The brain, a lung, a tumor: each contains a complex array of cell types arranged in an intricate spatial pattern that is critical to their role within the tissue. While work is underway to create an atlas of cellular diversity across tissues, traditional methods either have sufficiently high multiplexing capacity but lack spatial context, or they provide spatial information but lack the multiplexing capacity to identify subtle variations in cell type or state. This limitation precludes a comprehensive view across a whole tissue, including rare cells, as they exist within a tissue of interest. Since understanding how the molecular and cellular organization of a biological tissue contributes to its functionality is fundamental to developing treatments that target the cell biology underlying diseased states, improved tools that map biological systems with both high spatial resolution and high multiplexing capacity promise to deliver new, clinically relevant insights into the functional composition of biological systems. The recently developed multiplexed error-robust fluorescence in situ hybridization (MERFISH) method provides information about gene expression at subcellular resolution and reveals cell type, state, and cell-cell interaction by quantifying the expression and exact location of hundreds to thousands of transcripts within intact tissues. MERFISH represents a major advancement toward directly interrogating rich cellular heterogeneity and is being applied to gain insight into the biological complexity within and around us.

We examine a model of network formation in single-layer and multiplex networks in which individuals have positive incentives for social ties, closed triangles, and spillover edges. In particular, we investigate the influence of shocks to the network in which the cost of social ties changes after an initial equilibrium. We highlight the emergence of structural entrenchment: the retention of structural features, such as closed triangles and spillover edges, which are formed under historically different conditions from those currently driving network evolution. This work has broad implications for understanding path dependence in the structure and dynamics of single-layer and multiplex networks.

Cryptosporidium is a well-known cause of neonatal enteritis in cattle worldwide. Cattle are commonly infected with four different species of Cryptosporidium but only one of these, Cryptosporidium parvum, is associated with clinical disease. Identification of species in cases of calf scour can give an indication if Cryptosporidium is the causative agent or not. In addition, C. parvum is a zoonotic species and so has implications for human health, for this reason it is important to identify the species of Cryptosporidium infecting cattle particularly where a farm is implicated in an outbreak of cryptosporidiosis in humans. Here a multiplex PCR test, which can identify the four common cattle-adapted Cryptosporidium species, including C. parvum, has been developed. This test allows quick and accurate detection of Cryptosporidium species in cattle fecal samples including mixed infections, which could be missed by the more common method of sequencing the same gene.

Non-availability of adult worms from living hosts remains a key problem in population genetic studies of schistosomes. Indirect sampling involving passage through laboratory animals presents significant ethical and practical drawbacks, and may result in sampling biases such as bottlenecking processes and/or host-induced selection pressures. The novel techniques reported here for sampling, storage and multi-locus microsatellite analysis of larval Schistosoma mansoni, allowing genotyping of up to 7 microsatellite loci from a single larva, circumvent these problems. The utility of these assays and the potential problems of laboratory passage, were evaluated using 7 S. mansoni population isolates collected from school-children in the Hoima district of Uganda, by comparing the associated field-collected miracidia with adult worms and miracidia obtained from a single generation in laboratory mice. Analyses of laboratory-passaged material erroneously indicated the presence of geographical structuring in the population, emphasizing the dangers of indirect sampling for population genetic studies. Bottlenecking and/or other sampling effects were demonstrated by reduced variability of adult worms compared to their parent field-collected larval samples. Patterns of heterozygote deficiency were apparent in the field-collected samples, which were not evident in laboratory-derived samples, potentially indicative of heterozygote advantage in establishment within laboratory hosts. Genetic distance between life-cycle stages in the majority of isolates revealed that adult worms and laboratory-passaged miracidia clustered together whilst segregating from field miracidia, thereby further highlighting the utility of this assay.

This study reports the development of a novel multiplex PCR assay based on SCAR (Sequence-Characterised Amplified Region) markers for the simultaneous diagnosis of the 7 Eimeria species that infect domestic fowl. Primer pairs specific for each species were designed in order to generate a ladder of amplification products ranging from 200 to 811 bp. Sensitivity tests for each species were carried out, showing a detection threshold of 1–5 pg, which corresponds approximately to 2–8 sporulated oocysts. Distinct isolates of the 7 Eimeria species from different geographical sources were tested and successfully detected by the assay. All the species were amplified homogeneously, whether or not one of them was present in a high quantity, indicating that there was no cross-interference. The assay was also tested with different sources of Taq DNA polymerase and thermocycler models, confirming the high reproducibility of the reaction. The economy of consumables and labour represented by a single-tube reaction greatly facilitates the molecular diagnosis of a large number of samples, making it appropriate for field epizootiological surveys. We propose the use of this multiplex PCR assay as a rapid and cost-effective diagnostic method for the detection and discrimination of the 7 Eimeria species that infect domestic fowl.