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Three stage cultivation process of facultative strain of Chlorella sorokiniana for treating dairy farm effluent and lipid enhancement

Hena S, Fatihah N, Tabassum S, Ismail N

Water Res, 2015 Sep 1;80:346-56.
PMID: 26043271 DOI: 10.1016/j.watres.2015.05.001

Reserve lipids of microalgae are promising for biodiesel production. However, economically feasible and sustainable energy production from microalgae requires optimization of cultivation conditions for both biomass yield and lipid production of microalgae. Biomass yield and lipid production in microalgae are a contradictory problem because required conditions for both targets are different. Simultaneously, the mass cultivation of microalgae for biofuel production also depends extremely on the performance of the microalgae strains used. In this study a green unicellular microalgae Chlorella sorokiniana (DS6) isolated from the holding tanks of farm wastewater treatment plant using multi-step screening and acclimation procedures was found high-lipid producing facultative heterotrophic microalgae strain capable of growing on dairy farm effluent (DFE) for biodiesel feedstock and wastewater treatment. Morphological features and the phylogenetic analysis for the 18S rRNA identified the isolated strains. A novel three stage cultivation process of facultative strain of C. sorokiniana was examined for lipid production.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Fulltext Detection and molecular identification of blood parasites in rodents captured from urban areas of southern Sarawak, Malaysian Borneo

Perison PWD, Amran NS, Adrus M, Anwarali Khan FA

Vet Med Sci, 2022 Sep;8(5):2059-2066.
PMID: 35636429 DOI: 10.1002/vms3.849

BACKGROUND: Rodent species are well known for their potential as hosts and reservoirs for various zoonotic diseases. Studies on blood parasite infection in small mammals focused on urban cities in Peninsular Malaysia and have been conducted over the years. In contrast, there are information gaps related to molecular detection of blood parasites in urban areas of Sarawak that are associated with veterinary importance and zoonotic spillover potential. Increasing prevalence and transmission of blood parasite diseases is the most crucial public health issue, particularly in developing urban areas of Sarawak. Therefore, molecular identification studies were performed to determine and identify the blood parasites infecting rodents.
METHODS: A total of 40 rodent blood samples were analysed for blood parasite infection and a combined approach using polymerase chain reaction-based technique, and traditional microscopic examination (blood smear test) was conducted. 18s rRNA (Plasmodium spp.) and cytochrome b (Hepatocystis spp.) gene marker were used to identify the blood parasites.
RESULTS: Note that 67.5% (n = 27) blood samples were tested negative for blood parasites, while 32.5% (n = 13) blood samples collected were infected with at least one protozoan parasite. Out of 13 samples, 69.2% (n = 9) were detected with Hepatocystis sp., while 15.4% (n = 2) were positive with Hepatozoon ophisauri. Two individuals had multiple infections from both species. No Plasmodium spp. have been detected throughout this study using universal primer (targeted Plasmodium spp.); however, different parasite species which were H. ophisauri were detected.
CONCLUSION: Although there is no evidence of human infection from H. ophisauri and Hepatocystis sp. detected from the study, the data show the host species are heavily infected, and the information is essential for future prevention of zoonotic outbreaks and surveillance programmes. Therefore, it is suggested that the surveillance programmes should be incorporated in targeted areas with a high risk of disease emergence.

Matched MeSH terms: RNA, Ribosomal, 18S
Fulltext Microbiota of Palm Oil Mill Wastewater in Malaysia

Bala JD, Lalung J, Al-Gheethi AAS, Hossain K, Ismail N

Trop Life Sci Res, 2018 Jul;29(2):131-163.
PMID: 30112146 MyJurnal DOI: 10.21315/tlsr2018.29.2.10

