Kaplan, Oktay I. et al. (2019) International Worm Meeting "Functional characterisation of Joubert Syndrome associated CEP-41 and ARMC-9."

Kilic, Atiye, Altunkaynak, Betul, Pir, Mustafa S., Pir, Betul, Cevik, Sebiha, Yenisert, Ferhan, Kaplan, Oktay I.

[International Worm Meeting, 2019]

Joubert syndrome is a rare genetic disorder with an occurrence of approximately 1/100 000 live births. Developmental defects in cerebellum and brainstem are observed in patients with JS, and JS is defined as a neurodevelopmental disorder. Up to now 35 different genes have been linked to JS, and the functions of many of these genes including CEP41 and ARMC9 remain uncharacterised. To elucidate the functions of Joubert Syndrome associated CEP-41 (F42G8.19) and ARMC-9 (F59G1.4) genes, we have generated transgenics containing their own promoter driven GFP labelled constructs and generated/obtained null mutants for these genes. CEP-41 was found to be exclusively expressed in the ciliated sensory neurons, and its localisation is restricted to the middle segment of cilia while ARMC-9 localizes to the entire length of the ciliary axonemes. While null mutants for these genes possess no abnormal cilia structures, a number of double mutant combinations of cep-41 with other ciliopathy genes reveal defects in cilia structure in C. elegans.

Cevik, Sebiha et al. (2021) MicroPubl Biol

Cevik, Sebiha, Kaplan, Oktay I.

[MicroPubl Biol, 2021]

We report the identification of C. elegans F42G8.19 as the ortholog of human CEP41. This conclusion was based on database searches, reciprocal BLAST analysis, and cilia-specific localization. To look for the orthologous gene of human CEP41 in C. elegans, we used three databases: the model organism Alliance of Genome Resources (https://www.alliancegenome.org/, Release 3.2.0), OrthoList 2 (http://ortholist.shaye-lab.org/, Release 2017), and ConVarT (https://convart.org/, Release 2020) (Kim et. al., 2018; Pir et. al., 2021). On the Alliance of Genome Resources and ConVarT, we discovered that C. elegans F42G8.19 is the ortholog of human CEP41. To validate this conclusion, we ran a manual reciprocal BLAST analysis (the Reciprocal Best Hits BLAST (RBHB)). The protein-protein BLAST (BLASTp) search of human CEP41 protein sequence (NP 061188.1) revealed F42G8.19 as the top C. elegans hit (Altschul et. al., 1990). In the following step, the protein sequence from the best hit C. elegans F42G8.19 (NP_001294206.1) was compared to human proteins, with human CEP41 emerging as the best match. C. elegans F42G8.19 encodes a 182-amino-acid protein that is shorter than the 373-amino-acid human CEP41 protein (Figure 1A). We subsequently performed the amino acid alignments of human CEP41 (169373 amino acid) and C. elegans F42G8.19 (1182 amino acid), which revealed over 24 % identity to each other (Figure 1A). We eventually determined whether C. elegans F42G8.19 has a rhodanese domain as human CEP41 (169-266 aa), and our query showed that C. elegans F42G8.19 has a rhodanese domain (30-121 aa) (Figure 1A) (Lee et. al., 2012). Taken together, our analysis revealed that C. elegans F42G8.19 is orthologous to human CEP41, and C. elegans F42G8.19 was therefore assigned CEPH-41 (CEntrosomal Protein Homolog 41).

Cevik, Sebiha et al. (2021) MicroPubl Biol

Cevik, Sebiha, Kaplan, Oktay I.

