Molecular typing ofGiardia duodenalisin humans in Queensland – first report of Assemblage E

Parasitology ◽  
2017 ◽  
Vol 144 (9) ◽  
pp. 1154-1161 ◽  
Author(s):  
ALIREZA ZAHEDI ◽  
DANIEL FIELD ◽  
UNA RYAN
Keyword(s):  
Molecular Typing ◽  
Quantitative Polymerase Chain Reaction ◽  
Giardia Duodenalis ◽  
Protozoan Parasite ◽  
Mixed Infections ◽  
Chain Reaction ◽  
Specific Primers ◽  
First Report ◽  
Polymerase Chain ◽  
Assemblage B

SUMMARYLittle is known about the genetic diversity of the protozoan parasite,Giardia duodenalis,infecting humans in Queensland, Australia. The present study typed 88 microscopicallyGiardia-positive isolates using assemblage-specific primers at the triose phosphate isomerase (tpi) gene and sequenced a subset of isolates at the glutamate dehydrogenase (gdh) gene (n= 30) andtpilocus (n= 27). Using thetpi-assemblage specific primers,G. duodenalisassemblage A and assemblage B were detected in 50% (44/88) and 38·6% (34/88) of samples, respectively. Mixed infections with assemblages A and B were identified in 4·5% (4/88) and assemblage E was identified in 6·8% (6/88) of samples. Sequence analysis at thegdhandtpiloci also confirmed the presence of assemblage E in these isolates. Cyst numbers per gram of feces (g−1) were determined using quantitative polymerase chain reaction and of the isolates that were typed as assemblage E, cyst numbers ranged 13·8–68·3 × 106cysts g−1. This is the first report of assemblage E in humans in Australia, indicating that in certain settings, this assemblage may be zoonotic.

Mixed Giardia duodenalis assemblage infections in children and adults in South India

2012 ◽  
Vol 140 (11) ◽  
pp. 2023-2027 ◽  
Author(s):  
S. LAISHRAM ◽  
A. KANNAN ◽  
P. RAJENDRAN ◽  
G. KANG ◽  
S. S. R. AJJAMPUR
Keyword(s):  
South India ◽  
Triosephosphate Isomerase ◽  
Fragment Length Polymorphism ◽  
Gastrointestinal Symptoms ◽  
Giardia Duodenalis ◽  
Chain Reaction ◽  
Pcr Rflp ◽  
Polymerase Chain ◽  
Restriction Fragment Length ◽  
Assemblage B

SUMMARYThe assemblages of Giardia duodenalis in 25 children with and 25 children without diarrhoea and 24 adults with gastrointestinal symptoms in South India were determined. Polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) targeting the glutamate dehydrogenase (gdh), β-giardin and triosephosphate isomerase (tpi) genes was used. The tpi PCR was the most sensitive and detected G. duodenalis in all 74 microscopy-positive samples, while gdh and β-giardin PCR were positive in 62·2% and 56·8% of the samples. Assemblage B was predominant in both children and adults (82·4%) followed by assemblage AII (9·4%); assemblage AI was not detected. Infections with both assemblages A and B (detected by tpi PCR–RFLP) were seen exclusively in children and the mixed assemblage BIII and BIV (detected by gdh PCR–RFLP) was more common in children than adults (P=0·058).

scholarly journals Incidence of Multiple and Distinct Species of Caulimoviruses in Dahlia (Dahlia variabilis)

HortScience ◽  
2009 ◽  
Vol 44 (5) ◽  
pp. 1498-1500 ◽  
Author(s):  
Sahar Eid ◽  
Keri L. Druffel ◽  
Dayle E. Saar ◽  
Hanu R. Pappu
Keyword(s):  
Mosaic Virus ◽  
Distinct Species ◽  
The United States ◽  
Mixed Infections ◽  
Symptom Expression ◽  
Chain Reaction ◽  
Specific Primers ◽  
Polymerase Chain ◽  
Serious Disease ◽  
Free Material