This study was aimed at identifying indigenous microorganisms from palm oil mill effluent (POME) and to ascertain the microbial load. Isolation and identification of indigenous microorganisms was subjected to standard microbiological methods and sequencing of the 16S rRNA and 18S rRNA genes. Sequencing of the 16S rRNA and 18S rRNA genes for the microbial strains signifies that they were known as Micrococcus luteus 101PB, Stenotrophomonas maltophilia 102PB, Bacillus cereus 103PB, Providencia vermicola 104PB, Klebsiella pneumoniae 105PB, Bacillus subtilis 106PB, Aspergillus fumigatus 107PF, Aspergillus nomius 108PF, Aspergillus niger 109PF and Meyerozyma guilliermondii 110PF. Results revealed that the population of total heterotrophic bacteria (THB) ranged from 9.5 × 105 - 7.9 × 106 cfu/mL. The total heterotrophic fungi (THF) ranged from 2.1 × 104 - 6.4 × 104 cfu/mL. Total viable heterotrophic indigenous microbial population on CMC agar ranged from 8.2 × 105 - 9.1 × 106 cfu/mL and 1.4 × 103 - 3.4 × 103 cfu/mL for bacteria and fungi respectively. The microbial population of oil degrading bacteria (ODB) ranged from 6.4 × 105 - 4.8 × 106 cfu/mL and the oil degrading fungi (ODF) ranged from 2.8 × 103 - 4.7 × 104 cfu/mL. The findings revealed that microorganisms flourish well in POME. Therefore, this denotes that isolating native microorganisms from POME is imperative for effectual bioremediation, biotreatment and biodegradation of industrial wastewaters.

Matched MeSH terms: RNA, Ribosomal, 18S
Fulltext Light microscopy and molecular identification of Sarcocystis spp. in meat producing animals in Selangor, Malaysia

Latif B, Kannan Kutty M, Muslim A, Hussaini J, Omar E, Heo CC, et al.

Trop Biomed, 2015 Sep;32(3):444-52.
PMID: 26695204 MyJurnal

One thousand and forty-five tissue samples of skeletal muscles, tongue, heart, diaphragm and esophagus were collected from 209 animals (43 sheep, 89 goats and 77 cattle) from an abattoir in Selangor between February and October, 2013. Each sample was divided into three pieces with each piece measuring 2-3 mm3. Each piece was then squeezed between two glass slides and examined microscopically at x 10 magnification for the presence of sarcocystosis. Three positive samples from each animal species were then fixed in 10% formalin for histological processing. Seven positive samples collected from each animal species were preserved at -80°C or 90% ethanol for gene expression studies. Microsarcocysts were detected in 114 (54.5%) animals by light microscopy (LM). The infection rates in sheep, goat and cattle were 86, 61.8 and 28.6% respectively. The highest rate of infection was in the skeletal muscles of sheep (64.9%) and goats (63.6%) and in the heart of cattle (63.6%). The cysts were spindle to oval in shape and two stages were recognized, the peripheral metrocytes and centrally located banana-shaped bradyzoites. 18S rRNA gene expression studies confirmed the isolates from the sheep as S. ovicanis, goats as S. capracanis and cattle as S. bovicanis. This, to the best of our knowledge, is the first molecular identification of an isolate of S. ovicanis and S. capracanis in Malaysia. Further studies with electron microscopy (EM) are required in the future to compare the features of different types of Sarcocysts spp.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Fulltext Molecular epidemiology of Cryptosporidium in HIV/AIDS patients in Malaysia

Asma I, Sim BL, Brent RD, Johari S, Yvonne Lim AL

Trop Biomed, 2015 Jun;32(2):310-22.
PMID: 26691260 MyJurnal

Cryptosporidiosis is a particular concern in immunocompromised individuals where symptoms may be severe. The aim of this study was to examine the epidemiological and molecular characteristics of Cryptosporidium infections in HIV/AIDS patients in Malaysia in order to identify risk factors and facilitate control measures. A modified Ziehl-Neelsen acid fast staining method was used to test for the presence of Cryptosporidium oocysts in the stools of 346 HIV/AIDS patients in Malaysia. Standard coproscopical methods were used to identify infections with other protozoan or helminths parasites. To identify the species of Cryptosporidium, DNA was extracted and nested-PCR was used to amplify a portion of the SSU rRNA gene. A total of 43 (12.4%) HIV-infected patients were found to be infected with Cryptosporidium spp. Of the 43 Cryptosporidium-positive HIV patients, 10 (23.3%) also harboured other protozoa, and 15 (34.9%) had both protozoa and helminths. The highest rates of cryptosporidiosis were found in adult males of Malay background, intravenous drug users, and those with low CD4 T cell counts (i.e., < 200 cells/mm3). Most were asymptomatic and had concurrent opportunistic infections mainly with Mycobacterium tuberculosis. DNA sequence analysis of 32 Cryptosporidium isolates identified C. parvum (84.3%), C. hominis (6.3%), C. meleagridis (6.3%), and C. felis (3.1%). The results of the present study revealed a high prevalence of Cryptosporidium infection in hospitalized HIV/AIDS patients. The results also confirmed the potential significance of zoonotic transmission of C. parvum in HIV infected patients, as it was the predominant species found in this study. However, these patients were found to be susceptible to a wide range of Cryptosporidium species. Epidemiological and molecular characterization of Cryptosporidium isolates provides clinicians and researchers with further information regarding the origin of the infection, and may enhance treatment and control strategies.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Fulltext High prevalence of muscular sarcocystosis in cattle and water buffaloes from Selangor, Malaysia