[MicroPubl Biol, 2021]

C. elegans EGL-36 displays extensive similarity to human KCNC proteins, the conserved potassium channel subfamily KV3 (the Shaw subfamily). This potassium channel subfamily contains four genes KCNC1, KCNC2, KCNC3, and KCNC4 encoding KV3.1 (Shaker), KV3.2 (Shab), KV3.3 (Shaw), and KV3.4 (Shal), respectively. Previous studies have demonstrated that C. elegans EGL-36 is expressed in the egg-laying muscles, ventral cord neurons, and the subset of ciliated sensory neurons including ADEL, ADER, PHB through its subcellular localization in the ciliated sensory neurons remains unknown (Elkes et al. 1997; Johnstone et al. 1997). To observe the subcellular localization of EGL-36 in the ciliated sensory neurons in C. elegans, we tagged green fluorescent protein (GFP) at the C-termini of EGL-36 with a cilia-specific promoter. We observed overexpressed EGL-36::GFP in the subsets of sensory neurons in the head and tail including PHB sensory neurons in the tail but not in the PHA sensory neuron, which is in line with previous findings (Elkes et al. 1997; Johnstone et al. 1997) (Figure 1A). Confocal laser scanning microscopy shows that EGL-36::GFP is predominantly located at the base of the cilium. The EGL-36::GFP signal can be found in the neuronal cell body, with weak dendrite staining, where they may function in the dendritic excitability (Figure 1A).

Zorluer, Atiyye et al. (2021) International Worm Meeting "Use of C. elegans for investigating functional consequence of orthologous variants"

Kaplan, Oktay I., Zorluer, Atiyye, Cevik, Sebiha

[International Worm Meeting, 2021]

The widespread use of next-generation sequencing technologies has resulted in the emergence of millions of new genetic variants in humans and non-human species. One of the most important key tasks in genomics is to differentiate harmful genetic variants from non-harmful variants to diagnose disease better. Computational prediction tools and experimental data have helped to shed light on the functional importance of human genetic variants in this respect. However, mutants bearing human equivalent variants, known as orthologous variants (OrthoVar), have gotten less attention, even though they can be beneficial resources for unraveling the functional data for human OrthoVars. Here we used ConVarT (https://convart.org/), an OrthoVar search engine, to look for human OrthoVars in C. elegans. We selected ift-140, which encodes ciliopathy associated intraflagellar transport 140, because a null mutation causes easily detectable cilia abnormality in C. elegans. We identified conserved amino-acid positions that undergo amino acid changes within IFT-140 and discovered seven different variants that fall into this category, three of which have human OrthoVars. We obtained variant carrying mutants from CGC generated by the Million Mutation project and performed a fluorescent dye uptake assay, which indirectly assesses cilia structure. Except for V444I, none of these mutants exhibit Dye uptake defect, indicating that these variants are likely non-harmful variants. V444I mutants have a temperature-sensitive Dye uptake defect as well as IFT protein accumulations in the cilia, which is similar to the ift-140 null mutant phenotype. We are currently performing a rescue analysis for this variant. We then generated two mutants with human OrthoVars (G680S, P702A), and our thorough analysis found that the P702A variant (Human P726A, reported likely pathogenic) is a null mutant of ift-140, but not G680S (Human G704S, VUS). Our current OrthoVar-focused study has uncovered the functional implications of nine variants (5 human OrthoVars), and we believe that functional assessment of human OrthoVars can provide valuable insight into human variants.

Qi Z et al. (2021) Aging (Albany NY) "Sulforaphane promotes C. elegans longevity and healthspan via DAF-16/DAF-2 insulin/IGF-1 signaling."

Ji H, Li H, Wieland A, Bauer S, Le M, Herr I, Qi Z, Wink M, Liu L

[Aging (Albany NY), 2021]

