Assessing the efficacy and zootechnical performance in laying hens after administration of a 1% aqueous solution of fluralaner (Exzolt) for treating natural infestation by Menacanthus cornutus.

A brood of laying hens infested with the lice Menacanthus cornutus (Phthiraptera: Menoponidae) evidenced itching, irritation, and damage to their zootechnical performance. A study was conducted to evaluate the zootechnical performance and infestation control using a 1% solution of fluralaner in a brood of white laying hens infested naturally with lice. The experiment was carried out using 10,560 naturally infested chickens divided into 2 groups: a treatment group of animals that received a 1% solution of fluralaner in drinking water, at a dose of 0.05 mL/kg of body weight, in 2 administrations, 7 d apart; and a control group of infested and untreated chickens. The groups of chickens were followed for 120 d to evaluate the score of infestation and zootechnical performance. It was observed that birds in the treatment group became free of lice infestation 7 d after the administration of the first dose of a 1% solution of fluralaner. For up to 120 d after the experiment was initiated, there was no evidence of subsequent lice infestation, while continued infestation with all life stages of lice (adults, young, or eggs) was evident in the untreated control group, remaining stable during all evaluations performed. The birds in the treatment group showed improved zootechnical performance when compared to a 9.94% egg production decrease in the control group. The feed conversion and egg mass data showed statistically significant differences between the 2 groups. This study allows us to conclude that treatment with a 1% solution of fluralaner effectively controlled Menacanthus cornutus lice infestation and promoted recovery of egg production in a brood of laying hens treated with the test formulation.

Seroepidemiology of Hepatitis Viruses and Hepatitis B Genotypes of Female Marriage Immigrants in Korea.

PURPOSE: The Korean society has moved rapidly toward becoming a multicultural society. This study aimed to estimate the seroprevalence of hepatitis viruses and investigate hepatitis B virus (HBV) genotypic diversity in female marriage immigrants. MATERIALS AND METHODS: Screening program was conducted at support centers for multicultural families in 21 administrative districts in Korea between July 2011 and January 2017. A total of 963 female marriage immigrants were included in this study. Blood samples were tested for hepatitis viral markers and HBV genotype. RESULTS: Subjects' median age was 33 years (20-40 years), and they originated from nine countries including Vietnam (n=422, 43.8%), China (n=311, 32.3%), the Philippines (n=85, 8.8%), Cambodia (n=58, 6.0%), and Japan (n=39, 4.0%). About 30% (n=288) of subjects required hepatitis A vaccination. HBsAg positive rate was 5.4% (n=52). Positive HBsAg results were the highest in subjects from Southeast Asia (6.6%, n=38). Anti-HBs positive rate was 60.4% (n=582). About 34% (n=329) of subjects who were negative for anti-HBs and HBsAg required HBV vaccinations. Genotypes B and C were found in 54.6% (n=12) and 45.4% (n=10) of the 22 subjects with HBV, in whom genotypes were tested. Eight (0.8%) subjects were positive for anti-HCV. Positive anti-HCV results were the highest in subjects from Central Asia (7.9%, n=3). CONCLUSION: Testing for hepatitis viral marker (hepatitis A virus IgG and HBsAg/anti-HBs) is needed for female marriage immigrants. Especially, HBV genotype B is different from genotype C of Koreans. Therefore, interest and attention to vaccination programs for female marriage immigrants are necessary for both clinicians and public health institutes.

Increased Mitochondrial Genetic Diversity in Persons Infected With Hepatitis C Virus.

