Blood Polymerase Chain Reaction Research Articles

Detection of Mycobacterium avium subsp. paratuberculosis by several diagnostics techniques in clinical suspected sheep

A total of thirty sheep with clinical symptomatology of paratuberculosis (Johne’s disease) were subjected to four diagnostic techniques: histopathological examination, bacteriological culture (in faeces and tissues), polymerase chain reaction (PCR) (in blood, tissue and faecal samples) and antibody responses (ELISA). Twenty-one (70.0%) animals showed histological lesions. Bacterial culture of both faeces and tissue revealed that 2 (6.7%) and 6 (20.0%) of the 30 sheep were infected, respectively. Mycobacterium avium subsp. 24 paratuberculosis (Map) was identified in 4 animals via PCR of faeces (13.3%), and in 19 (63.3%) by PCR in tissues. PCR in blood revealed 7 (23.3%) infected animals. Three (10.0%) animals showed antibodies against Map. A greater number of positive animals were detected by histopathological examination and PCR in tissues than by culture or ELISA. This study confirmed the clinical findings and results suggest that histopathology, PCR in tissues and in blood can improve the detection of Map in physically suspected animals and should be considered useful tools in the diagnosis of Map in suspected sheep.

Detection of Ehrlichia, Anaplasma, and Babesia spp. in dogs of Cebu, Philippines.

Background:Ehrlichia, Anaplasma, and Babesia spp. are canine pathogens transmitted by the Rhipicephalus sanguineus tick which can cause varied clinical signs. These pathogens have been investigated in the Philippines, but coinfection has not been reported yet.Aim:The aim of this study was to evaluate the presence of Ehrlichia/Anaplasma and Babesia spp. in Philippine dogs.Materials and Methods:A total of 100 dogs from seven different veterinary establishments in Cebu, Philippines, were examined for Ehrlichia/Anaplasma and Babesia spp. infection using peripheral blood smear examination and polymerase chain reaction (PCR). Inclusion criteria included a history or presence of tick infestation, anemia, and/or thrombocytopenia. Clinical signs were recorded. Statistical analyses were performed between PCR positivity and clinical signs and hematological results.Results:A total of 10 and 18 dogs were found to be positive for Ehrlichia/Anaplasma and Babesia spp., respectively. One animal was PCR positive for both pathogens, which is the first report of coinfection in the country. The most common clinical signs observed include inappetence (89%), lethargy (80%), thrombocytopenia (85%), and anemia (74%). Analyses revealed that inappetence (p=0.044) and weight loss (p=0.028) were found statistically significant with Ehrlichia/Anaplasma infection. Basophil (p=0.001) and eosinophil counts (p=0.000) were also found significantly different between Ehrlichia/Anaplasma spp.-positive and -negative dogs. On the other hand, differential monocyte count (p=0.009) was found significantly different between Babesia spp.-positive and -negative dogs.Conclusion:The present study showed low infection rates of canine ehrlichiosis/anaplasmosis and babesiosis and provided additional evidence for the presence of the pathogens in the area.

Examination of methylation changes of VIM, CXCR4, DOK7, and SPDEF genes in peripheral blood DNA in breast cancer patients.

Studying whole blood DNA methylation as a risk marker has valuable applications in either diagnosis or staging of breast cancer. We investigated whole blood DNA methylation status of VIM, CXCR4, DOK7, and SPDEF genes in breast cancer patients in comparison to healthy control subjects. 60 patients with breast cancer and 40 healthy controls were examined. Genomic DNA isolated from peripheral blood and restriction enzyme polymerase chain reaction (REP) method was applied for analysis. Real-time PCR was used to confirm methylation status of the aforementioned genes and therefore to find out the methylation differences between normal and breast cancer subjects. Level of DOK7 promoter hypomethylation in normal and breast cancer samples was significant (P-value = 0.001). The study, also, showed that hypomethylation of VIM and CXCR4 genes are significant in patients compared with normal cases (P-value < 0.05). Furthermore, SPDEF promoter hypomethylation was not significantly differed between normal and breast cancer samples (P-value = 0.2). Hypermethylation of DOK7 gene in DNA from patients affected with breast cancer offers a biomarker for diagnosis of the breast cancer. This study indicates that methylation status of VIM and CXCR4 genes changes in breast cancer; so, they can be used as molecular biomarkers in breast cancer prognosis.