The broccoli-derived isothiocyanate sulforaphane inhibits inflammation, oxidative stress and cancer, but its effect on healthspan and longevity are unclear. We used the <i>C. elegans</i> nematode model and fed the wildtype and 9 mutant strains +/-sulforaphane. The lifespan, phenotype, pharyngeal pumping, mobility, lipofuscin accumulation, and RNA and protein expression of the nematodes were assessed by using Kaplan-Meier survival analysis, <i>in vivo</i> live imaging, fluorescence microscopy, and qRT-PCR. Sulforaphane increased the lifespan and promoted a health-related phenotype by increasing mobility, appetite and food intake and reducing lipofuscin accumulation. Mechanistically, sulforaphane inhibited DAF-2-mediated insulin/insulin-like growth factor signaling and its downstream targets AGE-1, AKT-1/AKT-2. This was associated with increased nuclear translocation of the FOXO transcription factor homolog DAF<i>-</i>16. In turn, the target genes <i>sod-3</i>, <i>mtl-1</i> and <i>gst-4</i>, known to enhance stress resistance and lifespan, were upregulated. These results indicate that sulforaphane prolongs the lifespan and healthspan of <i>C. elegans</i> through insulin/IGF-1 signaling. Our results provide the basis for a nutritional sulforaphane-enriched strategy for the promotion of healthy aging and disease prevention.

Souza ACR et al. (2018) Genes (Basel) "Pathogenesis of the Candida parapsilosis Complex in the Model Host Caenorhabditis elegans."

Colombo AL, Fuchs BB, Jayamani E, Mylonakis E, Souza ACR, Alves VS

[Genes (Basel), 2018]

<i>Caenorhabditis</i><i>elegans</i> is a valuable tool as an infection model toward the study of <i>Candida</i> species. In this work, we endeavored to develop a <i>C</i>. <i>elegans</i>-<i>Candida</i><i>parapsilosis</i> infection model by using the fungi as a food source. Three species of the C. parapsilosis complex (<i>C.</i><i>parapsilosis</i> (<i>sensu</i><i>stricto</i>), <i>Candida</i><i>orthopsilosis</i> and <i>Candida</i><i>metapsilosis</i>) caused infection resulting in <i>C. elegans</i> killing. All three strains that comprised the complex significantly diminished the nematode lifespan, indicating the virulence of the pathogens against the host. The infection process included invasion of the intestine and vulva which resulted in organ protrusion and hyphae formation. Importantly, hyphae formation at the vulva opening was not previously reported in <i>C</i>. <i>elegans</i>-<i>Candida</i> infections. Fungal infected worms in the liquid assay were susceptible to fluconazole and caspofungin and could be found to mount an immune response mediated through increased expression of <i>cnc</i>-<i>4</i>, <i>cnc</i>-<i>7</i>, and <i>fipr</i><i>-</i><i>22</i>/<i>23</i>. Overall, the <i>C</i>. <i>elegans</i>-<i>C</i>. <i>parapsilosis</i> infection model can be used to model <i>C</i>. <i>parapsilosis</i> host-pathogen interactions.

Fourie R et al. (2021) Front Cell Infect Microbiol "Candida albicans SET3 Plays a Role in Early Biofilm Formation, Interaction With Pseudomonas aeruginosa and Virulence in Caenorhabditis elegans."

Sebolai O, Fourie R, Albertyn J, Pohl CH, Gcilitshana O

[Front Cell Infect Microbiol, 2021]

The yeast <i>Candida albicans</i> exhibits multiple morphologies dependent on environmental cues. <i>Candida albicans</i> biofilms are frequently polymicrobial, enabling interspecies interaction through proximity and contact. The interaction between <i>C. albicans</i> and the bacterium, <i>Pseudomonas aeruginosa</i>, is antagonistic <i>in vitro, with P. aeruginosa</i> repressing the yeast-to-hyphal switch in <i>C. albicans</i>. Previous transcriptional analysis of <i>C. albicans</i> in polymicrobial biofilms with <i>P. aeruginosa</i> revealed upregulation of genes involved in regulation of morphology and biofilm formation, including <i>SET3</i>, a component of the Set3/Hos2 histone deacetylase complex (Set3C). This prompted the question regarding the involvement of <i>SET3</i> in the interaction between <i>C. albicans</i> and <i>P. aeruginosa</i>, both <i>in vitro</i> and <i>in vivo.</i> We found that <i>SET3</i> may influence early biofilm formation by <i>C. albicans</i> and the interaction between <i>C. albicans</i> and <i>P. aeruginosa</i>. In addition, although deletion of <i>SET3</i> did not alter the morphology of <i>C. albicans</i> in the presence of <i>P. aeruginosa</i>, it did cause a reduction in virulence in a <i>Caenorhabditis elegans</i> infection model, even in the presence of <i>P. aeruginosa.</i>