Latif B, Vellayan S, Heo CC, Kannan Kutty M, Omar E, Abdullah S, et al.

Trop Biomed, 2013 Dec;30(4):699-705.
PMID: 24522140 MyJurnal

The prevalence of sarcocystosis in cattle and water buffaloes from peninsular Malaysia was investigated in abattoirs in Selangor state, February, 2011, to March, 2012. Fresh muscle samples were collected from the tongue, heart, oesophagus, diaphragm and skeletal muscles of 102 cattle and 18 water buffaloes. Each sample was initially screened by light microscopy and then fixed for further histopathological analysis. Out of 120 animals examined, 49 (40.8%) harboured the microscopic type of Sarcocystis spp. The positivity rate for cattle was 36.2% and for water buffaloes 66.7%. In cattle, the organs highly infected were the skeletal muscles and diaphragm (27% each), followed by tongue and esophagus (24.3% each), and the heart (8%). In water buffaloes, the heart was most often infected (66.7%), followed by the oesophagus (50%) and skeletal muscle (33.3%); no sarcocysts were detected in the tongue and diaphragm. The shape of the sarcocyst was fusiform to oval with a mean cyst size of 151.66 x 75.83 μm and wall thickness of 2.47 μm in cattle, and 114 x 50.81 μm cyst size and the wall thickness of 1.11 μm in water buffaloes, consistent with Sarcocystis cruzi and Sarcocystis levinei, respectively. Remaining tissue from cattle was subjected to parasite specific 18S rRNA gene PCR and Sarcocystis cruzi was confirmed, at least exemplarily. The peripheral metrocytes and the banana-shaped bradyzoites (15.23 x 2.2 μm in cattle and 11.49 x 2.45 μm in water buffalo hosts) were easily recognized. In conclusion, a high positivity rate was found in Malaysian meat-producing animals with possible implications for meat consumption and human health.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Fulltext Inferring the phylogenetic position of Brugia pahangi using 18S ribosomal RNA (18S rRNA) gene sequence

Fong MY, Asha T, Azdayanti M, Yee LL, Sinnadurai S, Rohela M

Trop Biomed, 2008 Apr;25(1):87-92.
PMID: 18600209 MyJurnal

This paper presents the first reported use of 18S rRNA gene sequence to determine the phylogeny of Brugia pahangi. The 18S rRNA nucleotide sequence of a Malaysian B. pahangi isolate was obtained by PCR cloning and sequencing. The sequence was compared with 18S rRNA sequences of other nematodes, including those of some filarial nematodes. Multiple alignment and homology analysis suggest that B. pahangi is closely related to B. malayi and Wuchereria bancrofti. Phylogenetic trees constructed using Neighbour Joining, Minimum Evolution and Maximum Parsimony methods correctly grouped B. pahangi with other filarial nematodes, with closest relationship with B. malayi and W. bancrofti. The phylogeny of B. pahangi obtained in this study is in concordance with those previously reported, in which the 5S rRNA gene spacer region and cytochrome oxidase subunit I (COI) sequences were used.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics*
Molecular detection of Candidatus Anaplasma camelii in camels (Camelus dromedarius) from Asir Province, Saudi Arabia

Alshahrani MY, Alanazi AD, Alouffi AS, Abdullah HHAM, Allam AM, Mahmoud MS, et al.