A survey of biochemical and acute phase proteins changes in sheep experimentally infected with Anaplasma ovis

Objective: To determine the biochemical and acute phase proteins changes in sheep experimentally infected with Anaplasma ovis (A. ovis). Methods: One Iranian sheep naturally infected with A. ovis (parasitemia 0.02%) but with no other blood parasites based on blood smear and polymerase chain reaction methods was selected as donor, and it was splenectomized to induce high level of parasitemia. Then, three weeks after splenectomy when parasitemia was 6%, donor’s blood was intravenously administered to each recipient animal. Five 5-6 months old Iranian male sheep without any blood parasites were selected as recipient animals. The percent of parasites, packed cell volume, serum biochemical parameters (urea, creatinine, bilirubin, aspartate aminotransferase activity, cholesterol, total protein, albumin, globulin, Fe), acute phase proteins (haptoglobin, total iron binding capacity, fibrinogen), were evaluated in sheep before and after being experimentally infected with A. ovis (until day 38). In addition, body weights of sheep were measured on days 0, 20 and 38. Results: In recipient sheep, microscopic examination of erythrocytes revealed a significant rise of parasitemia on days 12 and 15. The lowest level of packed cell volume in sheep was seen on day 15 post infection. A significant rise existed in mean urea and bilirubin (total, direct and indirect) on days 15 and 20. The increase of indirect bilirubin level was higher than direct bilirubin. Furthermore, serum Fe significantly increased on days 20 and 23. The mean total protein concentration significantly increased on day 38. A significant increase was found in the serum globulin concentration from days 20 and 27 to 38. Maximum values of haptoglobin were observed on days 27 and 30. Moreover, aspartate aminotransferase activity (from days 20-30) and cholesterol concentration (on day 20) significantly decreased. However, no significant changes were found in other parameters. Conclusions: Experimental ovine anaplasmosis caused by A. ovis could be associated with some changes in measured parameters, which presumably could be helpful for evaluation on staging of disease.

Prevalence of carrier state theileriosis in lactating cows.

Aim:The objective of this study was to examine the carrier status of theileriosis among apparently healthy cross-bred jersey cattle population of Odisha using conventional blood smear examination and polymerase chain reaction (PCR).Materials and Methods:A total of 34 apparently healthy cross-bred Jersey lactating cows were considered in this study. Blood samples were subjected to microscopic examination after staining with Giemsa stain and PCR based molecular diagnosis using two sets of primer, i.e., N516/N517 and TorF1/TorF2 specific for Theileria annulata and Theileria orientalis, respectively.Results:Examination of blood samples revealed presence of theileria parasites to a magnitude of 20.59% for T. annulata, 8.82% for T. orientalis, and 2.94% for both.Conclusion:Molecular diagnosis was found to be much more sensitive than conventional method for diagnosis of theileriosis. T. annulata was found to be the predominant species affecting the exotic cattle. T. orientalis was detected in apparently healthy cows.

Congenital Transmission of Trypanosoma cruzi in Argentina, Honduras, and Mexico: An Observational Prospective Study.

Compared with South America, there is a lack of epidemiologic studies about the risk of congenital transmission of Trypanosoma cruzi in Central America and Mexico. It has been suggested that T. cruzi genotypes might differ by region and that congenital transmission might vary according to the parasite's genotype. Our objective was to compare T. cruzi congenital transmission rates in three countries. We performed an observational prospective study in 2011-2014 enrolling women at delivery in one hospital in Argentina, two hospitals in Honduras, and two hospitals in Mexico. Congenital T. cruzi infection was defined as the presence of one or more of the following criteria: presence of parasites in cord blood (direct parasitological microscopic examination) with positive polymerase chain reaction (PCR) in cord blood, presence of parasites in infant's blood at 4-8 weeks (direct parasitological microscopic examination), and persistence of T. cruzi-specific antibodies at 10 months, as measured by at least two tests. Among 28,145 enrolled women, 347 had at least one antibody rapid test positive in cord blood and a positive enzyme-linked immunosorbent assay in maternal blood. PCR in maternal blood was positive in 73.2% of the cases, and genotyping identified a majority of non-TcI in the three countries. We found no (0.0%; 95% confidence interval [CI]: 0.0, 2.0) confirmed congenital case in Honduras. Congenital transmission was 6.6% (95% CI: 3.1, 12.2) in Argentina and 6.3% (95% CI: 0.8, 20.8) in Mexico. Trypanosoma cruzi non-TcI predominated and risks of congenital transmission were similar in Argentina and Mexico.

