is poorly understood. Here, we reveal parasite invasion with high temporal and spatial resolution using rigorous molecular markers. We find that sequential discharge of specialized secretory organelles anchors the parasite within the host prior to internalization. The single Cryptosporidium rhoptry persists beyond its initial pre-invasion discharge, acting as a conduit for the secretion of multiple waves of effector proteins. Ultimately, the rhoptry membrane gives rise to the feeder organelle that separates host from parasite cytoplasm. These findings lead us to propose a mechanistic model of Cryptosporidium invasion and intracellular parasitism.

microbiology↗

bioRxiv2026

Dual species interactions shield Campylobacter against multiple antibiotics

BackgroundCampylobacter is a major food-borne pathogen causing diarrhea. In severe cases, treatment typically involves fluoroquinolones (FQ) and macrolides. However, high proportions of FQ resistance (FQR) have made macrolides the remaining first-line treatment option. As part of the Campylobacter surveillance program in Germany, we investigated multidrug resistance (MDR) in clinical Campylobacter samples collected 2010 to 2022. MethodsA total of 6,980 samples was single-colony purified and analyzed for antimicrobial resistance phenotypes. Genome sequencing was performed for 2,912 samples, and antimicrobial resistance genes were mapped. Cultures showing MDR were further analyzed for contaminating DNA and studied using scanning electron microscopy (SEM). FindingsWe found that 453 (6%) Campylobacter samples were resistant to both FQ and macrolides. Two of the C. jejuni samples were resistant to antibiotics from ten different classes. Genome analysis revealed that these samples, despite being derived from single colonies, contained >10% Enterococcus DNA reads. SEM confirmed the presence of coccoid bacteria interspersed with spiral-shaped Campylobacter. Additional culture-based purification resulted in pure C. jejuni isolates that retained FQR but lost macrolide resistance. We further showed that presence of MDR Enterococcus spp. in the mixed samples protected C. jejuni from above-MIC (minimum inhibitory concentration) of several ribosome-targeting antimicrobials whereas pure Campylobacter were susceptible. These findings revealed that Campylobacter may gain antimicrobial resistance advantages through close interactions with Enterococcus spp. InterpretationMDR phenotypes in Campylobacter cultures may arise through close, dual-species interactions with other highly resistant intestinal bacteria, such as MDR Enterococcus spp. and may have practical implications for antibiotic treatment. FundingThis study was funded by the German Ministry of Health dedicated to the National Reference Centre for Salmonella and other Bacterial Enteric Pathogens.

microbiology↗

bioRxiv2026

Label-free detection of individual virus-infected cells using deep learning

Numerous applications in research and medicine rely on reliable identification and quantification of virus-infected cells. Current methods either apply reporter viruses, that often differ from clinical isolates (e. g. cell tropism, immune evasion) or staining approaches, that prevent live-cell experiments and may introduce biases through manual counting. We present a deep learning model for the label-free identification of virus-infected cells on light microscopy images (VAIruScope). To overcome limitations, our pipeline enables an automated quantification of virus-infected cells based on the recognition of cytopathic effects. The method was applied to different cell models and four clinically relevant prototype viruses representing RNA- (influenza A virus), DNA- (human cytomegalovirus, herpes simplex virus-1) and retroviruses (human immunodeficiency virus-1). VAIruScope identified infected cells achieving classification accuracies of up to 96 %. As proof-of-concept, the method was validated using electron microscopy for a wild-type HSV-1. VAIruScope may be applicable to live-cell imaging to investigate infection dynamics.

bioinformatics↗

bioRxiv2024

Recruitment of the cellular lipid transport protein CERT to C. psittaci inclusions regulates the timing of bacterial egress

Egress of intracellular pathogens is highly regulated and carefully timed. For the zoonotic bacterium C. psittaci, the predominant egress pathway is Chlamydia-containing sphere (CCS) formation, a calcium-dependent sequential mechanism including protease activity, inclusion membrane destabilization, intracellular calcium increase, and plasma membrane blebbing. How egress is regulated to ensure that it takes place only after C. psittaci intracellular development is thus far unknown. Here, we show that C. psittaci recruits the cellular ceramide transporter CERT to its inclusion during intracellular development, but this recruitment is reduced at late time points prior to egress. In addition, an early loss of CERT at the inclusion membrane induced by CERT-KO induces premature egress by CCS formation. Complementation of the CERT-KO with different CERT-GFP variants prevents premature egress, except of complementation with a variant lacking the inclusion targeting PH domain, showing that the localization of CERT is critical for CCS formation. The CERT-KO induced premature CCS are formed by the sequential process described for mature CCS, but they contain mostly RBs and are predominantly non-infectious. Thus, our findings suggest that the timing of C. psittaci egress by CCS formation is regulated by the recruitment of CERT to the inclusion. We propose that CERT stabilizes the chlamydial inclusion by the formation of ER-inclusion membrane contact sites during intracellular development, and the loss of CERT recruitment facilitates inclusion membrane destabilization and CCS formation.