Trop Biomed, 2020 Sep 01;37(3):587-598.
PMID: 33612774 DOI: 10.47665/tb.37.3.587

Knowledge of molecular identification of tick-borne pathogens in camels in Saudi Arabia is very limited; few molecular epidemiological studies have been under taken. This study was to detect Anaplasma spp. and Piroplasma spp. in camels from Asir Province, Saudi Arabia. A total of 150 blood samples were collected from camels in Asir Province and investigated by polymerase chain reaction (PCR) that targeted 18S rRNA and 23S rRNA to detect the DNA of Piroplasma spp. and Anaplasma spp., respectively. The positive samples for 23S rRNA were assayed again by PCR targeting the 16S rRNA. All the blood samples were free from Piroplasma spp. infection. Three camels (2%) were found to be positive for Anaplasma infection through use of PCR that targeted the 23S rRNA gene. There were no significant differences between ages or sexes in the camels that tested positive for Anaplasma. All positive Anaplasma infections were recorded in camels that were infested by ticks. Two Anaplasma sequences for the16S rRNA gene were deposited in GenBank with accession numbers MN882724 and MN882725. They recorded 99.16% and 99.34% similarities (respectively) with KF843825.1 (Candidatus Anaplasma camelii reported in Unizah, Saudi Arabia). Phylogenetic analyses revealed that the two sequences recorded in this study were close to each other; both were located in one cluster with Candidatus Anaplasma camelii isolates that were recorded before in the adjacent areas of Unizah in Saudi Arabia and Iran. In conclusion: two new Anaplasma genotypes close to Candidatus Anaplasma camelii were found in camels in Asir Province, Saudi Arabia for the first time. The camels in this province were found to be free of Piroplasma infection.

Matched MeSH terms: RNA, Ribosomal, 18S
Evaluation of the inhibitory effects of drugs on the growth of Babesia gibsoni using relative quantification real-time PCR

He WH, Feng XX, Wu X, Zhai XH, Li YY, Zhang B, et al.

Trop Biomed, 2020 Dec 01;37(4):871-876.
PMID: 33612740 DOI: 10.47665/tb.37.4.871

To evaluate the inhibitory effects of drugs on the growth of Babesia gibsoni, relative quantification real-time PCR method was developed in this study. The 18S rRNA gene was used as a target gene for the 2-ΔΔCt method analysis. Additionally, chicken RNA was added to the parasitized blood before total RNA extraction. The chicken β-actin gene was selected as an internal control gene for the 2-ΔΔCt method analysis. The 100 µL parasitized blood samples with different percentages of parasitized erythrocytes (PPEs) (3%, 1.5%, 0.75%, 0.375% and 0.1875%) were prepared for relative quantification of B. gibsoni. Regression analysis results revealed significant linear relationships between the relative quantification value and parasitemia. 18S rRNA gene expression was significantly decreased after treatment with diminazene aceturate and artesunate in vitro drug sensitivity test. This result suggested that this relative quantification real-time PCR method can be used to evaluate the effects of drug inhibition.

Matched MeSH terms: RNA, Ribosomal, 18S
Identification of microbial agents in culture-negative brain abscess samples by 16S/18S rRNA gene PCR and sequencing

John DV, Aryalakshmi B, Deora H, Purushottam M, Raju R, Mahadevan A, et al.

Trop Biomed, 2022 Dec 01;39(4):489-498.
PMID: 36602206 DOI: 10.47665/tb.39.4.002

Despite clinical suspicion of an infection, brain abscess samples are often culture-negative in routine microbiological testing. Direct PCR of such samples enables the identification of microbes that may be fastidious, non-viable, or unculturable. Brain abscess samples (n = 217) from neurosurgical patients were subjected to broad range 16S rRNA gene PCR and sequencing for bacteria. All these samples and seven formalin-fixed paraffin-embedded tissue (FFPE) samples were subjected to species-specific 18S rRNA PCR for neurotropic free-living amoeba that harbour pathogenic bacteria. The concordance between smear and/or culture and PCR was 69%. One-third of the samples were smear- and culture-negative for bacterial agents. However, 88% of these culture-negative samples showed the presence of bacterial 16S rRNA by PCR. Sanger sequencing of 27 selected samples showed anaerobic/fastidious gram negative bacteria (GNB, 38%), facultative Streptococci (35%), and aerobic GNB (27%). Targeted metagenomics sequencing of three samples showed multiple bacterial species, including anaerobic and non-culturable bacteria. One FFPE tissue revealed the presence of Acanthamoeba 18S rRNA. None of the frozen brain abscess samples tested was positive for 18S rRNA of Acanthamoeba or Balamuthia mandrillaris. The microbial 16/18S rRNA PCR and sequencing outperformed culture in detecting anaerobes, facultative Streptococci and FLA in brain abscess samples. Genetic analyses of 16S/18S sequences, either through Sanger or metagenomic sequencing, will be an essential diagnostic technology to be included for diagnosing culture-negative brain abscess samples. Characterizing the microbiome of culture-negative brain abscess samples by molecular methods could enable detection and/or treatment of the source of infection.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Nested polymerase chain reaction for detection of Plasmodium falciparum infection in Malaysia