Infección natural por Cytauxzoon felis en onza parda (Puma concolor) de vida libre proveniente de la región suroeste de Mato Grosso do Sul, Brasil

Several factors such as deforestation and expansion of agriculture and livestock has promoted the close contact between wild and domestic animals, leading to the sharing of pathogens, including the Cytauxzoon felis. The aim of this study is to report a case of cytauxzoonose in a free-living Puma concolor from the southwest region of Mato Grosso do Sul, Brazil. The diagnosis was obtained through microscopy of peripheral blood smear and polymerase chain reaction, that in this report, was characterized as accidental finding, since clinical manifestations associated to disease were not observed. The detection of Cytauxzoon felis highlights the need to establish the epidemiological importance of wild animals and the risk that they represent to domestic and captive wild populations in relation to tick-borne pathogens transmitted.

Two randomized, controlled studies to assess the efficacy and safety of lotilaner (Credelio\u2122) in preventing Dermacentor reticulatus transmission of Babesia canis to dogs

BackgroundDogs worldwide are at risk of Babesia spp. infections. Preventive efficacy of lotilaner tablets (Credelio™, Elanco) against Babesia canis was evaluated in two studies.MethodsSixteen dogs in Study 1 and 12 dogs in Study 2, all seronegative and polymerase chain reaction (PCR) negative for B. canis, were randomized to a sham-treated control group or a lotilaner (20–43 mg/kg) treatment group, administered on Day 0 (Study 1: n = 8/group; Study 2: n = 6/group). Dogs were each infested with 50 Dermacentor reticulatus, a percentage of which (Study 1: 8.0–30.0%; Study 2: 12.2%) were infected with B. canis, in Study 1 on Days 2, 7, 14, 21 and 28, and in Study 2 on Day 28. Ticks were removed and counted on Day 30 in Study 1, and Day 34 in Study 2. Blood was collected for Babesia detection via smear, PCR and immunofluorescence assay (IFA) in Study 1 on Day 2, then approximately weekly through Day 56, and in Study 2 at weekly intervals between Days 28 to 49, and on Days 63 and 91. Additional samples were collected from dogs with body temperature > 39.4 °C (measured three times weekly, from Days 7 to 56 in Study 1 and from Days 35 to 56 in Study 2) and positive for B. canis on blood smear. Dogs with confirmed infections were rescue-treated, removed from the study and, in Study 1, replaced.ResultsAcross both studies B. canis infection of ticks ranged between 8.0–30.0%. In Study 1, all control dogs were positive for B. canis on blood smear and PCR on Day 10 and IFA on Day 21; on Day 21 seven of eight replacement control dogs were B. canis-positive; no replacement dogs were B. canis-positive following tick removal on Day 30. In Study 2, all control dogs were B. canis-positive on Day 56. All lotilaner-treated dogs remained B. canis-negative at all assessments in both studies.ConclusionLotilaner efficacy was 100% in preventing establishment of B. canis infection, despite post-treatment challenge with infected ticks on Days 2, 7, 14, 21 and 28.

