Comparison between the counter immunoelectrophoresis test and mouse neutralization test for the detection of antibodies against rabies virus in dog sera

The detection of rabies antibodies is extremely valuable for epidemiological studies, determination of immune status in man, animals, and for the diagnosis of the disease. Several serological procedures have been described for this purpose. The present study reports a comparison between counterimmunoelectrophoresis test (CIET) and mouse neutralization test (MNT) in the detection of antibodies against rabies virus from 212 serum samples of vaccinated dogs. The agreement between both techniques was 79.7% and a significative association was demonstrated. The correlation coefficients between MNT and the CIET titers was determined considering 88 samples showing positive results in both techniques [CIET = 2 and MNT = 5 (0.13 IU/ml)] and resulted r² = 0.7926 (p < 0.001). The performance of CIET system was technically simple, cheap and rapid, and thereby it could be useful for serological monitoring of dog vaccination campaigns as well as for individual analysis.

Rabies neutralizing antibody detection by indirect immunperoxidase serum neutralization assay performed on chicken embryo related cell line

The aim of this study was to evaluate the indirect immunoperoxidase virus neutralization (IPVN) and mouse neutralization test (MNT) to detect antibodies against rabies virus from vaccinated dogs and cattle. The IPVN was set up for the ability to measure 0.5 International Units/ml (IU) of antibody required by the World Health Organization and the Office International des Epizooties as the minimum response for proof of rabies immunization. IPVN was developed and standardized in chicken embryo related (CER) cell line when 141 dog and 110 cattle sera were applied by serial five-fold dilutions (1:5, 1:25, 1:125) as well as the positive and negative reference controls, all added in four adjacent wells, of 96-well microplates. A 50 µl amount of CVS32 strain dilution containing 50-200 TCID50/ml was mixed to each serum dilution, and after 90 min 50 µl of 3 x 10(5) cells/mlcell suspension added to each well. After five days of incubation, the monolayers were fixed and the IPVN test performed. The correlation coefficient between the MNT and IPVN performed in CER cells was r = 0.9949 for dog sera (n = 100) and r = 0.9307 for cattle sera (n = 99), as well as good specificity (94.7%), sensitivity (87.5%), and agreement (96.6%) were also obtained. IPVN technique can adequately identify vaccinated and unvaccinated animals, even from low-responding vaccinated animals, with the advantage of low cost and faster then MNT standard test.

Chicken embryo related (CER) cell line for quantification of rabies neutralizing antibody by fluorescent focus inhibition test

Levels of rabies virus neutralization antibody in sera from vaccinated dogs and cattle were either measured by mouse neutralization test (MNT) or by rapid fluorescent focus inhibition test (RFFIT), performed on CER monolayers. The two tests were compared for their ability to detect the 0.5 International Units/ml (I.U.) recommended by the World Health Organization (WHO) as the minimum response for proof of rabies immunization. A significant correlation was found between the two tests (n = 211; r = 0.9949 in dogs and 0.9307 in cows, p < 0.001), good sensitivity (87.5%), specificity (94.7%) and agreement (96.6%) as well. RFFIT method standardized on CER cell system for neutralizing antibodies detection turns the diagnosis easier and less expensive, specially when a great number of samples must be tested from endemic areas as commonly found in Brazil. (c) 2005 the International Association for Biologicals. Published by Elsevier Ltd. All rights reserved.

Comparison between RT-PCR and the mouse inoculation test for detection of rabies virus in samples kept for long periods under different conditions

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Estudo Comparativo de Três Diferentes Modalidades de Elisa para Medir os Anticorpos Contra o Vírus da Bronquite Infecciosa em Frangos Vacinados e Não Vacinados

No presente estudo, foram vacinados 150 frangos de corte de um dia de idade com sorotipo H120, e, após 28 dias desafiados à vacinação com o sorotipo M 41 do vírus da bronquite infecciosa das aves. da mesma forma, foram obtidos 150 soros de aves não vacinadas para a análise. Os respectivos soros foram analisados 28, 34 e 46 dias após o desafio, examinados através das técnicas de ELISA indireto (ELISA-I), Sandwich ELISA (S-ELISA) e ELISA com bloqueio de fase líquida (ELISA-BFL) e comparados com a técnica padrão de soroneutralização (SN) para efeito de cálculo da especificidade e sensibilidade relativas, bem como os valores predictivos positivos e negativos. O cálculo do coeficiente de correlação também foi empregado para a análise de concordância. Assim, os valores da correlação encontrados foram r = 0.98 entre ELISA-BFL x SNT, r = 0.79 entre S-ELISA x SNT e r = 0.74 ELISA-I x SNT. No entanto, quando comparamos as técnicas de ELISA entre si, ELISA-BFL x S-ELISA, ELISA-BFL x ELISA-I e S-ELISA x ELISA-I os valores encontrados foram r = 0.75, r = 0.69 e r = 0.79. A técnica de ELISA-BFL demonstrou melhor sensibilidade relativa que as técnicas de S-ELISA e ELISA-I, mesmo 46 dias após o desafio com a estirpe heteróloga. Entretanto, apesar das técnicas de S-ELISA e ELISA-I apresentarem especificidade realtiva superiores, a melhor correlação observada foi entre as técnicas de ELISA-BFL e a SN.