BACKGROUND & AIMS: The host genetic environment contributes significantly to the outcomes of hepatitis C virus (HCV) infection and therapy response, but little is known about any effects of HCV infection on the host beyond any changes related to adaptive immune responses. HCV persistence is associated strongly with mitochondrial dysfunction, with liver mitochondrial DNA (mtDNA) genetic diversity linked to disease progression. METHODS: We evaluated the genetic diversity of 2 mtDNA genomic regions (hypervariable segments 1 and 2) obtained from sera of 116 persons using next-generation sequencing. RESULTS: Results were as follows: (1) the average diversity among cases with seronegative acute HCV infection was 4.2 times higher than among uninfected controls; (2) the diversity level among cases with chronic HCV infection was 96.1 times higher than among uninfected controls; and (3) the diversity was 23.1 times higher among chronic than acute cases. In 2 patients who were followed up during combined interferon and ribavirin therapy, mtDNA nucleotide diversity decreased dramatically after the completion of therapy in both patients: by 100% in patient A after 54 days and by 70.51% in patient B after 76 days. CONCLUSIONS: HCV infection strongly affects mtDNA genetic diversity. A rapid decrease in mtDNA genetic diversity observed after therapy-induced HCV clearance suggests that the effect is reversible, emphasizing dynamic genetic relationships between HCV and mitochondria. The level of mtDNA nucleotide diversity can be used to discriminate recent from past infections, which should facilitate the detection of recent transmission events and thus help identify modes of transmission.

Accurate Genetic Detection of Hepatitis C Virus Transmissions in Outbreak Settings.

Hepatitis C is a major public health problem in the United States and worldwide. Outbreaks of hepatitis C virus (HCV) infections are associated with unsafe injection practices, drug diversion, and other exposures to blood and are difficult to detect and investigate. Here, we developed and validated a simple approach for molecular detection of HCV transmissions in outbreak settings. We obtained sequences from the HCV hypervariable region 1 (HVR1), using end-point limiting-dilution (EPLD) technique, from 127 cases involved in 32 epidemiologically defined HCV outbreaks and 193 individuals with unrelated HCV strains. We compared several types of genetic distances and calculated a threshold, using minimal Hamming distances, that identifies transmission clusters in all tested outbreaks with 100% accuracy. The approach was also validated on sequences obtained using next-generation sequencing from HCV strains recovered from 239 individuals, and findings showed the same accuracy as that for EPLD. On average, the nucleotide diversity of the intrahost population was 6.2 times greater in the source case than in any incident case, allowing the correct detection of transmission direction in 8 outbreaks for which source cases were known. A simple and accurate distance-based approach developed here for detecting HCV transmissions streamlines molecular investigation of outbreaks, thus improving the public health capacity for rapid and effective control of hepatitis C.

Evaluating Protection Against Infectious Bronchitis Virus by Clinical Signs, Ciliostasis, Challenge Virus Detection, and Histopathology.

In this study, we examined the association among clinical signs, ciliostasis, virus detection, and histopathology for evaluating protection of vaccinated chickens against homologous and heterologous infectious bronchitis virus (IBV) challenge. At 5 days following challenge with IBV, we found a good correlation among clinical signs, ciliostasis in the trachea, challenge virus detection, and microscopic lesions in the trachea, with all four criteria being negative in fully protected birds and positive in fully susceptible birds. In partially protected birds we observed clinical signs and detected challenge virus; however, the ciliated epithelium was intact. In a second experiment, we challenged fully protected, partially protected, and fully susceptible birds with IBV, and then at 5 days postchallenge we gave the birds an opportunistic bacterium intranasally. Twenty Bordetella avium colonies were recovered from one of five fully protected birds, and only five colonies were isolated from two of five partially protected birds without ciliostasis, whereas in birds with ciliostasis, numerous colonies were isolated. Obviously, decreasing IBV infection and replication in the upper respiratory tract will decrease transmission and mutations, leading to variant viruses, and herein we demonstrate that protection of the cilia will decrease secondary bacterial infections, which have been shown to lead to condemnations and increased mortality. Thus, it appears that examining both criteria would be important when evaluating IBV vaccine efficacy.