The challenging diagnosis of hepatosplenic cat scratch disease in children

Cat scratch disease (CSD) can present as a systemic disease in 5–10% of cases leading to various entities including hepatosplenic disease with abscesses detected at abdominal imaging in more than 50% of patients. We describe a challenging diagnosis of hepatosplenic CSD, and review of literature. A previously healthy 12 y.o. girl was admitted for prolonged fever. She lived in contact with a cat. She had already received a 7-day unsuccessful antibiotic therapy (amoxicillin-clavulanate/ceftriaxone). Clinical/laboratory examination showed hepatosplenomegaly and no enlarged lymph nodes, CRP = 44 mg/L; ESR = 110 mm/h. An abdominal ultrasound evidenced multiple intrahepatic/intrasplenic hypoechoic lesions compatible with abscesses. Tests for main causes of liver abscesses ruled out included antibodies for Bartonella Henselase (BH) (enzyme immunoassay, EIA), Francisella Tularensis, Echinococcus Granulosus, Brucella spp; Entamoeba Histolitica microscopic identification; Tubercolosis skin tests and interferon-gamma release assays. Cultures, bone marrow aspiration, and alphafetoprotein were also normal. Due to a cat-pet at home CSD was reconsidered. IgG/IgM anti-BH were retested by indirect fluorescence (IFA) and found positive (title > 1:64). BH blood polymerase chain reaction (PCR) was negative. Home cat was found colonized by BH. The combination of contact history, serological, clinical, and imaging criteria were considered compatible with hepatosplenic CSD (Margileth diagnostic criteria). She was prescribed a combination of Azithromycin/Doxycycline with progressive improvement of all parameters.

Borrelia miyamotoi Polymerase Chain Reaction Positivity on a Tick-Borne Disease Panel in an Endemic Region of Rhode Island

Background Borrelia miyamotoi is a newer cause of ixodid (hard-bodied) tick-borne illness. Because symptoms are generally nonspecific and pathognomonic clinical features are lacking, whole blood polymerase chain reaction (PCR) may be useful for confirming a diagnosis. A tick-borne disease panel (“TBD-3”) bundled with Borrelia species (including B. miyamotoi) PCR (“TBD-3M”) was offered for patients with suspicion of TBD during 2014–2015 at South County Hospital in Rhode Island, an endemic region for many tick-borne illnesses. Materials and Methods Imugen laboratory was queried for all TBD-3M panels and Borrelia PCR’s sent from South County Hospital patients between January 1, 2014, and August 31, 2015. One positive whole blood B. miyamotoi PCR sample (genus and species specific) was considered evidence for B. miyamotoi infection. Demographics, clinical presentations, coinfections, laboratory abnormalities, and treatment courses for B. miyamotoi disease were examined. Results A total of 2324 patients had a bundled tick-borne panel performed for unexplained fever and other symptoms. In addition, 258 patients had individual Borrelia PCR’s performed. Overall, 78 patients were positive for B. burgdorferi PCR (3.0% positivity rate) and 18 patients were positive for B. miyamotoi PCR (0.7% positivity rate). Most patients presented with a “summer influenza”–like illness (94% reporting fever). Sixteen of these 18 patients were empirically prescribed doxycycline. One patient was concomitantly positive for B. miyamotoi PCR, B. microti PCR, and IgM Lyme antibodies. Three additional B. miyamotoi positive patients had positive Lyme serology. Conclusions Borrelia whole blood PCR was bundled to a pre-existing tick-borne panel to potentially account for unexplained fevers and other nonlocalizing symptoms in high-risk patients. Treatment recommendations for B. miyamotoi are based on case reports of successful treatments in other causes of relapsing fever, and the same antibiotics regimens are used for both. The utility of B. miyamotoi PCR in first-line screening of patients with unexplained fever may be minimal, given the low positivity rate observed and uniform effective empiric treatments prescribed.

Comparison of three methods for recovery of Brucella canis DNA from canine blood samples

Brucella canis, a gram-negative, facultative intracellular and zoonotic bacterium causes canine brucellosis. Direct methods are the most appropriate for the detection of canine brucellosis and bacterial isolation from blood samples has been employed as gold-standard method. However, due to the delay in obtaining results and the biological risk of the bacterial culturing, the polymerase chain reaction (PCR) has been successfully used as an alternative method for the diagnosis of the infection. Sample preparation is a key step for successful PCR and protocols that provide high DNA yield and purity are recommended to ensure high diagnostic sensitivity. The objective of this study was to evaluate the performance of PCR for the diagnosis of B. canis infection in 36 dogs by testing DNA of whole blood obtained through different extraction and purification protocols. Methods 1 and 2 were based on a commercial kit, using protocols recommended for DNA purification of whole blood and tissue samples, respectively. Method 3 was an in-house method based on enzymatic lysis and purification using organic solvents. The results of the PCR on samples obtained through three different DNA extraction protocols were compared to the blood culture. Of the 36 dogs, 13 (36.1%) were positive by blood culturing, while nine (25.0%), 14 (38.8%), and 15 (41.6%) were positive by PCR after DNA extraction using methods 1, 2 and 3, respectively. PCR performed on DNA purified by Method 2 was as efficient as blood culturing and PCR performed on DNA purified with in-house method, but had the advantage of being less laborious and, therefore, a suitable alternative for the direct B. canis detection in dogs.