Zhu Q et al. (2020) Oxid Med Cell Longev "A Dihydroflavonoid Naringin Extends the Lifespan of C. elegans and Delays the Progression of Aging-Related Diseases in PD/AD Models via DAF-16."

Zhou XG, Qu Y, Luo HR, Zhu Q, Wu GS, Chen JN

[Oxid Med Cell Longev, 2020]

Naringin is a dihydroflavonoid, which is rich in several plant species used for herbal medicine. It has a wide range of biological activities, including antineoplastic, anti-inflammatory, antiphotoaging, and antioxidative activities. So it would be interesting to know if naringin has an effect on aging and aging-related diseases. We examined the effect of naringin on the aging of <i>Caenorhabditis elegans</i> (<i>C</i>. <i>elegans</i>). Our results showed that naringin could extend the lifespan of <i>C</i>. <i>elegans</i>. Moreover, naringin could also increase the thermal and oxidative stress tolerance, reduce the accumulation of lipofuscin, and delay the progress of aging-related diseases in <i>C</i>. <i>elegans</i> models of AD and PD. Naringin could not significantly extend the lifespan of long-lived mutants from genes in insulin/IGF-1 signaling (IIS) and nutrient-sensing pathways, such as <i>daf</i>-<i>2</i>, <i>akt</i>-<i>2</i>, <i>akt</i>-<i>1</i>, <i>eat</i>-<i>2</i>, <i>sir</i>-<i>2</i>.<i>1</i>, and <i>rsks</i>-<i>1</i>. Naringin treatment prolonged the lifespan of long-lived <i>glp</i>-<i>1</i> mutants, which have decreased reproductive stem cells. Naringin could not extend the lifespan of a null mutant of the fox-head transcription factor DAF-16. Moreover, naringin could increase the mRNA expression of genes regulated by <i>daf</i>-<i>16</i> and itself. In conclusion, we show that a natural product naringin could extend the lifespan of <i>C</i>. <i>elegans</i> and delay the progression of aging-related diseases in <i>C</i>. <i>elegans</i> models via DAF-16.

Coomansingh-Springer C et al. (2019) Heliyon "Internal parasitic burdens in brown rats (Rattus norvegicus) from Grenada, West Indies."

Sharma RN, Armstrong E, Vishakha V, Acuna AM, Coomansingh-Springer C

[Heliyon, 2019]

This study identified the endoparasites in Brown rat (<i>Rattus norvegicus)</i> during May to July 2017 in Grenada, West Indies. A total of 162 rats, 76 females and 86 males were trapped from St. George and St. David parishes in Grenada. The collected fecal samples were examined for parasitic eggs and/or oocysts using simple fecal flotation technique. Adult parasites found in the intestinal tract were examined for identification. The overall prevalence of intestinal parasites among rats was 79 %. Ten helminth species were recovered, several of which were reported for the first time in rodents in Grenada. The internal parasites consist of seven nematodes (<i>Angiostrongylus</i> spp., <i>Nippostrongylus braziliensis</i>, <i>Heterakis spumosa</i>, <i>Strongyloides ratti</i>, <i>Aspiculuris tetraptera</i>, <i>Syphacia</i> spp. and <i>Protospirura</i> spp.), one cestode (<i>Hymenolepsis diminuta</i>), one acanthocephalan (<i>Moniliformis moniliformis</i>) and one protozoa species (<i>Eimeria</i> spp.). The most prevalent zoonotic species were <i>Angiostrongylus</i> spp. (35.2%), <i>Hymenolepsis diminuta</i> (7.4%) and <i>Moniliformis moniliformis</i> (3.1%). Several nonzoonotic endoparasites; which included <i>Nippostrongylus braziliensis</i> (50.6%), <i>Heterakis spumosa</i> (15.4%), <i>Strongyloides ratti</i> (43.2%), <i>Aspiculuris tetraptera</i> (2.5%), <i>Syphacia</i> spp<i>.</i> (1.9%), <i>Protospirura</i> spp. (1.2%) and <i>Eimeria</i> spp. (4.7%) were also identified. The most prevalent parasites were <i>Nippostrongylus brasiliensis</i> (50.6%), <i>Strongyloides ratti</i> (43.2%) and <i>Angiostrongylus spp.</i> (35.2%). Co-infections occurred with up to six species per rat showing different combinations of parasitic infections.