Interval between first dose and booster affected antibody production in cattle vaccinated against rabies

In this study, we compared the levels of neutralizing antibodies induced by inactivated rabies vaccine in cattle by using three alternative immunization procedures. Forty-five bovines (breed nelore) were then organized in three groups (A, B and C, with 15 animals/group). Group A received only one vaccine dose at day zero and Group B received the first dose at day zero and then another dose at day 30 (early booster). Group C was also immunized with two doses; however, the booster was postponed until day 180 after the first dose (delayed booster). Blood samples were withdrawn at days zero (before the first dose) and 30, 210, 390, and 540 after the beginning of immunization and the antibody titers were evaluated by mouse neutralization test. The protocol used to immunize Group C (booster at day 180) was clearly more efficient. In this group, antibody levels were higher and also remained higher for longer periods in comparison with the other two groups. These results show that booster timing significantly affected antibody levels. Therefore, programs addressed to control this disease in cattle should consider not only the use of a booster but also its administration time.

Physical environment modulates the behavioral responses induced by chemical stimulation of dorsal periaqueductal gray in mice

In order to investigate the relationship between behaviors elicited by chemical stimulation of the dorsal periaqueductal gray (dorsal PAG) and spontaneous defensive behaviors to a predator, the excitatory amino acid D,L-homocysteic acid (5 nmol in 0.1 mu l), was infused into the dorsal PAG and behavioral responses of mice were evaluated in two different situations, a rectangular novel chamber or the Mouse Defense Test Battery (MDTB) apparatus. During a 1-min period following drug infusion, more jumps were made in the chamber than in the MDTB runway but running time and distance traveled were significantly higher in the runway. Animals were subsequently tested using the standard MDTB procedure (anti-predator avoidance, chase and defensive threat/attack). No drug effects on these measures were significant. In a further test in the MDTB apparatus, the pathway of the mouse during peak locomotion response was blocked 3 times by the predator stimulus (anesthetized rat) to determine if the mouse would avoid contact. Ninety percent of D,L-homocysteic treated animals made direct contact with the stimulus (rat), indicating that D,L-homocysteic-induced running is not guided by relevant (here, threat) stimuli. These results indicate that running as opposed to jumping is the primary response in mice injected with D,L-homocysteic into the dorsal PAG when the environment enables flight. However, the lack of responsivity to the predator during peak locomotion suggests that D,L-homocysteic-stimulation into the dorsal PAG does not induce normal antipredator flight. (c) 2006 Published by Elsevier B.V.

Botulism by Clostridium botulinum type C in goats associated with osteophagia

An outbreak of botulism was detected in goats in the semiarid region of Brazil. In a flock of 460 goats, 38 does were affected and 37 died. Kids and younger goats were not affected. The main clinical signs were flaccid tetraparesis leading to tetraplegia that was often accompanied by twisted neck, tongue paralysis, and muscle tremors. At the time of the visit, 4 out of 11 affected goats were recumbent. Ambulatory goats had uncoordinated and swaying gaits with hypometria and weakness, mainly of the hind limbs. Two recumbent and four ambulatory goats showed twisted neck. Two recumbent goats were euthanized and necropsied. Non-significant gross and histologic lesions were observed. Samples of the liver, gut and rumen content were collected from the two goats and examined for botulinum toxins using the mouse serum neutralization test. The three samples from one goat were positive for type C toxin. Marked osteophagia was observed when the goats had access to bones in the pasture, and the farmer mentioned that osteophagia was common among goats of the flock. A sample of the plant Hybantus ipecaconha, the most abundant forage available for the goats, contained 2800 mg/kg of Ca and 450 mg/kg of P. One soil sample contained 58.12 mg/kg of Ca and 2.02 mg/kg of P. It was concluded that in this outbreak, botulism was associated with osteophagia probably due to phosphorus deficiency. (C) 2012 Elsevier B.V. All rights reserved.