Dahlia mosaic is a serious disease affecting dahlias. In addition to the Dahlia mosaic virus (DMV) reported previously, we characterized two putative new caulimoviruses, tentatively designated as DMV-D10 and Dahlia common mosaic virus (DCMV), from dahlia. To better understand their relative incidence in dahlia, a total of 213 samples were collected during 2007 and 2008 from several varieties of cultivated dahlia (D. variabilis) in the United States. Samples were tested for the three caulimoviruses using virus-specific primers in a polymerase chain reaction. Amplicons were cloned and sequenced to confirm the infection of dahlia with these viruses. Results showed that DMV-D10 was the most prevalent (94%) followed by DCMV (48.5%) and DMV (23%). Mixed infections were common and viruses were detected irrespective of symptom expression at the time of sampling. Two percent of the samples were not infected by any of the three tested caulimoviruses. Results suggest that caulimovirus infections are widespread in dahlia and highlight the need for testing and production of virus-free material to reduce their spread.

scholarly journals First Report of a Lettuce-Infecting Sequivirus in Chile

Plant Disease ◽  
2005 ◽  
Vol 89 (10) ◽  
pp. 1129-1129 ◽  
Author(s):  
R. Krause-Sakate ◽  
A. S. Jadão ◽  
A. C. Firmino ◽  
M. A. Pavan ◽  
F. M. Zerbini ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Mosaic Virus ◽  
Plant Viruses ◽  
Yellow Mosaic Virus ◽  
Mixed Infections ◽  
Replicase Gene ◽  
Rt Pcr ◽  
Chain Reaction ◽  
First Report ◽  
Polymerase Chain

Sequiviruses are isometric aphidborne plant viruses. Dandelion yellow mosaic virus (DaYMV), genus Sequivirus, was isolated from dandelion and lettuce in Europe. Lettuce mottle virus (LeMoV), a putative sequivirus, is often found in mixed infections with Lettuce mosaic virus (LMV) in Brazil (3). DaYMV, LeMoV and LMV cause similar mosaics in field-grown lettuce. Differences in biology and sequence suggest that DaYMV and LeMoV are distinct species (2). Forty-two and 101 lettuce samples with mosaic symptoms collected from two locations near Santiago during a survey of lettuce viruses in Chile in 2002 and 2003, respectively, were analyzed for the presence of LeMoV using reverse transcription polymerase chain reaction (RT-PCR). Total RNA was extracted (1) and used for RT-PCR with the specific LeMoV primers pairs Lmo3 (5′ ACATGAGCACTAGTGAGG 3′) and Lmo4 (5′ AGATAGAGCCGTCT GGCG 3′) (2). One of the 42 and three of the 101 samples produced the expected 300-bp fragment. Isometric particles of 30 nm diameter, typical of a sequivirus, were visualized by transmission electron microscopy. These samples were tested using RT-PCR for the presence of LMV and Cucumber mosaic virus (CMV), but no mixed infections were observed. One isolate, Ch36, was reamplified with the degenerate primer pairs DALE 1 (5′ GARTTCAACATGCACGCCAG 3′) and DALE 2 (5′ TTTTTCTCCCCATYCGTCAT 3′) which amplify part of the putative replicase gene (2) and produced a 563-bp fragment that was cloned on pGEM-T Easy (Promega, Madison, WI) and sequenced. The Ch36 product (EMBL Accession No. AM039965) showed 97% amino acid identity with LeMoV from Brazil, 79% with DaYMV, 72% with the sequivirus Parsnip yellow fleck virus, and 34% with the waikavirus Maize chlorotic dwarf virus. To our knowledge, this is the first report of a sequivirus in field lettuce in Chile, and although the virus was found at low incidence, this report extends the range of LeMoV to the western side of the Cordillera de Los Andes. The impact of LeMoV needs to be further analyzed in Chile, Brazil, and possibly other South American countries. References: (1) Y. D. Bertheau et al. DNA amplification by polymerase chain reaction (PCR) 1998. In: Methods for the Detection and Quantification of Erwinia carotovora subsp. atroseptica on potatoes. M. C. N. Perombelon and J. M. van der Wolff, eds. Scott. Crop Res. Inst. Occasional Publ., Dundee, 1998. (2) A. S. Jadão. Caracterização parcial e desenvolvimento de oligonucleotídeos específicos para detecção de sequivirus infectando alface. Ph.D. thesis. FCA-UNESP-Botucatu, Brazil, 2004. (3) O. Stangarlin et al. Plant Dis. 84:490, 2000.