Wu CJ et al. (2019) Appl Environ Microbiol "Genotypic and Phenotypic Characteristics of Aeromonas Isolates from Fish and Patients in Tainan City."

Shu CY, Li CW, Ko WC, Su YC, Chen YW, Lee NY, Su SL, Wu CJ, Chen PL, Li MC, Lin YT

[Appl Environ Microbiol, 2019]

The present study aimed to isolate <i>Aeromonas</i> from fish sold in the markets as well as in sushi and seafood shops and compare their virulence factors and antimicrobial characteristics with those of clinical isolates. Among the 128 fish isolates and 47 clinical isolates, <i>A. caviae</i>, <i>A. dhakensis</i>, and <i>A. veronii</i> were the principal species. <i>A. dhakensis</i> isolates carried at least 5 virulence genes, more than other <i>Aeromonas</i> species. The predominant genotype of virulence genes was <i>hlyA/lip/alt/col/el</i> in both <i>A. dhakensis</i> and <i>A. hydrophila</i> isolates, <i>alt/col/ela</i> in <i>A. caviae</i> isolates, and <i>act</i> in <i>A. veronii</i> isolates. <i>A. dhakensis</i>, <i>A. hydrophila</i>, and <i>A. veronii</i> isolates more often exhibited hemolytic and proteolytic activity and showed greater virulence than <i>A. caviae</i> in <i>Caenorhabditis elegans</i> and the C2C12 cell line. However, the link between the genotypes and phenotypes of the studied virulence genes in <i>Aeromonas</i> species is not evident. Among the four major clinical <i>Aeromonas</i> species, nearly all (99.0%) <i>A. dhakensis</i>, <i>A. hydrophila</i>, and <i>A. veronii</i> isolates harbored <i>bla</i>_CphA, which encodes a carbapenemase, but only a minority (6.7%, 7/104) were nonsusceptible to carbapenem. Regarding AmpC -lactamase genes, <i>bla</i>_AQU-1 was exclusively found in <i>A. dhakensis</i> isolates and <i>bla</i>_MOX3 only in <i>A. caviae</i> isolates, but only 7.6% (6) of the 79 <i>Aeromonas</i> isolates carrying <i>bla</i>_AQU-1 or <i>bla</i>_MOX3 exhibited a cefotaxime resistance phenotype. In conclusion, fish <i>Aeromonas</i> isolates carry a variety of combinations of virulence and B-lactamase resistance genes and exhibit virulence phenotypes and antimicrobial resistance profiles similar to those of clinical isolates.<b>IMPORTANCE</b><i>Aeromonas</i> species can cause severe infections in immunocompromised individuals upon exposure to virulent pathogens in the environment, but the characteristics of environmental <i>Aeromonas</i> species remain unclear. Our study showed several pathogenic <i>Aeromonas</i> species possessing virulence traits and antimicrobial resistance similar to those of <i>Aeromonas</i> isolates causing clinical diseases were present in fish intended for human consumption in Tainan City.