Khoo A, Furuta T, Abdullah NR, Bah NA, Kojima S, Wah MJ

Trans R Soc Trop Med Hyg, 1996 1 1;90(1):40-1.
PMID: 8730308
Matched MeSH terms: RNA, Ribosomal, 18S/genetics*
Detection of Cryptosporidium parvum in HIV-infected patients in Malaysia using a molecular approach

Zaidah AR, Chan YY, Asma HS, Abdullah S, Nurhaslindawati AR, Salleh M, et al.

Southeast Asian J. Trop. Med. Public Health, 2008 May;39(3):511-6.
PMID: 18564692

This cross-sectional study determined the prevalence of cryptosporidiosis in HIV-infected patients using polymerase chain reaction (PCR). Stool specimens were collected from HIV infected patients who were admitted to Hospital Raja Perempuan Zainab II, Kota Bharu, Malaysia, for various indications from December 2004 to December 2005. A modified acid-fast stain was performed on the direct stool smears, then the stool specimens were further tested using nested PCR targeting the 18S rRNA gene of Cryptosporidium parvum, with a built-in internal control (IC). Out of 59 samples, 11 were positives. Nested PCR identified a total of nine samples (16%) compared to microscopy, which identified only three samples. All PCR negative results showed IC amplicons, suggesting that these samples were true negatives and were not due to inhibition of PCR. This study highlights the importance of molecular diagnosis in determining the true prevalence and epidemiology of C. parvum.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Fulltext Detection of a Babesia sp. genotype closely related to marsupial-associated Babesia spp. in male Haemaphysalis shimoga from Sarawak, Malaysian Borneo

Lau AC, Qiu Y, Moustafa MAM, Nakao R, Shimozuru M, Onuma M, et al.

J Vet Med Sci, 2022 Nov 01;84(11):1469-1473.
PMID: 36123016 DOI: 10.1292/jvms.22-0304

In this study, Babesia screening was conducted in 55 rodents and 160 tick samples collected from primary forests and an oil palm plantation in Sarawak, Malaysian Borneo. PCR targeting the 18S ribosomal DNA revealed the presence of Babesia spp. DNA detected in two questing male Haemaphysalis shimoga ticks collected from the oil palm plantation. Sequence analysis revealed that both sequences were identical and had 98.6% identity to a Babesia macropus sequence obtained from Eastern grey kangaroos (Macropus giganteus) in Australia. Phylogenetic tree revealed clustering with marsupial-associated Babesia spp. in the Babesia sensu stricto clade. Whether or not H. shimoga is the competent vector and the importance of the Babesia sp. detected in this study warrants more investigation.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
A new rhabditoid nematode species in Asian sciurids, distinct from Strongyloides robustus in North American sciurids

Sato H, Torii H, Une Y, Ooi HK

J Parasitol, 2007 Dec;93(6):1476-86.
PMID: 18314696 DOI: 10.1645/GE-1106.1

Strongyloides callosciureus n. sp. (Nematoda: Rhabditoidea), from Asian sciurids, is described based on morphology, morphometry, and the small and large subunit (SSU/LSU) ribosomal RNA gene (rDNA) sequences. This new species was collected from Pallas's squirrels (Callosciurus erythraeus) in the central part of mainland Japan (Honshu), which were originally introduced from Taiwan some decades ago, and plantain squirrels (Callosciurus notatus) imported from Malaysia as personal pets. For comparison, Strongyloides robustus Chandler, 1942 was collected from American red squirrels (Tamiasciurus hudsonicus) and southern flying squirrels (Glaucomys volans) imported from the United States as personal pets. The parasitic females found in North American and Asian sciurids shared some key morphological features such as the ovary running spirally around the gut, and the shapes of the stoma in the apical view and the tail. However, morphometric features of parasitic females in North American and Asian sciurids differed significantly from each other; the former was larger than the latter, and the relative position of the vulva to the whole body length from the mouth was different. The SSU/LSU rDNA sequences supported the division of sciurid Strongyloides isolates by geographical distribution of the host and morphological features, leading us to propose the erection of new species.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Effect of condensed tannins on bovine rumen protist diversity based on 18S rRNA gene sequences