scholarly journals First Report of Tomato infectious chlorosis virus in Spain

Plant Disease ◽  
2002 ◽  
Vol 86 (6) ◽  
pp. 696-696 ◽  
Author(s):  
M. I. Font ◽  
P. Martínez-Culebras ◽  
M. C. Jorda ◽  
D. Louro ◽  
A. M. Vaira ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
The United States ◽  
Mixed Infections ◽  
Chain Reaction ◽  
First Report ◽  
Dna Fragment ◽  
Polymerase Chain ◽  
Plant Pathol ◽  
Tomato Chlorosis Virus ◽  
Tomato Infectious Chlorosis Virus

During the summer and autumn of 2001, symptoms of interveinal yellowing, bronzing, brittleness, and rolling of lower leaves were observed in greenhouse- and field-grown tomato (Lycopersicon esculentum) plants in Castellon Province in eastern Spain. Symptoms resembled those caused by the whitefly-transmitted criniviruses (1,2). Total RNA was extracted from 28 samples of symptomatic leaves collected in three greenhouses and one field and analyzed by reverse transcription-polymerase chain reaction using primers specific for Tomato chlorosis virus (ToCV) (1) and Tomato infectious chlorosis virus (TICV) (2). The 501-bp TICV-specific DNA fragment was amplified in four samples collected during the summer in three greenhouses and one field, and the 439-bp ToCV-specific DNA fragment was amplified in 15 samples collected during the autumn in the same three greenhouses; no mixed infections were found. The DNA fragments amplified from TICV were sequenced and showed 99 to 100% identity with the TICV isolates (GenBank Accession Nos. U67449 and AY048855) from the United States and Italy, respectively, confirming the diagnosis. One sequence was deposited as GenBank Accession No. AF479662. To our knowledge, this is the first report of TICV in Spain and the second in Europe. References: (1) D. Louro et al. Eur. J. Plant Pathol. 106:539, 2000. (2) A. M. Vaira et al. Phytoparasitica. In Press.

scholarly journals First Report of Cacopsylla picta as a Vector of Apple Proliferation Phytoplasma in Germany

Plant Disease ◽  
2003 ◽  
Vol 87 (1) ◽  
pp. 101-101 ◽  
Author(s):  
B. Jarausch ◽  
N. Schwind ◽  
W. Jarausch ◽  
G. Krczal ◽  
E. Dickler ◽  
...  
Keyword(s):  
Northern Italy ◽  
Insect Vector ◽  
Apple Proliferation ◽  
Chain Reaction ◽  
Specific Primers ◽  
Molecular Analyses ◽  
First Report ◽  
Micropropagated Plants ◽  
Polymerase Chain ◽  
Test Plants

Since 2000, a serious epidemic of apple proliferation (AP) reappeared in southwestern Germany. Molecular analyses revealed that the AP phytoplasma is associated with this disease. Since no curative treatments or resistant cultivars exist, the only means to reduce spread of the disease is the control of the insect vector. Recently, Frisinghelli et al. (1) identified Cacopsylla costalis as a vector of AP phytoplasma in northern Italy. Following this result, transmission trials with C. picta (synonym C. costalis) were conducted in southwestern Germany at Neustadt (Rheinland-Pfalz) and Dossenheim (Baden-Württemberg) since 2001. Overwintering psyllids were captured from March to May in different orchards. Groups of 5 to 30 C. picta were caged for 2 to 4 weeks on apple seedlings or healthy micropropagated plants. Leaf midribs of test plants were sampled 2 to 3 months after inoculation feeding and tested by polymerase chain reaction (PCR) for AP phytoplasma with specific primers AP5/AP4 (2). In 2001, 1 of 10 test plants, and in 2002, 7 of 40 test plants became AP infected. In 2002, one to four C. picta specimens fed on plants which became infected were tested AP phytoplasma positive by PCR while all psyllids recollected from PCR-negative plants were tested negative. Transmission of the AP phytoplasma was successful at both sites. To our knowledge, this is the first report of C. picta as a vector of the AP phytoplasma in Germany. References: (1) C. Frisinghelli et al. J. Phytopathol. 148:425, 2000. (2) W. Jarausch et al. Appl. Environ. Microbiol. 60:2916, 1994.