microbiology↗

bioRxiv2023

The formation of Chlamydia-containing spheres, a novel non-lytic egress pathway of the zoonotic pathogen Chlamydia psittaci

Egress of intracellular bacteria from host cells and cellular tissues is a critical process during the infection cycle. This egress process is essential for bacteria to spread inside the host and can influence the outcome of an infection. For the obligate intracellular Gram-negative zoonotic bacterium Chlamydia psittaci little is known about the mechanisms resulting in chlamydial egress from the infected epithelium. Here, we describe and characterize a novel non-lytic egress pathway of C. psittaci by formation of Chlamydia-containing spheres (CCS). CCS are spherical, low phase contrast structures surrounded by a phosphatidylserine exposing membrane with specific barrier functions. They contain infectious progeny and morphologically impaired cellular organelles. The formation of CCS shares characteristics of apoptotic cell death including a proteolytic cleavage of the peptide DEVD albeit independent of active caspase-3, an increase in the intracellular calcium concentration of infected cells, followed by blebbing of the plasma membrane and rupture of the inclusion membrane. Finally, infected blebbing cells detach and leave the monolayer thereby forming CCS. These results support that Chlamydia psittaci egresses the epithelial cell by a novel non-lytic egress pathway, a process beneficial for the bacterium, which might influence the outcome of the infection in organisms. ImportanceHost cell egress is essential for intracellular pathogens to spread within an organism and for host-to-host-transmission. Here, we describe CCS formation as a novel egress pathway for the intracellular, zoonotic bacterial pathogen C. psittaci. This non-lytic egress pathway is fundamentally different from previously described Chlamydia egress pathways. Interestingly, CCS formation shares several characteristics of apoptotic cell death. However, the sequence of proteolytic activity, followed by plasma membrane blebbing and the final detachment of a whole phosphatidylserine exposing former host cell is unique for C. psittaci. Thus, CCS formation represents a new egress pathway for intracellular pathogens that could possibly be linked to C. psittaci biology including host tropism, protection from host cell defense mechanisms or bacterial pathogenicity.

microbiology↗

bioRxiv2023

Dynamic ventral disc contraction is necessary for Giardia attachment and host pathology

Giardia lamblia is a common parasitic protist that infects the small intestine and causes giardiasis, resulting in diarrhea, vomiting, weight loss, and malabsorption. Giardiasis leads to cellular damage, including loss of microvilli, disruption of tight junctions, impaired barrier function, enzyme inhibition, malabsorption, and apoptosis. In the host, motile Giardia trophozoites attach to the duodenal microvilli using a unique microtubule organelle called the ventral disc. Despite early observations of disc-shaped depressions in microvilli after parasite detachment, little is known about disc-mediated attachment mechanisms and there little direct evidence showing that parasite attachment causes cellular damage. However, advancements in in vitro organoid models of infection and genetic tools have opened new possibilities for studying molecular mechanisms of attachment and the impact of attachment on the host. Through high-resolution live imaging and a novel disc mutant, we provide direct evidence for disc contraction during attachment, resolving the long-standing controversy of its existence. Specifically, we identify three types of disc movements that characterize contraction, which in combination result in a decrease in disc diameter and volume. Additionally, we investigate the consequences of attachment and disc contractility using an attachment mutant that has abnormal disc architecture. In a human organoid model, we demonstrate that this mutant has a limited ability to break down the epithelial barrier as compared to wild type. Based on this direct evidence, we propose a model of attachment that incorporates disc contraction to generates the forces required for the observed "grasping" of trophozoites on the host epithelium. Overall, this work highlights the importance of disc contractility in establishing and maintaining parasite attachment, leading to intestinal barrier breakdown.

microbiology↗

bioRxiv2023

The domed architecture of Giardias ventral disc is necessary for attachment and host pathogenesis

After ingestion of dormant cysts, the widespread protozoan parasite Giardia lamblia colonizes the host gastrointestinal tract via direct and reversible attachment using a novel microtubule organelle, the ventral disc. Extracellular attachment to the host allows the parasite to resist peristaltic flow, facilitates colonization and is proposed to cause damage to the microvilli of host enterocytes as well as disrupt host barrier integrity. The 9 {micro}m in diameter ventral disc is defined by a highly complex architecture of unique protein complexes scaffolded onto a spiral microtubule (MT) array of one hundred parallel, uniformly spaced MT polymers that bend approximately one and a quarter turns to form a domed structure. To investigate the role of disc-mediated attachment in causing epithelial cell damage, we used a new approach to rapidly create a stable quadruple knockout of Giardia of an essential ventral disc protein, MBP, using a new method of CRISPR-mediated gene disruption with multiple positive selectable markers. MBP quadruple KO mutant discs lack the characteristic domed architecture and possess a flattened "crescent" or horseshoe-shaped conformation that lacks the overlapping region, with severe defects in the microribbon-crossbridge (MR-CB) complex structure. MBP KO mutants are also unable to resist fluid flow required for attachment to inert surfaces. Importantly, MBP KO mutants have 100% penetrance off positive selection, which is essential for quantification of in vivo impacts of disc and attachment mutants with host cells. Using a new gastrointestinal organoid model of pathogenesis, we found that MBP KO infections had a significantly reduced ability to cause the barrier breakdown characteristic of wild-type infections. Overall, this work provides direct evidence of the role of MBP in creating the domed disc, as well as the first direct evidence that parasite attachment is necessary for host pathology, specifically epithelial barrier breakdown.

