Metabolomics to unveil and understand phenotypic diversity between pathogen populations

Visceral leishmaniasis is caused by a parasite called Leishmania donovani, which every year infects about half a million people and claims several thousand lives. Existing treatments are now becoming less effective due to the emergence of drug resistance. Improving our understanding of the mechanisms used by the parasite to adapt to drugs and achieve resistance is crucial for developing future treatment strategies. Unfortunately, the biological mechanism whereby Leishmania acquires drug resistance is poorly understood. Recent years have brought new technologies with the potential to increase greatly our understanding of drug resistance mechanisms. The latest mass spectrometry techniques allow the metabolome of parasites to be studied rapidly and in great detail. We have applied this approach to determine the metabolome of drug-sensitive and drug-resistant parasites isolated from patients with leishmaniasis. The data show that there are wholesale differences between the isolates and that the membrane composition has been drastically modified in drug-resistant parasites compared with drug-sensitive parasites. Our findings demonstrate that untargeted metabolomics has great potential to identify major metabolic differences between closely related parasite strains and thus should find many applications in distinguishing parasite phenotypes of clinical relevance

A single methyltransferase YefA (RlmCD) catalyses both m5U747 and m5U1939 modifications in Bacillus subtilis 23S rRNA

Methyltransferases that use S-adenosylmethionine (AdoMet) as a cofactor to catalyse 5-methyl uridine (m5U) formation in tRNAs and rRNAs are widespread in Bacteria and Eukaryota, and are also found in certain Archaea. These enzymes belong to the COG2265 cluster, and the Gram-negative bacterium Escherichia coli possesses three paralogues. These comprise the methyltransferases TrmA that targets U54 in tRNAs, RlmC that modifies U747 in 23S rRNA and RlmD that is specific for U1939 in 23S rRNA. The tRNAs and rRNAs of the Gram-positive bacterium Bacillus subtilis have the same three m5U modifications. However, as previously shown, the m5U54 modification in B. subtilis tRNAs is catalysed in a fundamentally different manner by the folate-dependent enzyme TrmFO, which is unrelated to the E. coli TrmA. Here, we show that methylation of U747 and U1939 in B. subtilis rRNA is catalysed by a single enzyme, YefA that is a COG2265 member. A recombinant version of YefA functions in an E. coli m5U-null mutant adding the same two rRNA methylations. The findings suggest that during evolution, COG2265 enzymes have undergone a series of changes in target specificity and that YefA is closer to an archetypical m5U methyltransferase. To reflect its dual specificity, YefA is renamed RlmCD

The divergent eukaryote Trichomonas vaginalis has an m7G cap methyltransferase capable of a single N2 methylation

Eukaryotic RNAs typically contain 5′ cap structures that have been primarily studied in yeast and metazoa. The only known RNA cap structure in unicellular protists is the unusual Cap4 on Trypanosoma brucei mRNAs. We have found that T. vaginalis mRNAs are protected by a 5′ cap structure, however, contrary to that typical for eukaryotes, T. vaginalis spliceosomal snRNAs lack a cap and may contain 5′ monophophates. The distinctive 2,2,7-trimethylguanosine (TMG) cap structure usually found on snRNAs and snoRNAs is produced by hypermethylation of an m7G cap catalyzed by the enzyme trimethylguanosine synthase (Tgs). Here, we biochemically characterize the single T. vaginalis Tgs (TvTgs) encoded in its genome and demonstrate that TvTgs exhibits substrate specificity and amino acid requirements typical of an RNA cap-specific, m7G-dependent N2 methyltransferase. However, recombinant TvTgs is capable of catalysing only a single round of N2 methylation forming a 2,7-dimethylguanosine cap (DMG) as observed previously for Giardia lamblia. In contrast, recombinant Entamoeba histolytica and Trypanosoma brucei Tgs are capable of catalysing the formation of a TMG cap. These data suggest the presence of RNAs with a distinctive 5′ DMG cap in Trichomonas and Giardia lineages that are absent in other protist lineages

Methylated constituents of Aedes albopictus poly (A)-containing messenger RNA.

Poly (A)-containing mRNA prepared from cultured mosquito (Aedes albopictus) cells was found to contain methylated 5'-terminal "caps" as well as internal m6A residues. Both type I [m7G(5')ppp(5')Xmp] and type II [m7G(5')ppp(5')XmpYmp] caps were present, at molar ratio of ca five to one. All four common RNA bases were represented in the second position (Xm) of the caps, adenine being the most abundant and N6-methyladenine being absent. The four bases were also represented in the third position (Ym), but here uracil was the predominant base. There was approximately one internal m6A residue for every three caps. These studies demonstrate that mRNA from an invertebrate source can have a methylation pattern comparable with that of mammalian cells in it complexity

Sequence and structure of a methionine transfer RNA from mosquito mitochondria.