Prevalence and seasonality of Nosema species in Québec honey bees

2012 ◽  
Vol 144 (4) ◽  
pp. 577-588 ◽  
Author(s):  
T.R. Copley ◽  
H. Chen ◽  
P. Giovenazzo ◽  
E. Houle ◽  
S.H. Jabaji
Keyword(s):  
Polymerase Chain Reaction ◽  
Honey Bees ◽  
Quantitative Polymerase Chain Reaction ◽  
Conventional Polymerase Chain Reaction ◽  
Published Data ◽  
Seasonal Patterns ◽  
Mixed Infections ◽  
Chain Reaction ◽  
Nosema Species ◽  
Polymerase Chain

AbstractNosemosis is a disease of adult honey bees, Apis mellifera Linnaeus (Hymenoptera: Apidae), caused by two described species of Microsporidia: Nosema ceranae Fries and Nosema apis Zander. The epidemiology of N. apis is well understood; however, little is known about N. ceranae in Canadian apiaries. The following study aimed to determine the seasonal patterns of N. ceranae and N. apis in European honey bees in a Québec, Canada, apiary. Honey bees from six hives were sampled from 2008 to 2010 and the amount of spores quantified by both microscopic spore counts and duplex quantitative polymerase chain reaction (qPCR). Results demonstrated that duplex qPCR was the most sensitive technique and was able to detect N. ceranae in samples confirmed negative for microscopic spore counts and conventional polymerase chain reaction (PCR) detection. Results show that N. ceranae is the more prevalent parasite and was present in 75% of collections as single or mixed infections in the sampled apiary. The prevalence of N. apis was lower representing 29.7% throughout the 3 years of the study, and by 2010 was present only as mixed infections. Seasonal patterns of N. apis were consistent with previously published data with peaks in spring and autumn months, while N. ceranae peak infections varied throughout the 3-year study.

AN IMPROVED METHOD FOR INHIBITOR FREE NUCLEIC ACIDS FROM POLYPHENOL RICH LEAVES OF MUSA SPP

2017 ◽  
Vol 23 (1) ◽  
Author(s):  
N.NANDHA KUMAR ◽  
K. SOURIANATHA SUNDARAM ◽  
D. SUDHAKAR ◽  
K.K. KUMAR
Keyword(s):  
Polymerase Chain Reaction ◽  
Quantitative Polymerase Chain Reaction ◽  
Proteinase K ◽  
Chain Reaction ◽  
Banana Plant ◽  
High Molecular Weight Dna ◽  
Musa Spp ◽  
Leaf Tissues ◽  
Polymerase Chain

Excessive presence of polysaccharides, polyphenol and secondary metabolites in banana plant affects the quality of DNA and it leads to difficult in isolating good quality of DNA. An optimized modified CTAB protocol for the isolation of high quality and quantity of DNA obtained from banana leaf tissues has been developed. In this protocol a slight increased salt (NaCl) concentration (2.0M) was used in the extraction buffer. Polyvinylpyrrolidone (PVP) and Octanol were used for the removal of polyphenols and polymerase chain reaction (PCR) inhibitors. Proteins like various enzymes were degraded by Proteinase K and removed by centrifugation from plant extract during the isolation process resulting in pure genomic DNA, ready to use in downstream applications including PCR, quantitative polymerase chain reaction (qPCR), ligation, restriction and sequencing. This protocol yielded a high molecular weight DNA isolated from polyphenols rich leaves of Musa spp which was free from contamination and colour. The average yields of total DNA from leaf ranged from 917.4 to 1860.9 ng/ìL. This modified CTAB protocol reported here is less time consuming 4-5h, reproducible and can be used for a broad spectrum of plant species which have polyphenol and polysaccharide compounds.

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