Tan HY, Sieo CC, Abdullah N, Liang JB, Huang XD, Ho YW

J. Eukaryot. Microbiol., 2013 Jan-Feb;60(1):98-100.
PMID: 23205499 DOI: 10.1111/jeu.12011

Molecular diversity of protists from bovine rumen fluid incubated with condensed tannins of Leucaena leucocephala hybrid-Rendang at 20 mg/500 mg dry matter (treatment) or without condensed tannins (control) was investigated using 18S rRNA gene library. Clones from the control library were distributed within nine genera, but clones from the condensed tannin treatment clone library were related to only six genera. Diversity estimators such as abundance-based coverage estimation and Chao1 showed significant differences between the two libraries, although no differences were found based on Shannon-Weaver index and Libshuff.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Fulltext Loop-Mediated Isothermal Amplification Assay for Identification of Five Human Plasmodium Species in Malaysia

Lau YL, Lai MY, Fong MY, Jelip J, Mahmud R

Am J Trop Med Hyg, 2016 Feb;94(2):336-339.
PMID: 26598573 DOI: 10.4269/ajtmh.15-0569

The lack of rapid, affordable, and accurate diagnostic tests represents the primary hurdle affecting malaria surveillance in resource- and expertise-limited areas. Loop-mediated isothermal amplification (LAMP) is a sensitive, rapid, and cheap diagnostic method. Five species-specific LAMP assays were developed based on 18S rRNA gene. Sensitivity and specificity of LAMP results were calculated as compared with microscopic examination and nested polymerase chain reaction. LAMP reactions were highly sensitive with the detection limit of one copy for Plasmodium vivax, Plasmodium falciparum, and Plasmodium malariae and 10 copies for Plasmodium knowlesi and Plasmodium ovale. LAMP positively detected all human malaria species in all positive samples (N = 134; sensitivity = 100%) within 35 minutes. All negative samples were not amplified by LAMP (N = 67; specificity = 100%). LAMP successfully detected two samples with very low parasitemia. LAMP may offer a rapid, simple, and reliable test for the diagnosis of malaria in areas where malaria is prevalent.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Fulltext Sarcocystis nesbitti infection in human skeletal muscle: possible transmission from snakes

Lau YL, Chang PY, Tan CT, Fong MY, Mahmud R, Wong KT

Am J Trop Med Hyg, 2014 Feb;90(2):361-4.
PMID: 24420776 DOI: 10.4269/ajtmh.12-0678

Sarcocystis nesbitti is an intracellular protozoan parasite found as sarcocysts within muscle fibers of intermediate hosts (monkey and baboon). The definitive host is suspected to be the snake. We report two cases from a larger cohort of 89 patients who had fever, headache, and generalized myalgia after a trip to Pangkor Island, Malaysia. Sarcocysts were detected in skeletal muscle biopsy specimens by light and electron microscopy from these two patients. DNA sequencing based on the 18S ribosomal DNA region identified the Sarcocystis species as S. nesbitti. We also identified S. nesbitti sequences in the stools of a snake (Naja naja). Phylogenetic analysis showed that these sequences form a cluster with most of the other known Sarcocystis species for which the snake is a definitive host. We believe these two patients were likely to have symptomatic acute muscular sarcocystosis after S. nesbitti infection that may have originated from snakes.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics
Fulltext From 18S to 28S rRNA Gene: An Improved Targeted Sarcocystidae PCR Amplification, Species Identification with Long DNA Sequences

Lee FCH, Muthu V

Am J Trop Med Hyg, 2021 02 22;104(4):1388-1393.
PMID: 33617472 DOI: 10.4269/ajtmh.20-0767