EAN consensus review on prevention, diagnosis and management of tick-borne encephalitis.

Tick-borne encephalitis (TBE) is an infection of the central nervous system (CNS) caused by tick-borne encephalitis virus (TBEV) and transmitted by ticks, with a variety of clinical manifestations. The incidence of TBE in Europe is increasing due to an extended season of the infection and the enlargement of endemic areas. Our objectives are to provide recommendations on the prevention, diagnosis and management of TBE, based on evidence or consensus decisions. For systematic evaluation, the literature was searched from 1970 to 2015 (including early online publications of 2016), and recommendations were based on evidence or consensus decisions of the Task Force when evidence-based data were not available. Vaccination against TBE is recommended for all age groups above 1 year in highly endemic areas (≥5 cases/100 000/year), but also for individuals at risk in areas with a lower incidence. Travellers to endemic areas should be vaccinated if their visits will include extensive outdoor activities. Post-exposure prophylaxis after a tick bite is not recommended. A case of TBE is defined by the presence of clinical signs of meningitis, meningoencephalitis or meningoencephalomyelitis with cerebrospinal fluid (CSF) pleocytosis (>5 × 106 cells/l) and the presence of specific TBEV serum immunoglobulin M (IgM) and IgG antibodies, CSF IgM antibodies or TBEV IgG seroconversion. TBEV-specific polymerase chain reaction in blood is diagnostic in the first viremic phase but it is not sensitive in the second phase of TBE with clinical manifestations of CNS inflammation. Lumbar puncture should be performed in all patients with suspected CNS infection unless there are contraindications. Imaging of the brain and spinal cord has a low sensitivity and a low specificity, but it is useful for differential diagnosis. No effective antiviral or immunomodulating therapy is available for TBE; therefore the treatment is symptomatic. Patients with a potentially life threatening meningoencephalitis or meningoencephalomyelitis should be admitted to an intensive care unit. In the case of brain oedema, analgosedation should be deepened; osmotherapy and corticosteroids are not routinely recommended. If intracranial pressure is increased, therapeutic hypothermia or decompressive craniectomy might be considered. Seizures should be treated as any other symptomatic epileptic seizures. Tick-borne encephalitis is a viral CNS infection that may result in long-term neurological sequelae. Since its incidence in Europe is increasing due to broadening of endemic areas and prolongation of the tick activity season, the health burden of TBE is enlarging. There is no effective antiviral treatment for TBE, but the disease may be effectively prevented by vaccination.

Combined parasitological and molecular-based diagnostic tools improve the detection of Trypanosoma cruzi in single peripheral blood samples from patients with Chagas disease.

In order to detect Trypanosoma cruzi and determine the genetic profiles of the parasite during the chronic phase of Chagas disease (ChD), parasitological and molecular diagnostic methods were used to assess the blood of 91 patients without specific prior treatment. Blood samples were collected from 68 patients with cardiac ChD and 23 patients with an indeterminate form of ChD, followed by evaluation using blood culture and polymerase chain reaction. T . cruzi isolates were genotyped using three different genetic markers. Blood culture was positive in 54.9% of all patients, among which 60.3% had the cardiac form of ChD, and 39.1% the indeterminate form of ChD. There were no significant differences in blood culture positivity among patients with cardiac and indeterminate forms. Additionally, patient age and clinical forms did not influence blood culture results. Polymerase chain reaction (PCR) was positive in 98.9% of patients, although comparisons between blood culture and PCR results showed that the two techniques did not agree. Forty-two T . cruzi stocks were isolated, and TcII was detected in 95.2% of isolates. Additionally, one isolate corresponded to TcIII or TcIV, and another corresponded to TcV or TcVI. Blood culture and PCR were both effective for identifying T. cruzi using a single blood sample, and their association did not improve parasite detection. However, we were not able to establish an association between the clinical form of ChD and the genetic profile of the parasite.