Hatchery Spray Cabinet Administration Does Not Damage Avian Coronavirus Infectious Bronchitis Virus Vaccine Based on Analysis by Electron Microscopy and Virus Titration.

studies in our laboratory showed that the Arkansas-Delmarva Poultry Industry (Ark-DPI) vaccine given to 1-day-old chickens by hatchery spray cabinet replicated poorly and failed to adequately protect broilers against homologous virus challenge, whereas the same vaccine given by eye-drop did replicate and the birds were protected following homologous virus challenge. To determine if mechanical damage following spray application plays a role in failure of the Ark-DPI vaccine, we examined the morphology of three Ark-DPI vaccines from different manufacturers using an electron microscope and included a Massachusetts (Mass) vaccine as control. One of the Ark-DPI vaccines (vaccine A) and the Mass vaccine had significantly (P < 0.005) fewer spikes than the other two Ark-DPI vaccines. We also found that the Ark-DPI and Mass vaccines had significantly (P < 0.005) fewer spike proteins per virus particle when compared to their respective challenge viruses. This observation is interesting and may provide some insight into the mechanism behind infectious bronchitis virus attenuation. No obvious differences were observed in virus morphology and no consistent trend in the number of spikes per virion was found in before- and after-spray samples. We also determined the vaccine titer before and after spray in embryonated eggs and found that both Ark-DPI and Mass vaccines had a similar drop in titer, 0.40 logi and 0.310 logi, respec10ively. Based on these data, it appears that mechanical damage to the Ark-DPI vaccine is not occurring when delivered by a hatchery spray cabinet, suggesting that some other factor is contributing to the failure of that vaccine when given by that method.

Detection of infectious bronchitis virus with the use of real-time quantitative reverse transcriptase-PCR and correlation with virus detection in embryonated eggs.

Real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) assays have been used to detect the presence of challenge virus when the efficacy of infectious bronchitis virus (IBV) vaccine against field viruses is being experimentally evaluated. However, federal guidelines for licensing IBV vaccines indicate that challenge-virus detection following vaccination is to be conducted in embryonated eggs. In this study, we examined qRT-PCR data with the use of universal and type-specific primers and probe sets for IBV detection and compared those data with challenge-virus detection in embryonated eggs to determine if the two methods of evaluating vaccine efficacy are comparable. In addition, we tested the qRT-PCR assays on thermocyclers from two different manufacturers. We found the universal IBV primers and probe set to be comparable to challenge-virus detection in embryonated eggs. However, for some IBV types (Mass41 and Conn on the SmartCycler II and Ark, Mass41, Conn, and GA98 on the ABI 7500) the qRT-PCR assay was more sensitive than virus detection in embryonated eggs. This may simply be due to the universal IBV qRT-PCR assay being more sensitive than virus detection in eggs or to the assay detecting nucleic acid from nonviable virus. This finding is important and needs to be considered when evaluating challenge-virus detection for vaccination and challenge studies, because qRT-PCR could potentially identify positive birds that would otherwise be negative by virus detection in embryonated eggs; thus it could lead to a more stringent measure of vaccine efficacy. We also found that the IBV type-specific primers and probe sets designed in this study were in general less sensitive than the universal IBV primers and probe set. Only the Ark-DPI-spedcific assay on the SmartCycler II and the Ark-DPI-, Mass41-, and DE072/GA98- (for detection of GA98 virus only) specific assays on the ABI 7500 were comparable in sensitivity to virus detection in eggs. We found that a number of variables, including the virus type examined, primers and probe efficiency and stability, and assay conditions, including thermocycler platform, can affect the data obtained from qRT-PCR assays. These results indicate that qRT-PCR assays can be used to detect IBV challenge virus, but each assay, including the assay conditions and thermocycler, should be individually evaluated if those data are expected to be comparable to virus detection in embryonated eggs.

Simultaneous detection of five major serotypes of Avian coronavirus by a multiplex microsphere-based assay.

Avian coronavirus (commonly known as Infectious bronchitis virus [IBV]) is of major economic importance to commercial chicken producers worldwide. Due to the existence of multiple serotypes and variants of the virus that do not cross-protect, it is important to diagnose circulating serotypes and choose the right vaccine type for successful protection. In an effort to improve conventional diagnostic tests, a microsphere-based assay was developed and evaluated for simultaneous detection of the most common IBV vaccine serotypes in the United States: Arkansas (Ark), Connecticut (Conn), Massachusetts (Mass), Delaware (DE072), and Georgia 98 (GA98). The analytical specificity and sensitivity, and diagnostic specificity and sensitivity, were evaluated. The microsphere-based assay was highly specific to designated serotypes and generated reproducible data. Comparing the microsphere-based assay to nucleotide sequencing, the 2 methods agreed more than 93% (kappa value > .77). In addition, the microsphere-based assay could detect coinfections in clinical samples. The results demonstrate the utility of the microsphere-based assay as a rapid and accurate diagnostic tool with the potential for high throughput diagnosis.