Study of the presence of the spores of Clostridium botulinum in honey in Brazil

The isolation of Clostridium botulinum from honey samples is described. Botulism is characterized as an intoxication provoked by ingestion of contaminated foods with this toxin. Infant botulism happens by the ingestion of spores of C. botulinum together with food that in special conditions of the intestinal tract, such as those present in babies of less than 1 year old, will allow the germination and colonization of the intestine with production and absorption of botulinic toxin. The samples were subjected to dilution and to a thermal shock and cultivated in modified CMM (Difco). Cultures were subjected to Gram smears and toxicity tests in mice. The toxic cultures were purified in RFCA (Oxoid) plates and incubated in anaerobic jars. Positive samples were typed using the mouse assay neutralization test. From the 85 honey samples analyzed, six were positive for C. botulinum (7.06%), and identified as producers of type A, B, and D toxins.

Rabies surveillance in bats in Northwestern State of Sao Paulo

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Detection of antibodies against bovine respiratory syncytial virus (BRSV) in dairy cattle with different prevalences of bovine herpesvirus type 1 (BoHV-1) in São Paulo State, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Serological survey for rabies in serum samples from vampire bats (Desmodus rotundus) in Botucatu region, SP, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Distribution of rabies virus in infected mice, vaccinated and submitted to P-acnes as immunomodulator

The lethality and distribution of rabies virus were evaluated in swiss mice experimentally infected with street rabies virus, vaccinated and submitted to immunomodulation by P. acnes (formerly Corynebacterium parvum). The animals were sacrificed at different times,when the different tissues were collected and submitted to fluorescent antibody test (FAT) and mouse inoculation test (MIT). The group submitted to vaccination and P. acnes treatment presented a percentage of survival superior to that observed in infected mice only treated with P. acnes. Control infected animals had the lowest survival rates. The distribution of rabies virus in spleen of infected mice, vaccinated and submitted to P. acnes was superior to that verified in infected mice not treated with P. acnes. The increased survival correlated with the distribution of rabies virus in lymphoid tissues, could be interpreted as the consequence of P. acnes activity on macrophages. The results suggest the role of macrophages against rabies virus infection in mice and the importance of vaccination in the post expositive treatment of rabies. (C) 2002 Published by Elsevier B.V. Ltd.

Soroprevalência da arterite viral equina em mesorregiões paulistas entre 2007 e 2008

This study investigated the occurrence of equine arteritis virus antibodies using neutralization test in 1,400 horses from Campinas and macro metropolitan paulista mesoregions located at São Paulo State, between 2007 and 2008. Eighty (5.7%) samples showed antibodies to virus (titers from 4 to 4,096). Among the 42 cities, 15 (35.7%) presented at least one seropositive animal to equine arteritis virus. of the 238 farms analyzed 41 showed at least one seropositive animal. The occurrence was higher in sport horses like jumping horses, Quarter horses and was similar for female and male. The rate of seropositive was higher in animals older than 24 months of age. These results suggest the virus circulation among horse population in studied farms, and the need for continuing epidemiological surveillance studies.

Avaliação da resposta imune humoral em cães jovens imunizados contra a cinomose com vacina de vírus atenuado

Este estudo foi realizado com o objetivo de se investigar a cinética da resposta imune em cães jovens vacinados contra cinomose, com vírus atenuado, por meio do teste de soroneutralização. Onze cães sadios da raça Beagle, observados do nascimento até 30 meses de idade, isolados de outros cães, receberam vacina monovalente viva atenuada de cinomose canina aos 75, 105 e 135 dias de idade e doze meses após a terceira dose vacinal. O desenvolvimento de anticorpos foi mensurado pela reação de soroneutralização imediatamente antes da vacinação, trinta dias após a administração da primeira e terceira doses, noventa dias após a administração da terceira dose, nove e doze meses após a administração da terceira dose, trinta dias, seis e doze meses após a dose anual. Não havia títulos detectáveis de anticorpos contra cinomose canina antes da primovacinação. O título de anticorpos variou de 4,047 a 4,880 (em logaritmos na base 10), trinta dias após a administração da primeira dose e 6 meses após a dose anual, respectivamente. Todos os cães apresentaram uma resposta similar, embora com variações, com a produção de anticorpos com títulos maiores do que 2, considerado o título mínimo protetor. Doze meses após a dose anual, a maioria dos cães apresentou alto título de anticorpos, sugerindo que a revacinação anual poderia ser desnecessária.