Prevalence and distribution of malaria, Pfcrt and Pfmdr 1 genes in patients attending FUT Health Centre, Akure, Nigeria

This research evaluated the distribution of malaria, Plasmodium falciparum chloroquine resistance transporter (Pfcrt) and Plasmodium falciparum multidrug resistant (Pfmdr1) mutant genes among patients attending FUTA Health Centre. The research was carried out between December 2015 and February 2016. One hundred and forty (140) patients coming for malaria test were examined out of which 74 (52.86%) were males and 66 (47.14%) were females. Malaria test was done by thick blood film microscopy and Polymerase Chain Reaction (PCR) technique. PCR technique was used to detect Pfcrt and Pfmdr1 mutant genes of P. falciparum. Information about sex, age, status and date of collection of each patient were collected. Pearson’s Chi-square test was used for analysis. Out of 140 blood samples that were tested, 100 (71%) were positive for malaria infection. The percentage distribution of malaria infection among male and female patients was 54 (72.97%) and 46 (69.70%) respectively with no significant difference (P=0.668). The prevalence of Pfcrt (95%) and Pfmdr1 (45%) showed significant difference (P=0.001). The research revealed high prevalence of Pfcrt and Pfmdr1 mutant genes of P. falciparum among patients which may be as a result of treatment of malaria with Chloroquine and other medication which the parasite have formed resistance against. It is therefore important to administer other malaria drugs apart from the drugs the parasite have formed resistance against.

Preliminary data on microRNA expression profiles in a group of South African patients diagnosed with chronic myeloid leukaemia.

Micro-ribonucleic acids (miRNAs) are small functional non-coding RNAs that downregulate gene expression at the post-transcriptional level. Abnormal expression of specific miRNAs has been recorded in chronic lymphocytic leukaemia, other non-Hodgkin B-cell lymphomas, lung cancer and chronic myeloid leukaemia (CML). The aim of this study was to compare miRNA expression profiles among patients with newly diagnosed CML, those on established therapy with imatinib mesylate, and healthy individuals. The expression of 88 miRNAs was evaluated in a total of nine samples divided into three groups: Group 1 comprised three samples collected from newly diagnosed CML patients; group 2 consisted of three samples collected from patients on therapy; the remaining three samples were collected from healthy volunteers (control group). Total RNA was extracted from whole blood and reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) was performed on the LightCycler® 480 platform using Human Serum & Plasma miRNA PCR Arrays. In group 1, only SNORD44 was downregulated, while hsa-miR-372 and hsa-miR-375 were found to be significantly upregulated compared with the control group. By contrast, 49 miRNAs were significantly upregulated in group 2 compared with the control group. miRNAs hsa-miR-106b, hsa-miR-21, hsa-miR-221, hsa-miR-10a, hsa-miR-193a-5p and hsa-miR-30e were expressed in group 2. Therefore, miRNA expression profiles differed between the two patient groups.

Primary Pyomyositis as Unusual Cause of Limp: Three Cases in Immunocompetent Children and Literature Review

AbstractPyomyositis (PM) is an uncommon primary skeletal muscle infection caused mainly by Staphylococcus aureus that is characterized by single or multiple intramuscular abscess formation. In our ward, between 2013 and 2015, three children (two females and one male) aged from 2 to 12 years were diagnosed and treated for PM. Patients' medical records and imaging studies were examined retrospectively. All patients, otherwise healthy, complained of limp, fever, and severe lower limb pain. Skin scratch lesions were detected in two cases; one of them showed an edematous appearance of the affected area. Multifocal bilateral abscesses of gemini and gastrocnemius were detected in the youngest patient; right obturator and iliac muscles were affected in the second patient; and right gluteus and pyriform muscles were involved in the third patient. All patients showed elevated acute phase reactants and had normal serum creatinine kinase levels. Blood cultures and polymerase chain reaction (PCR) investigations were negative in all cases. Magnetic resonance imaging (MRI) findings included muscle enlargement, deep fascia, high signal in subcutaneous tissues, and postgadolinium abscess formation. No patient required surgical or percutaneous drainage. All three were treated conservatively with intravenous oxacillin, associated with ceftriaxone in the first patient and ceftazidime in the other two, followed by oral antibiotic therapy for a period ranging from 5 to 6 weeks. Pyomyositis must be considered as an unusual cause of limp in children of all ages. When promptly diagnosed and adequately treated, it has a favorable outcome without relapses or sequelae as occurred in all our patients.