Evaluation of infectious bronchitis virus Arkansas-type vaccine failure in commercial broilers.

Infectious bronchitis virus (IBV) causes an upper respiratory tract disease in chickens and is highly contagious. Many different types of the virus exist, but only a few types are used as attenuated live vaccines in the commercial poultry industry. Of the vaccine types used, the Arkansas (Ark)-type virus is most frequently reisolated from vaccinated broilers. Previous research has suggested that incomplete clearance of Ark-type vaccine virus plays a role in the inadequate protection observed when vaccinated broilers are challenged with pathogenic Ark virus. In this study, we examine routes of vaccine administration using multiple IBV types including Ark in an effort to understand why Ark vaccines do not provide good protection and persist in commercial broilers. We found that interference between different types of IBV vaccines was not occurring when combined and administered using a commercial hatchery spray cabinet. Also, Ark vaccine virus was not efficacious in 1-day-old broilers when sprayed using a hatchery spray cabinet, but it gave good protection when administrated by eyedrop inoculation. We also found that the amount of Ark vaccine virus was low or undetectable in choanal swabs out to 35 days postvaccination when vaccine was administered by eyedrop or drinking water. Alternatively, a subpopulation of the Ark vaccine isolated from a vaccinated bird, Ark-RI-EP1, showed a peak titer at 7-10 days of age when given by the same routes, suggesting that the Ark-RI-EP1 was more fit with regard to infection, replication in the birds, or both. Moreover, we found that detection of IBV vaccine virus early after administration, regardless of strain or route, correlated with protection against homologous challenge and may thus be a good indicator of vaccine efficacy in the field because humoral antibody titers are typically low or undetectable after vaccination. These experiments provided key findings that can be used to direct efforts for improving the efficacy of IBV Ark-type vaccines given in the hatchery. They also elucidated factors contributing to the persistence of Ark vaccine in the field.

Effects of DDA, CpG-ODN, and plasmid-encoded chicken IFN-gamma on protective immunity by a DNA vaccine against IBDV in chickens.

This study examined the adjuvant effects of dimethyl dioctadecyl ammonium bromide (DDA), CpG oligodeoxynucleotides (CpG-ODN), and chicken interferon-gamma (ChIFN-gamma) on a DNA vaccine (pcDNA-VP243) against the infectious bursal disease virus (IBDV). A plasmid encoding chicken IFN-ã was constructed. Twice at 2-week intervals, two-week-old chickens were injected intramuscularly and intraperitoneally with either a DNA vaccine alone or a DNA vaccine together with the respective adjuvants. On week 2 after the second immunization, the chickens were orally challenged with the highly virulent IBDV. The groups that received the DNA vaccines plus either DDA or CpG-ODN showed significantly lower survival rates than the group that received the DNA vaccine alone. However, the survival rates for the DNA vaccine alone and for the DNA vaccine plus ChIFN-gamma were similar. The chickens had no detectable antibodies to the IBDV before the challenge but all the surviving chickens in all groups except for the normal control group showed the induction of antibodies to the IBDV at day 10 after the challenge. As judged by the lymphocyte proliferation assays using the a WST-8 solution performed on the peripheral blood and splenic lymphocytes, the stimulation indices (SI) of the peripheral blood lymphocytes in all groups except for the normal control group were similar immediately before the challenge. At 10 days post-challenge, the SI for DNA vaccine plus either CpG-ODN or ChIFN-gamma was similar to that of the DNA vaccine control group. For splenic lymphocytes, the SI in the DNA vaccine plus CpG-ODN and DNA vaccine plus ChIFN-gamma groups were higher than for the DNA vaccine control. These results suggest that DDA actually compromises the protection against the IBDV by DNA vaccine, and CpG-ODN and IFN-gamma had no significant effect.