Sarcocystosis outbreaks in Tioman and Pangkor islands of Malaysia between 2011 and 2014 have raised the need to improve Sarcocystis species detection from environmental samples. In-house works found that published primers amplifying the 18S rRNA gene of Sarcocystis either could not produce the target from environmental samples or produced Sarcocystis DNA sequence that was insufficient for species identification. Using the primer pair of 18S S5 F (published) and 28S R6 R (new), this study improved the PCR amplification of Sarcocystidae to overcome these two difficulties. The PCR product spanned from the 18S to 28S rRNA genes, providing more information for species identification. The long DNA sequence allowed comparison between the "Ident" and "Query Cover" sorting in GenBank identity matching. This revealed the ambiguity in identity matching caused by different lengths of reference DNA sequences, which is seldom discussed in the literature. Using the disparity index test, a measurement of homogeneity in nucleotide substitution pattern, it is shown that the internal transcribed spacer (ITS)1-5.8S-ITS2 and 28S genes are better than the 18S gene in indicating nucleotide variations, implying better potentials for species identification. The example given by the handful of Sarcocystidae long DNA sequences reported herein calls for the need to report DNA sequence from the 18S to the 28S rRNA genes for species identification, especially among emerging pathogens. DNA sequence reporting should include the hypervariable 5.8S and ITS2 regions where applicable, and not be limited to single gene, per the current general trend.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics*
Gold-nanoparticle based electrochemical DNA sensor for the detection of fish pathogen Aphanomyces invadans

Kuan GC, Sheng LP, Rijiravanich P, Marimuthu K, Ravichandran M, Yin LS, et al.

Talanta, 2013 Dec 15;117:312-7.
PMID: 24209346 DOI: 10.1016/j.talanta.2013.09.016

Epizootic ulcerative syndrome (EUS) is a devastating fish disease caused by the fungus, Aphanomyces invadans. Rapid diagnosis of EUS is needed to control and treat this highly invasive disease. The current diagnostic methods for EUS are labor intensive. We have developed a highly sensitive and specific electrochemical genosensor towards the 18S rRNA and internal transcribed spacer regions of A. invadans. Multiple layers of latex were synthesized with the help of polyelectrolytes, and labeled with gold nanoparticles to enhance sensitivity. The gold-latex spheres were functionalized with specific DNA probes. We describe here the novel application of this improved platform for detection of PCR product from real sample of A. invadans using a premix sandwich hybridization assay. The premix assay was easier, more specific and gave higher sensitivity of one log unit when compared to the conventional method of step-by-step hybridization. The limit of detection was 0.5 fM (4.99 zmol) of linear target DNA and 1 fM (10 amol) of PCR product. The binding positions of the probes to the PCR amplicons were optimized for efficient hybridization. Probes that hybridized close to the 5' or 3' terminus of the PCR amplicons gave the highest signal due to minimal steric hindrance for hybridization. The genosensor is highly suitable as a surveillance and diagnostic tool for EUS in the aquaculture industry.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics*
Description of raabeia, synactinomyxon and neoactinomyxum developing stages of myxosporeans (Myxozoa) infecting Isochaetides michaelseni Lastočkin (Tubificidae) in Lake Balaton and Kis-Balaton Water Reservoir, Hungary

Borkhanuddin MH, Cech G, Molnár K, Németh S, Székely C

Syst Parasitol, 2014 Jul;88(3):245-59.
PMID: 24935127 DOI: 10.1007/s11230-014-9496-1

Molecular and morphometric investigations were conducted on the actinosporean morphotypes of myxosporeans surveyed in oligochaetes of Lake Balaton and Kis-Balaton Water reservoir. Oligochaetes belonging to the species Isochaetides michaelseni Lastočkin and Branchiura sowerbyi Beddard as well as to the genera Nais Dujardin, Dero Müller and Aeolosoma Ehrenberg were studied during an 18-month period. Actinosporeans were obtained exclusively from I. michaelseni (7,818 specimens) with very low prevalence (0.01-0.06%). Four new actinosporean morphotypes of the collective groups raabeia (2 types), synactinomyxon (1 type) and neoactinomyxum (1 type) were found and described, including the first synactinomyxon collective group from Hungarian biotopes and a new raabeia morphotype. Except for Synactinomyxon type 1, the 18S rDNA analysis revealed that the spores did not match any myxospore entity found in the GenBank.

Matched MeSH terms: RNA, Ribosomal, 18S/genetics

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