Congenital malaria: Is it really rare? A case report

A 7-day-old term male infant weighing 2.4 kg was admitted with fever, pallor, icterus, and splenomegaly for 3 days. The primi mother was treated for pyrexia in the last trimester. Investigation revealed anemia, hyperbilirubinemia, and thrombocytopenia and demonstrated Plasmodium vivax in peripheral blood smear and card test. C-reactive protein was raised and blood culture was sterile. The baby was responded well to intravenous (IV) artesunate. Recent studies suggest that congenital malaria (CM) is not as rare as previously thought. Cord blood shows greater parasitemia as compared to neonatal blood. Besides light microscopy, plasmodium antigen detection and polymerase chain reaction of blood may help in diagnosis. CM can be confused with toxoplasmosis, rubella, cytomegalovirus, herpes simplex syndrome, and neonatal sepsis.

Novel incidental finding of Hepatozoon canis infection in two dogs of the same household in Trinidad, West Indies.

A five-year old mixed breed bitch was presented to the veterinary clinic in lateral recumbency with a history of anorexia and muscle hyperaesthesia. Examination of the blood smear of this animal revealed the presence of Hepatozoon spp. gamonts in the neutrophils and monocytes with a parasitaemia level of approximately 2%. Complete blood count (CBC) revealed a neutrophilia, and a normocytic normochromic non-regenerative anaemia which were consistent with Hepatozoon spp. infections. Diagnosis was confirmed by polymerase chain reaction (PCR) amplification of the 18S rRNA gene followed by DNA sequencing of the amplicon. Although the other dog in the household appeared asymptomatic, Hepatozoon canis infection was confirmed by both microscopic examination of blood smear and PCR. Both dogs were infested with Rhipicephalus sanguineus ticks. Phylogenetic analysis revealed that the H. canis sequences from these two dogs were similar to those from Venezuela and St Kitts but not Brazil. This is the first reported case of Hepatozoon canis infections in dogs in Trinidad that were confirmed by molecular techniques.

Laboratory Diagnosis of Breakthrough Varicella in Children

Breakthrough varicella (BV) develops in vaccinated persons as a result of infection by wild-type varicella-zoster virus more than 42 days after varicella vaccination. The clinical symptoms are atypical, and clinical diagnosis can be difficult. We investigated laboratory-based diagnostic methods that are relatively simple and highly precise to conduct accurate surveillance. We enrolled 42 patients with suspected BV at 2 pediatric hospitals and performed a real-time polymerase chain reaction (PCR) on the skin lesions to confirm the BV diagnosis. We performed PCR on saliva and blood collected during the acute phase, as well as direct fluorescent antibody (DFA) imaging on lesions, and measured varicella-zoster virus immunoglobulin (Ig) G and IgM during the acute and convalescent phases. We confirmed the BV diagnosis in 31 of 42 enrolled patients. The sensitivity of DFA imaging of the lesion, and PCR of saliva and blood were 93.5%, 87.1% and 61.3%, respectively. IgM was detected in 12.9% of patients during the acute phase and in 65.5% during the convalescent phase. IgG increased more than 4-fold in 86.2% of patients between the acute and convalescent phases. The sensitivity and specificity of the assay were 83.9% and 81.8%, respectively, when the diagnostic criteria for IgG were set to greater than 20 during the acute phase. The gold standard of laboratory-based diagnosis of BV has been the PCR of samples taken from lesions. However, DFA of the lesion showed equivalent sensitivity when compared with PCR. PCR using saliva samples is an effective, noninvasive method of diagnosis. We found that high values of IgG during the acute phase can aid in the diagnosis of BV.

Rapid diagnostic tests for typhoid and paratyphoid (enteric) fever.

Rapid diagnostic tests for typhoid and paratyphoid (enteric) fever.