We have sequenced a methionine tRNA from mosquito mitochondria, and examined its structure using nucleases S1 and T1 under non-denaturing conditions. The sequence is highly homologous to a putative initiator methionine tRNA gene from Drosophila mitochondria. Its anticodon stem contains a run of three G-C base pairs that is characteristic of conventional initiator tRNAs; however, nuclease S1 analysis suggested an anticodon loop configuration characteristic of conventional elongator tRNAs. We propose that this tRNA can assume both initiator and elongator roles

[[alternative]]The Relationship Between Health Beliefs Of Caregivers And Early Childhood Caries Among Preschool Children

[[abstract]]目的：乳牙的健康影響恆牙發育，由於幼兒的口腔肌肉還沒有發育完全加上缺乏照顧自己的能力，特別需要照顧者協助與養成。本研究主要的目的在瞭解學齡前期幼兒主要照顧者的健康信念情形，進而探討健康信念與幼兒早期性齲齒的相關性。方法：採橫斷式、描述相關性研究設計，以方便取樣，用結構式問卷收集資料，共90位以台北市某醫學中心兒童牙科門診之六足歲以下純乳牙齒列兒童的主要照顧者，所蒐集資料以SPSS for Window17.0統計套裝軟體進行資料的分析，進行描述性和推論性統計分析。結果：1.主要照顧者對幼兒口腔保健的健康信念偏向負面：照顧者認為幼兒牙齒健康在遺傳的體質中已注定，後天的努力很難產生效果並且認為還會換牙，沒必要為幼兒進行乳牙的檢查；多數照顧者同意在照護乳牙方面的能力及時間方面有困難，且當幼兒口腔健康有問題時，政府加強辦理兒童健康保險勝過要求父母想辦法改善幼兒的口腔健康。2.照顧者重視幼兒的口腔醫療檢查更甚於親職清潔照顧：有91.1%的照顧者會帶幼兒去看牙醫接受口腔健康檢查，且定期檢查佔67.8%；雖然有97%的照顧者在睡前會幫幼兒清潔牙齒，但仍有18.9%的幼兒有夜奶的習慣。3.照顧者學習幼兒乳牙保健相關知識可預防早期性齲齒：照顧者有學習與幼兒乳牙保健相關的知識其幼兒患有早期性齲齒顆數較少。建議：針對照顧者，當幼兒進食後，以含氟牙膏刷牙，並於睡前幫幼兒使用牙線並且可使用含氟漱口水。有夜奶情形，可採毛巾代替牙刷來清潔牙齒。盡量減少讓攝取甜食或者採集中時間方式給予。幼兒園可請家長提供牙刷與含氟牙膏，讓幼兒養成刷牙的好習慣。以故事或影片教導幼兒學習牙齒健康的重要性。[[abstract]]Objective: Health of primary teeth affects permanent teeth development. Caregiver need to assist children in oral habit development as they lack of self-care abilities. The main purpose of this research is to understand the health beliefs of the primary caregivers, and investigate the relationship between the health beliefs and the occurrence of early childhood caries amongst preschool children.Method: A cross sectional, descriptive correlational research is used with convenious sampling method, and a structured questionnaire is used as data collection. A total of 90 primary caregivers caring for children under six, with complete primary dentition in a medical center in Taipei are surveyed. Data is then analyzed by SPSS for Window 17.0 to produce descriptive and inferential statistical.Results: 1. Primary caregivers oral health beliefs are somewhat negative, believing that dental check-up is unnecessary due to dentition transition, and majority of the caregivers are unable to invest time and effort into caring for primary teeth. They believe strengthening government’s children health insurance will be more effective than asking parents to improve the oral health of preschool children. 2. Caregivers regard oral examination to be more important than parental oral care: The percentage of caregivers that accompanied children to visit dentists on a regular basis is 67.8%. Although 97% of the caregivers assist with teeth brushing at bedtime, 18.9% of the children still had milk bottle feeding at night. 3. Educating caregivers about oral health care for children with primary dentition can effectively reduce the occurrence of early children caries.Suggestion: 1. Use fluoride toothpaste or mouthwash after eating and floss before sleeping. 2. For bottle feeding at night, a wet towel may replace use for toothbrush. 3. Avoid a grazing diet. 4. Kindergartens should promote toothbrush habit to children and use storytelling or videos to teach them to keep good oral hygiene