microbiology↗

bioRxiv2023

First virological and pathological study of Göttingen Minipigs with Dippity Pig Syndrome (DPS)

Dippity Pig Syndrome (DPS) is a well-known but rare complex of clinical signs affecting minipigs, which has not been thoroughly investigated yet. Clinically affected animals show acute appearance of red, exudating lesions across the spine. The lesions are painful, evidenced by arching of the back (dipping), and the onset of clinical symptoms is generally sudden. In order to understand the pathogenesis, histological and virological investigations were performed in affected and unaffected Gottingen Minipigs (GoMPs). The following DNA viruses were screened for using PCR-based methods: Porcine cytomegalovirus (PCMV), which is a porcine roseolovirus (PCMV/PRV), porcine lymphotropic herpesviruses (PLHV-1, PLHV-2, PLHV-3), porcine circoviruses (PCV1, PCV2, PCV3, PCV4), porcine parvovirus 1 (PPV1), and Torque Teno sus virus (TTSuV1, TTSuV2). Screening was also performed for integrated porcine endogenous retroviruses (PERV-A, PERV-B, PERV-C) and recombinant PERV-A/C and their expression as well as for the RNA viruses hepatitis E virus (HEV) and SARS-CoV-2. Eight clinically affected and one unaffected GoMPs were analyzed. Additional unaffected minipigs had been analyzed in the past. The analyzed GoMPs contained PERV-A and PERV-B integrated in the genome, which are present in all pigs and PERV-C, which is present in most, but not all pigs. In one affected GoMPs recombinant PERV-A/C was detected in blood. In this animal a very high expression of PERV mRNA was observed. PCMV/PRV was found in three affected animals, PCV1 was found in three animals with DPS and in the healthy minipig, and PCV3 was detected in two animals with DPS and in the unaffected minipig. Most importantly, in one animal only PLHV-3 was detected. It was found in the affected and unaffected skin, and in other organs. Unfortunately, PLHV-3 could not be studied in all other affected minipigs. None of the other viruses were detected and using electron microscopy, no virus particles were found in the affected skin. This data identified some virus infections in GoMPs with DPS and assign a special role to PLHV-3. Since PCMV/PRV, PCV1, PCV3 and PLHV-3 were also found in unaffected animals, a multifactorial cause of DPS is suggested. However, elimination of the viruses from GoMPs may prevent DPS.

microbiology↗

bioRxiv2020

Morphometry of SARS-CoV and SARS-CoV-2 particles in ultrathin sections of infected Vero cell cultures.

SARS-CoV-2 is the causative of the COVID-19 disease, which has spread pandemically around the globe within a few months. It is therefore necessary to collect fundamental information about the disease, its epidemiology and treatment, as well as about the virus itself. While the virus has been identified rapidly, detailed ultrastructural analysis of virus cell biology and architecture is still in its infancy. We therefore studied the virus morphology and morphometry of SARS-CoV-2 in comparison to SARS-CoV as it appears in Vero cell cultures by using conventional thin section electron microscopy and electron tomography. Both virus isolates, SARS-CoV Frankfurt 1 and SARS-CoV-2 Italy-INMI1, were virtually identical at the ultrastructural level and revealed a very similar particle size distribution with a median of about 100 nm without spikes. Maximal spike length of both viruses was 23 nm. The number of spikes per virus particle was about 30% higher in the SARS-CoV than in the SARS-CoV-2 isolate. This result complements a previous qualitative finding, which was related to a lower productivity of SARS-CoV-2 in cell culture in comparison to SARS-CoV.

microbiology↗

bioRxiv2020

Olfactory transmucosal SARS-CoV-2 invasion as port of Central Nervous System entry in COVID-19 patients

The newly identified severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes COVID-19, a pandemic respiratory disease presenting with fever, cough, and often pneumonia. Moreover, thromboembolic events throughout the body including the central nervous system (CNS) have been described. Given first indication for viral RNA presence in the brain and cerebrospinal fluid and in light of neurological symptoms in a large majority of COVID-19 patients, SARS-CoV-2-penetrance of the CNS is likely. By precisely investigating and anatomically mapping oro- and pharyngeal regions and brains of 32 patients dying from COVID-19, we not only describe CNS infarction due to cerebral thromboembolism, but also demonstrate SARS-CoV-2 neurotropism. SARS-CoV-2 enters the nervous system via trespassing the neuro-mucosal interface in the olfactory mucosa by exploiting the close vicinity of olfactory mucosal and nervous tissue including delicate olfactory and sensitive nerve endings. Subsequently, SARS-CoV-2 follows defined neuroanatomical structures, penetrating defined neuroanatomical areas, including the primary respiratory and cardiovascular control center in the medulla oblongata.

neuroscience↗